TRANSFORMATION AND AVAILABILITY OF COPPER

TO WHEAT (Triticum aestivum L) AS INFLUENCED BY

PHOSPHORUS FERTILIZATION

Thesis

Submitted to the Punjab Agricultural University

in partial fulfillment of the requirements

for the degree of

MASTER OF SCIENCE

in

SOIL SCIENCE (Minor Subject: Chemistry)

By

Harpreet Kaur

(L-2012-A-113-M)

Department of Soil Science College of Agriculture

©PUNJAB AGRICULTURAL UNIVERSITY

LUDHIANA-141004

2014

CERTIFICATE I

This is to certify that the thesis entitled, “Transformation and availability of

copper to wheat (Triticum aestivum L) as influenced by phosphorus fertilization”

submitted for the degree of Master of Science, in the subject of Soil Science (Minor subject:

Chemistry) of the Punjab Agricultural University, Ludhiana, is a bonafide research work

carried out by Harpreet Kaur (L-2012-A-113-M) under my supervision and that no part of

this thesis/dissertation has been submitted for any other degree.

The assistance and help received during the course of investigation have been fully

acknowledged.

_________________________

Major Advisor

(Dr. J. S. Manchanda)

Senior Soil Chemist

Department of Soil Science

Punjab Agricultural University

Ludhiana - 141004

CERTIFICATE II

This is to certify that the thesis entitled, “Transformation and availability of

copper to wheat (Triticum aestivum L) as influenced by phosphorus fertilization”

submitted by Harpreet Kaur (L-2012-A-113-M) to the Punjab Agricultural University,

Ludhiana, in partial fulfillment of the requirements for the degree of Master of Science, in

the subject of Soil Science (Minor subject: Chemistry) has been approved by the Student’s

Advisory Committee along with Head of the Department after an oral examination on the

same, in collaboration with an external examiner.

____________________ ____________________

(Dr. J. S. Manchanda) (Dr. I. M. Chhibba)

Major Advisor External Examiner

Senior Soil Chemist (Retd.)

112-C, Rajguru Nagar

Ludhiana - 141012

____________________

(Dr. H. S. Thind)

Head of the Department

_______________________

(Dr. Gursharan Singh)

Dean, Post Graduate Studies

4

ACKNOWLEDGEMENTS

First of all, I offer my humble thanks with folded hands and bowed head to the

Almighty for his grace, kindness and blessings due to which I have been able to accomplish

this important task of my life.

With immense pleasure, I wish to express my sincere thanks and profound sense

of gratitude to the major advisor Dr. J S Manchanda, Sr. Soil Chemist, Department of Soil

Science, Punjab Agricultural University, Ludhiana who provided me the research insights,

inspiring and affectionate guidance, constructive criticism, calm endurance, parental

attitude and keen interest throughout the course of investigation and preparation of this

manuscript. Working under his expertise has been a great learning experience.

I express my deep sense of gratitude to the respected members of my Advisory

Committee, Dr. D S Bhatti, Senior Soil Chemist, Department of Soil Science, Dr. M S

Hadda, Professor of Soil Conservation (Nominee Dean PGS Studies), Department of Soil

Science and Dr. (Mrs) Anita Garg, Chemist, Department of Chemistry, for their

encouragement, continuous support, immaculate guidance and help during the tenure of my

study.

I duly acknowledge the research facilities provided by the Dr. H S Thind, Professor

and Head, Department of Soil Science, Punjab Agricultural University, Ludhiana.

Invaluable help rendered by laboratory and field staff of the Department of Soil

Science is fully acknowledged.

I found no words in expressing my profound gratitude to my parents and family

members. Their endless love, affection, sacrifices and constant inspiration has enabled me

to reach the footsteps of my long cherished aspiration.

Harpreet Kaur

5

Title of the Thesis : Transformation and availability of copper to wheat

(Triticum aestivum L) as influenced by phosphorus

fertilization

Name of the Student : Harpreet Kaur

and Admission No. (L-2012-A-113-M)

Major Subject : Soil Science

Minor Subject : Chemistry

Name and Designation : Dr. J. S. Manchanda

of Major Advisor Sr. Soil Chemist

Degree to be Awarded : M.Sc

Year of award of Degree : 2014

Total Pages in Thesis : 131+Vita

Name of University : Punjab Agricultural University, Ludhiana – 141004,

Punjab, India

ABSTRACT

A pot experiment was conducted to study the effect of P and Cu application on transformation

and availability of Cu to wheat. The soils used were i) calcareous loamy sand (ls) Typic Ustipssament

(pH 8.1, EC 0.375 dS m-1

, OC 0.15%, available P 8.5, CaCO3 0.13%, DTPA-Cu 0.18 and DTPA-Fe

3.20 mg kg-1

soil) and ii) non calcareous sandy loam (sl) Typic Haplustept (pH 6.5, EC 0.295 dS m-1

,

OC 0.38%, available P 12.65, DTPA-Cu 1.05 and DTPA-Fe 51.6 mg kg-1

soil). Six levels of P (0, 25,

50, 100, 200 and 400 mg P kg-1

soil) as monocalcium phosphate monohydrate and four levels of Cu (0,

5, 10 and 20 mg Cu kg-1

soil) as Cu-EDTA were applied in all possible combinations to eight kg of

each soil per pot with three replications. Wheat (cv HD 2967) was grown and soil, root, grain and straw

samples were collected at maturity. Soil (pH, available P, DTPA-Cu, DTPA-Fe and chemical pools of

Cu) and plant samples (Total P, Cu and Fe) were processed and analysed. Soil pH decreased with Cu

application while EC and Olsen P increased with P application in both soils. DTPA-Cu and Fe

decreased up to a level of about 75 mg P kg-1

soil and thereafter followed an upward trend. In ls soil P

application decreased exchangeable and specifically adsorbed Cu but increased carbonate bound-Cu.

However in sl soil, applied P decreased the content of Cu in exchangeable, amorphous and crystalline oxides

but increased in organically bound Cu. Growth and yield of wheat improved significantly with graded

levels of applied P. However, when any level of P was combined with 20 mg Cu kg-1

soil, severe Fe

chlorosis of leaves, a drastic reduction in growth, chlorophyll content and increase in activity of super

oxide dismutase was observed in calcareous ls only. The results indicated that it was Cu and not P that

induced Fe deficiency in wheat grown in alkaline calcareous soil and the Cu requirement of the crop

seemed to be much lower in the light textured soils. Root dry matter, grain and straw yield decreased

with increasing levels of applied Cu in ls but in sl a maximum increase of 62.5, 74.3 and 63.7 per cent

in root, grain and straw yield was observed with a combination of P400Cu5 over P0Cu0. Path coefficient

analysis revealed the importance of DTPA-Cu, exchangeable, specifically adsorbed and amorphous

oxides bound Cu in effecting the grain yield of wheat in ls soil and that of oxides and organically

bound Cu in sl. Phosphorus uptake by each plant part decreased with increase in DTPA-Cu while Fe

uptake increased with increase in Olsen P but decreased with increase in DTPA-Cu. Accumulation of

Cu in root decreased the Fe absorption by roots. The root Cu: Fe concentration ratio at which severe

Fe chlorosis of leaves was observed varied between 0.304 to 0.429. About 6.56, 5.39 and 5.37 mg

DTPA-Cu kg-1

in soil and 436, 11.04 and 19.33µg Cu g-1

in root, grain and straw produced 50 per cent

reduction from the maximum yield of root, grain and straw, respectively which may be considered as

the upper critical values for wheat.

Key words: Wheat, Cu, P, Fe Chlorosis, Chemical Pools of Cu

________________________ _____________________

Signature of Major Advisor Signature of the Student

6

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7

CONTENTS

CHAPTER TOPIC PAGE(S)

I INTRODUCTION 1-2

II REVIEW OF LITERATURE 3-12

III MATERIALS AND METHODS 13-18

IV RESULTS AND DISCUSSION 19-115

V SUMMARY 116-119

REFERENCES 120-131

VITA

CHAPTER I

INTRODUCTION

Phosphorus has been considered to be the kingpin in agriculture because it plays a

pivotal role in increasing crop production and improving the quality of crops. India is the

world’s third largest producer of phosphatic fertilizers and second largest consumer after

China (Prasad 2012). If large amounts of P are supplied in soils, its luxury uptake may disturb

the ratios of P to other nutrients including micronutrients (Tagliavini et al 1991). The present

day modern agriculture aims at achieving maximum production per unit area per unit time per

unit cost. This has created an imbalance of nutrients in soils and crops not only with respect to

macronutrients but also the micronutrients. Rice-wheat is the major cropping system of

Punjab and normally P is applied to wheat. However, in some areas farmers are applying P to

both the crops. But only one- third of applied P is taken up by the current crop that may lead

to the buildup of P in plough layer of soils over a period of time. At present only 36% of the

area in Punjab is low in P supplying capacity (Sharma et al 2011). Build up of P in soils can

affect pH, CEC and surface charge of soils which in turn may alter the equilibria of

micronutrients among their various chemical pools. Besides leading to various types of

nutrient interactions in soils and plants. Nutrient interactions in crop plants are probably one

of the most important factors affecting the yields of annual crops. An interaction takes place

when the supply of one of the nutrients affects the absorption, distribution or function of the

other and it may be negative, positive or neutral (Fageria 2001). Thus, depending upon the

nutrient supply, the interactions between nutrients can either induce deficiencies or toxicities

and can modify the growth response.

Intensive cultivation of high yielding cultivars on coarse textured soils and heavy

applications of macronutrients such as N, P, and K fertilizers to the crops leads to the

occurrence of micronutrient deficiencies. Recent surveys have demonstrated that about 3-4

per cent soils in Punjab have become deficient in available Cu (Anonymous 2008). Copper is

one of the essential nutrient elements required by plants and plays an important role in the

nutrition of crops. In modern agriculture, an increased cropping intensity, excessive use of

high analysis chemically pure nitrogenous and phosphatic fertilizers, cultivation of high

yielding varieties and extension of rice cultivation to coarse textured soils had led to the

depletion of micronutrients to a level which has become a serious problem for obtaining the

optimum yield of crops (Dar 2004, Barik and Chandel 2001). In plants, Cu plays an essential

role in chlorophyll formation, photosynthesis, respiration, energy transfer, cell wall

lignification, protein metabolism, maintaining the sterility of male flower besides helping in

seed production (Fageria 2009, Rattan and Goswami 2002). Under Cu deficient conditions the

activities of super oxide dismutase and ascorbate oxidase enzymes are reduced (Salama

2

2001). Apart from meeting the nutritional requirement of crops, Cu is also known to control

some diseases of plants. Foliar application of Cu in rice reduced the incidence of brown leaf

spot of rice (Manchanda et al 2008). Wheat plants with insufficient Cu have been found to be

susceptible to stem melanosis (Piening et al 1989).

Copper deficiency is more frequently observed in crops grown in organic soils. But its

deficiency has also been observed in coarse textured, alkaline calcareous mineral soils and

usually occurs in irregular patches within the fields (Kruger et al 1985). Its deficiency in crops

is expected when total Cu in soils is 5-25 mg kg-1 soil and the profile distribution of Cu shows a

decreasing trend up to 60 cm soil depth. Copper deficiency in Indian soils vary from <1% in

Andhra Pradesh, Assam, Orissa and West Bengal to as high as 31% in Kerala (Singh 2009).

The content of Cu in plants varies from 5-30 mg kg-1

dry matter. Among the various

crops wheat, oats, onion, tomato, alfalfa, lettuce, spinach and sugar beet are highly responsive

to Cu fertilization. Wheat has a special need for Cu and has been classified as a relatively

sensitive crop to its deficiency which has been reported in countries like Zimbabwe (Tanner

and Cooper 1984, Tanner et al 1981), Canada (Malhi et al 2004) and Australia (Best et al

1985, Robson et al 1984). In India Cu deficiency in wheat has been reported as early as 1969

(Grewal et al 1969). In Punjab wheat is grown on an area of about 3.49 m ha with an annual

production of 15.5 m tonnes. Only a few studies indicated as to how P affects the availability

of Cu in Punjab soils by redistributing it into its various chemical forms. Thus, keeping in

view the above points the present study has been designed with the following objectives:

i) To study interactive effects of P and Cu on availability of Cu to wheat.

ii) To study the effect of applied P on redistribution of Cu into various chemical pools.

CHAPTER II

REVIEW OF LITERATURE

The relevant research studies carried out by various workers on the different aspects

of the present investigation entitled “Transformation and availability of Cu to wheat (Triticum

aestivum L) as influenced by P fertilization in Punjab soils” has been reviewed under the

following headings:

2.1 Availablity of micronutrient cations in soils as influenced by P fertilization

2.2 Importance of Cu in plant nutrition

2.3 Yield and nutrition of crops as influenced by Cu fertilization

2.4 Transformation of micronutrient cations in soils

2.1 Availability of micronutrient cations in soils as influenced by phosphorus fertilization

In an incubation study, Mandal and Haldar (1980) observed that application of P @ 5

and 10 mg kg-1

soil in lowland rice soils of West Bengal significantly decreased the contents

of DTPA-extractable Zn , Cu, Fe and Mn. The rate of decrease gradually reduced with

increasing period of incubation from 10 to 70 days. They further observed that the depressive

effect of P on extractable Zn was more pronounced on native rather than applied Zn.

Manchanda et al (2012) observed a steep fall in DTPA- Zn when P: Mn ratio in soil was 6.0

and reported that a P: Mn ratio of 3.28 in soil produced 80% of the maximum dry matter yield

of shoot of wheat. Awan and Abbasi (2000) observed that P application increased P

concentration and decreased the Cu concentration in the maize plants in sandy loam soi,

indicating that an interaction between P and Cu in soil occurs which effect the production of

maize fodder significantly.

Singh et al (2005) observed that availability of Mn in a near neutral non-calcareous

soil was increased when the build up of available P in soil exceeded 60 mg P kg-1

soil.

Chatterjee et al (1983) reported a significant decrease in DTPA extractable Mn with P

application @ 100 mg P kg-1

in an acidic soil (pH 6.5) incubated for 30 days. However, in an

alkaline soil (pH = 7.6), Mamo and Parson (1987) reported a significant increase in DTPA

extractable Mn with P application @ 400 mg P kg-1

soil incubated for 28 days. Misra and

Mishra (1968) reported that P application to alkali soils decreased the retention of Mn by soil

colloids and increased the availability of Mn. However, in near neutral soil (pH = 7.3) having

11.5 ppm DTPA-Mn, the availability of Mn in soil was not influenced by P application even

up to 120 kg P2O5 ha-1

(Rao et al 1984).

Nutrient interactions in crop plants are probably one of the most important factors

affecting the yields of annual crops. An interaction takes place when the supply of one of the

nutrients affects the absorption, distribution or function of the other and it may be negative,

positive or neutral (Fageria 2001). Thus, depending upon the nutrient supply, the interactions

4

between nutrients can either induce deficiencies or toxicities and can modify the growth

response. India is the world’s third largest producer of phosphatic fertilizers and second

largest consumer after China (Prasad 2012). If large amounts of P are supplied in soils, its

luxury uptake may disturb the ratios of P to other nutrients including micronutrients

(Tagliavini et al 1991). Thus, an understanding of the interaction of P with other nutrients can

be of help to maintain a balanced supply of nutrients to get the optimum crop yields.

Numerous field and green house studies suggest that high levels of available P in soils

may depress Zn concentrations in plant tissues and thus increase the Zn requirements by

various crops (Haldar and Mandal 1981, Dwivedi et al 1975, Manchanda et al 2012, Deo and

Khandelwal, 2009). But some reports are there that within the normal rates of their

application P x Zn interaction may be synergistic in calcareous soil (Yadav et al 1991).

Furthermore, P-Zn relationship in soils and plants may also be governed by factors like

organic matter (Nayak and Gupta 2000), plant parts (Reddy and Yadav 1994), plant species

and growth stages (Islam et al 2005). Mandal and Haldar (1980) reported that the decrease in

uptake of Zn, Cu and Fe by rice because of P application was due to decrease in the

availability of these nutrients in soils. In a greenhouse study, Haldar and Mandal (1981)

observed that application of P in alluvial rice soils significantly increased the dry matter yield

of root, shoot and grain, but it decreased the concentration of Zn, Cu, Fe and Mn in both roots

and shoots. This decrease in the concentration of elements in shoots was not due to the

dilution effect or to the reduced rate of translocation from roots to tops. They observed that

this decrease was more due to the changes in their availability in soil due to P application.

Dwivedi et al (1975) observed P induced Zn deficiency in corn when P was applied

@ 120 mg kg-1

soil in a Zn deficient soil (DTPA- Zn 0.2 mg kg-1

soil). They further observed

that high levels of P rendered the applied Zn unavailable to plants by immobilizing almost

40% or more of the total Zn absorbed in roots and 20 per cent or more at the nodes of the

stem. Rahman et al (2011) observed that a combined application of P and Zn @ 50 and 10 kg

ha-1

, respectively produced a significant positive effect on the yield (5.97 t ha-1

) and yield

components of rice in soils of Bangladesh. In a nutrient culture experiment, Soltangheisi et al

(2014) observed that Zn deficiency in sweet corn can enhance P uptake and translocation to

such an extent that P may accumulate to toxic levels in its leaves. They further reported that

the lowest dry matter yield of corn was produced when P was applied @ 80 mg L-1

in the

absence of applied Zn. Application of 100 kg P ha-1

significantly decreased the Cu

concentration in maize plants (Awan and Abassi 2000). Similarly, an antagonistic effect of P

on Cu concentration of wheat (Shukla and Singh 1979, Javadi et al 1991) has been observed.

Singh et al (2005) observed a significant increase in dry matter yield of root and

shoot of wheat with graded levels of applied P in a Typic Haplustept but P and Mn

concentration in both root and shoot were inversely related with each other. An antagonistic

5

effect of P on Fe concentration in moong bean has been observed (Yadav et al 2002). A

synergistic (Kuo and Mikkelsen 1981) as well antagonistic (Soni et al 2000) effect of P

fertilization on Mn availability in soils to plants has been reported. Manchanda et al (2012)

observed that total Zn uptake by wheat root plus shoot decreased steeply when Olsen P at

harvest was >15 mg kg-1

soil.

Application of Zn (4-32 kg ha-1

) and P (30-60 kg P2O5ha-1

) at all levels increased the

grain and straw yield of wheat under field conditions. Optimum dose of Zn varied with the

level of P. Zinc content of grain and straw decreased as the level of P increased (Singh and

Singh 1979). Mishra and Abidi (2010) reported that P and Zn application had a synergistic

impact on the 1000-seed weight and protein content of the wheat varieties.

Das et al (2005) conducted a green house experiment on stevia plants and reported

that separate applications of P and Zn either as soil or foliar spray increased their content

while same levels of P and Zn when applied in combination decreased their content both in

soil and plants suggesting a mutual antagonistic effect between Zn and P.

Shittu and Ogunwale (2013) reported that application of more than 2 kg Zn ha-1

significantly decreased P uptake by soybean plants and maximum grain yield was produced

with a combined application of 30 kg P ha-1

and 2 kg Zn ha-1

. Soni et al (2000) reported that

when P and Mn was applied @ 0 to 60 mg P kg-1

soil and 0 to 50 mg Mn kg-1

soil respectively

in a reclaimed sodic soil (pH = 8.7) then increasing levels of P decreased the concentration of

Mn in wheat at each level of applied Mn. Recovery of added Mn was lower at higher levels of

P application.

Adriano et al (1971) reported that in corn seedlings with high P levels, shoot growth

was increased only by high Zn. The most marked interaction at high P levels was between Fe

and Zn which mutually antagonized translocation more than absorption. Dev et al (1983)

reported that application of 7.5 mg P kg-1

soil P enhanced the Mn concentration of chickpea

while its concentration decreased when P was applied @ 15 and 30 mg kg-1

soil.

In a field experiment Zhang et al (2012) observed that application of P up to 400 kg P

ha-1

did not influence the DTPA- extractable Zn, Cu, Fe and Mn in a alkaline calcareous loam

soil under wheat crop. However, they observed a significant reduction in the concentration of

Zn in wheat grain by 17 to 56 per cent with P application. But the accumulation of shoot Fe,

Cu and Mn was increased with applied P. Li et al (2007) also did not observed any significant

effect of P application on DTPA extractable micronutrient cations in soil under long term

inorganic and organic fertilizer application.

Goel and Duhan (2014) observed a significant decrease in DTPA extractable Zn, Cu,

Fe and Mn with application of varying levels of P (0-37.5 kg P ha-1

) to a Typic

Torripsamment of Hisar. In an alkaline calcareous soil of Pakistan, Ali et al (2014) observed a

6

decrease in the availability of Zn and Cu but an increase in the availability of Fe and Mn with

that application of P (0-150 kg P ha-1

).

2.2 Role of copper in plant growth and development

Copper plays an important role in growth and development of plants. Bortels (1927),

a German scientist, provided the first credible evidence that Cu was an essential element in

the nutrition of lower plants. Later on its essentiality for higher plants was established by

Lipman and Mackinney (1931) and Sommer (1931). Copper is a constituent of several

proteins and enzymes. These Cu containing enzymes play a key role in respiration,

photosynthesis, lignifications, phenol metabolism, protein synthesis and regulation of auxins

(Maksymiec 1997). In nearly all cases, Cu enzymes are involved in oxidation- reduction

reactions. Copper is a constituent of plastocyanin, hence it plays a role in photo-

phosphorylation and electron transport chain (Barr and Crane 1976; Solberg et al 1999).

Droppa et al (1984) reported a reduced rate of chlorophyll synthesis in the Cu deficient sugar

beet leaves. Copper ions readily form a complex with amino acids and these are more stable

than those involving any other metal. It is also involved in lignin biosynthesis, which not only

provides strength to cell walls but also prevents wilting (Taiz and Zeiger, 2010).

Copper is immobile in plants and the movement of Cu appears to be strongly

dependent on Cu status of plants. In wheat plants well supplied with Cu, movement readily

occurs from leaves to the grain, but in deficient plants, Cu is relatively immobile. Copper

deficiency in wheat and other cereals produces characteristic symptoms of yellowing and

curling of young leaves, pig tailing of leaf tips, limpness or wilting, delay in heading aborted

heads and spikelets, head and stem bending (Graham and Nambiar 1981; Piening and

McPherson 1985). Copper deficiency symptoms were described by Evans et al (1994) and

Solberg et al (1999). Symptoms occurred as irregular patches, sometimes with a browning

discoloration called “stem melanosis” (Piening et al 1987) or ergot infection. Graham (1975)

reported that Cu deficiency leads to pollen sterility.

2.3 Yield and nutrition of crops as influenced by copper fertilization

Wheat crop has a special need for copper and has been classified as a relatively

sensitive crop to its deficiency (Brown and Clark 1977). Copper deficiency in wheat and other

cereals produces characteristic symptoms of yellowing and curling of young leaves, pig

tailing of leaf tips, limpness or wilting delay in heading, aborted heads and spikelet’s, head

and stem bending (Graham and Nambiar, 1981; Robson and Reuter1981; Piening and

McPherson, 1985). Sinclair and Withers (1995) while studying the role of Cu in cereal

nutrition and the effects of Cu deficiency on yield and quality of cereals concluded that yield

reductions due to shortage of Cu can occur even without visual deficiency symptoms and

application of Cu gives a more increase in grain yield and improves the protein levels in

grains. Solberg et al (1999) reported that application of Cu fertilizer on Cu deficient soils

7

improved the crop quality and yield of cereals. It was observed that with Cu application, the

yield of barley increased from 41.5 to 61.6 q ha-1

and in park wheat the yield increased from

10.0 to 24.1 q ha-1

. Robson et al (1984) conducted a pot experiment to diagnose the Cu

deficiency in wheat and found that Cu deficiency can resist growth of the plants and

whenever the growth of the crop was reduced the average concentration of Cu in the youngest

fully emerged leaf was less than 1.3 mg kg-1

. Malakauti and Ziaeian (2002) reported that the

application of Cu, Zn, Fe and Mn caused a significant increase in grain and straw yield, 1000-

grain weight and the number of seeds per spikelet in wheat. With the application of these

micronutrients their concentration and total uptake by grain and concentration in flag leaves

increased significantly.

Brennan (1990) carried out a field experiment in Western Australia where wheat

was grown on Cu deficient yellow loamy sand soil and Cu sulphate was applied as foliar

sprays at 0, 21, 63, 125, 250 and 375 g Cu ha-1

. Copper deficiency symptoms were observed

in no Cu plots receiving no Cu whereas the highest Cu rates eliminated all deficiency

symptoms without causing foliar damage, although some damage occurred when a second

spray at high Cu rate (375 g Cu ha-1

) was applied. It was concluded that the quantity of Cu

required to achieve maximum grain yield was 250 g Cu ha-1

. A greenhouse experiment was

conducted by Kumar et al (1990) with a Cu deficient sandy soil using wheat crop. Copper

was applied at different concentrations such as 0, 5, 10 and 20 mg kg-1

soil. An increase in

dry matter yields of shoots and roots of wheat was observed up to a level of 5 mg kg-1

soil,

but decreased at higher level of Cu. Maximum yield of the crop was observed at 5 µg Cu g-1

soil level. Schmidt et al (2000) studied the effect of foliar application of Cu in the form of

Cu tetramine hydroxide on the yield of winter wheat. In spite of the fact that soil analysis

did not indicate an expressed Cu deficiency, the application of Cu in the form of Cu

tetramine hydroxide increased the yield of winter wheat. The Cu levels of 0.3 to 1.0 kg ha-1

increased the yield significantly in most of the cases and there was a strong significant

relationship between yield and dose of Cu application. Dwivedi and Shankar (1977)

reported that wheat responded to an application of 5 kg Cu sulphate ha-1

when soil had 0.5

mg Cu kg-1

soil as extracted by 1 N neutral ammonium acetate in soils of Kanpur. Brar and

Sekhon (1978) reported that application of Cu @ 2.5 mg kg-1

soil increased the dry matter

yield of wheat significantly but dry weight was significantly reduced with application of 10

mg Cu kg-1

soil.

Barnes and Cox (1973) observed that application of Cu materials on Cu deficient

organic soils increased the grain yield of wheat and soybean crop. Turvey (1984) reported a

significant increase in growth of the wheat crop with Cu treatments on Cu deficient soils of

Victoria, Australia. Galrao (1988) conducted a field experiment in an organic paddy soil to

evaluate the effect of Cu on wheat yield. The treatments consisted of five levels of Cu (0, 2, 4,

8

8, 16 kg Cu ha-1

), which were broadcasted as Cu sulphate. The treatments with no Cu

produced the lowest yield. Cu application at 2 kg Cu ha-1

increased the grain yield of wheat

significantly. Sakal et al (1981) reported that Cu application @ 5 kg Cu sulphate ha-1

significantly increased the Cu concentration in wheat grain and straw without any significant

increase in grain yield in a soil having 1.8 mg DTPA- Cu kg-1

soil.

Slight deficiency of Cu result in10-20%, moderate deficiency results in 20-50% and

severe deficiency results in 50-100 reduction in yield. Malhi et al (1989) conducted a field

experiment to study the effects of Cu deficiency on Park Wheat (Triticum aestivum cv. Park),

barley and oats. In 1985, six cultivars of wheat were sown in the field with treatments of

control (no Cu) and soil application of Cu chelate @ 4 kg Cu ha-1

. It was observed that stem

melanosis occurred under the control treatment while the Cu treated wheat was less subjected

to the stem melanosis disease. In the following year, wheat, barley and oats cultivars were

raised in similar conditions. It was observed that the wheat cultivars were more susceptible to

stem melanosis disease when grown on Cu-deficient soils and reported that when Cu sulphate

was applied at the rate of 10-20 kg Cu ha-1

, the incidence of the disease was reduced resulting

in improved grain yield.

Gupta and Macleod (1970) conducted a greenhouse experiment to determine the

optimum levels of Cu in wheat, barley and oats crops. The treatments consisted of four

levels of Cu (0, 0.5, 1 and 2 ppm Cu) added as CuSO4.5H2O. It was observed that the

application of 0.5 ppm Cu to the soil, under conditions increased the grain yield of wheat by

38 per cent, barley by 180 per cent and that of oats by 500 per cent over control while under

control condition (0 ppm Cu) the emergence of barley heads was delayed by 10 to 14 days.

Copper added at 1.0 ppm significantly increased barley kernel yields. A similar tendency

was noted with the 2 ppm Cu treatment, but the increase was short of statistical significance

at the 5% level.

Kumar et al (1990) conducted a pot experiment using a nitrogen and Cu deficient

sandy soil to study the effect of application of nitrogen fertilizers along with Cu fertilizers.

The sources of nitrogen used were Ca(NO3)2, NH4Cl and NH4NO3 applied in amounts

necessary to establish 120 ppm of soil nitrogen and using a control (0 ppm N). Copper was

applied, as Cu chloride, to give soil Cu levels of 0, 5, 10 and 20 ppm. Up to a level of 5

ppm Cu, the dry matter yield of shoot and root increased, but decreased at higher levels of

Cu. Increasing Cu levels significantly decreased the available nitrogen in soil after harvest

and also the concentration of N in the plants. At the same time the concentration of Cu in

shoot and root and available Cu in the soil was increased. Nitrogen and Cu were found to

have a mutually antagonistic effect on each other's concentration in the plants.

In a field experiment, Karimian et al (2001) reported that build up of P in plough

layer of soils may interfere with nutrition of wheat thereby increasing the need for

9

supplemental micronutrients. Zhu et al (2002) reported that higher P availability in soil

significantly reduced the uptake of Cu by eight cultivars of barley. Sharma and Singh (2012)

observed significant increase in the grain and straw yield of wheat cultivars with Cu

application of 5 kg ha-1

but a significant decline in yield was observed when Cu was applied

@ 10 kg ha-1

in an alkaline soil of Uttar Pradesh. They further observed that P uptake by

wheat was improved when Cu was applied @ 2.5 kg ha-1

. They further observed an adverse

effect of Cu in P utilization by wheat crop. Singh and Parkash (2009) reported that application

of Cu up to 2.5 kg ha-1

significantly increased the grain and straw yield of wheat in alluvial

soil but application of Cu reduced the available P in soil. Manivasagaperumal et al (2011)

observed that application of 50 mg Cu kg-1

soil resulted in a significant increase in the overall

growth dry matter yield and nutrient content of green gram whereas higher concentration up

to 250 mg Cu kg-1

soil decreased the growth and nutrient content of green grams.

Elevated levels of Cu in soils may prove to be toxic for plant growth. Copper in excess

interferes with plant’s capacity to absorb and /or translocate other nutrients, inhibits root hair

growth, root elongation and adversely affects the permeability of root cell membrane (Fageria

2014, Kopittke and Menizes 2006, Sheldon and Menizes 2005). The effects of Cu toxicity are

expected to be much higher in P deficient soils as compared to P sufficient soils. Northfield et al

(2011) observed that the alleviation of P deficiency in soil though improved the overall growth

of cowpea plants but it did not influence the apparent toxicity of Cu and they concluded that

reduced growth at higher level of Cu were associated with reduction in shoot tissue Fe

concentration. Michaud et al (2007) also reported that risks of Cu phytotoxicity to wheat

(Triticum durum) in situ might be greater in calcareous soils due to interaction with Fe nutrition.

In a pot experiment with rice, Brown et al (1954) observed that rice yield decreased with

increasing levels of P (0-720 mg kg-1

soil) and Cu (0-180 mg kg-1 soil) and caused the plants to

develop chlorosis. However, the crop growth was good at all levels of applied P in the absence

of applied Cu. In another study, Brown et al (1955) observed that increasing additions of P

varying from 0-800 kg ha-1

did not affect the absorption or utilization of Fe to the extent that can

lead to the development of chlorosis in wheat, rice and soybean unless Cu was also present in

the growth media. Copper and P were effective in producing Fe chlorosis if applied together

than if either element was applied separately. Similarly, in a field study Sahu et al (1988)

reported that excess of P and Cu in leaves of garden peas interfere with metabolic translocation

of Fe and render it inactive for chlorophyll synthesis. Kumar et al (2009) observed a significant

decrease in Fe concentration in leaves, grain and straw of wheat with increasing levels (0-2.5

mg kg-1

soil) of Cu application.

2.4 Transformation of micronutrient cations in soils

Generally, < 10 per cent of the total micronutrients associated with the solid phases

are in soluble and exchangeable form that may become available for plant uptake (Lake et al

10

1984). However, their redistribution among different pools due to changes in soil properties

brought about by natural and/or anthropogenic activities make the so called capacity factor

important for micronutrients. Some of the soil properties that affect the transformation and

availability of Zn, Cu, Fe and Mn in soils is reviewed.

Shuman (1985) reported that Mn was primarily in the organic and Mn oxide fractions

whereas Cu and Zn were found mainly in the crystalline Fe oxide, silt and clay fractions in the

fine textured soils. In sandy soils Mn was predominantly in sand and Cu was in Fe oxide

fraction, whereas Fe was relatively higher in organic matter and Zn was higher in

exchangeable and organic fractions in sandy soils compared with the fine textured soils.

Jessica et al (1993) observed higher content of Zn in all fractions in fine textured soils as

compared to coarse textured soils.

Soil pH is one of the most important factors that affect the availability of

micronutrients to crops by redistributing them in to the various solid phases. The content of

water soluble and exchangeable Zn decreased by 60 per cent and complexed Zn by 80 per

cent as the pH of soil samples collected from a contiguous belt of salt affected area in

Varanasi district (UP) increased from 7.6 to 10.3 (Srivastava and Srivastava 1992). There

was two times reduction in water soluble Mn, 6.4 fold reduction in exchangeable Mn and 2.6

fold increase in easily reducible Mn with increase in soil pH from 7.2 to 10.3, EC from 1.46

to 20.82 dS m-1

and ESP from 8.3 to 70 (Srivastava and Srivastava 1994). They observed a

significant and positive coefficient of correlation between pH and amorphous and crystalline

Fe and Al oxides bound Zn.

Sims (1986) reported that soil pH markedly altered the distribution of Zn and Mn in

soil fractions but had a little effect on Cu. The majority of soil Cu was in the organic fraction

but considerable percentages were found in amorphous Fe oxide fraction. Plant uptake of Mn

and Zn was primarily related to exchangeable fraction.

Increasing rates of application of lime decreased exchangeable Zn and increased

organic fraction of Zn and Mn. This increase in Zn and Mn in the organic fraction, as pH is

increased showed that pH does not influence metals in some fractions in the same manner as

it does plant availability. Iron decreased in exchangeable and organic fractions as pH

increased due to lime application (Shuman 1986).

Shuman (1988a) studied the effect of addition of wheat straw as OM on the

transformation of Zn, Cu, Fe and Mn under green house conditions. He observed that Fe and

Mn moved from less soluble forms to more plant available forms as the rates of OM

applications increased. However, Cu was not affected significantly, but Zn bound to Mn

oxides fraction increased at the expense of Zn in other fractions. He concluded that

accumulated OM near the soil surface can increase the plant availability of Fe and Mn and

possibly decrease that of Zn, by causing a redistribution of elements between soil fractions.

11

Singh et al (1988) studied the distribution and forms of Zn, Cu Fe and Mn in

calcareous soils in India. They observed that 62, 52, 53 and 82 per cent of the total soil Cu,

Fe, Mn and Zn, respectively, was present in residual fraction, and 17, 41, 11 and 7 per cent of

total soil Cu, Fe, Mn and Zn respectively, was associated with crystalline Fe oxide fraction.

The amorphous Fe oxide fraction averaged to 12, 6, 9 and 5 per cent of total soil Cu, Fe, Mn

and Zn, respectively. The amounts of metals in OM fraction were generally <1 per cent.

Carbonate bound Cu, Fe, Mn and Zn accounted for 6, < 0.1, 4 and 2 per cent of total present

in soils. Exchangeable Cu and Zn averaged to 2 per cent whereas exchangeable Fe and Mn

were in non significant fractions averaging to <0.1 percent of the total.

Shuman (1988b) reported the effect of added P on transformation of micronutrients

under green house conditions. Increasing rates of P application from 0 to 60 mg P kg-1

soil as

KH2PO4 moved Mn from less soluble fractions (Crystalline Fe oxides and residual) to the

intermediately soluble fractions (Mn oxides and Amorphous Fe oxides) and had a little effect

on plant available fractions (water soluble and exchangeable). Copper moved to the

exchangeable fraction from residual fraction in fine textured soils, whereas Zn moved from

Mn oxide and crystalline Fe oxide fractions to the exchangeable fraction for all soils studied.

Singh et al (2005) reported that application of P (0-400 mg kg-1

soil) moved Mn from less

plant available forms (crystalline oxides and Mn oxides bound) to more plant available forms

(exchangeable and specifically adsorbed). However, Manchanda et al (2012) observed that

application of P transformed Zn in soil from exchangeable and Fe oxides bound pools to Mn

oxides bound, organic matter bound and specifically adsorbed fractions. Brar et al (1986)

observed that a marked release of Zn from acid soluble fraction was the reason for lack of

response of maize and wheat to Zn application and significant positive correlations between

acid soluble Zn and Zn uptake by wheat and maize were reported. Many other workers

(Mandal et al 1988, Murthy 1982, Dhane and Shukla 1995) have reported significant and

positive correlations among different fractions of metals, which suggested that various forms

of the elements are in dynamic equilibrium with each other and changes in the concentration

of one pool will be accompanied by changes in other fractions in order to maintain the

equilibrium depending upon the prevailing soil environment.

Jalali and Moharami (2010) reported that application of mono potassium phosphate

(MPP) and diammonium phosphate (DAP) in a calcareous soil altered the distribution of Zn and

Cu in soil fractions. Diammonium phosphate significantly increased the concentration of Zn and

Cu in exchangeable fraction. They further observed that in a leaching experiment with MPP and

DAP the concentration of Cu and Zn in carbonate fraction increased significantly over control.

Behera et al (2009) reported that residual Cu fraction was the dominant pool of total

Cu in all the four layers (0-15, 15-30, 30-45 and 45-60 cm) of soil under maize-wheat

cropping system. DTPA-Cu was predominantly influenced by the sorbed Cu, though the

12

contribution of other fractions was also dominant. Pietrzak and McPhail (2004) reported a

decline in organically bound Cu with increasing depth and lower contents of organic matter in

vine yard soils of Victoria, Australia. Zhu and Alva (1993) reported that organically bound

form was the dominant form of Cu in some sandy soils, with varying pH, trace metal

concentration, and organic matter in Florida under citrus production.

Saha and Mandal (2000) reported that under submerged soil conditions, Cu was

mobilized from water soluble plus exchangeable, organically complexed, and crystalline Fe

oxides bound fractions to amorphous Fe oxides and residual fractions, the rate of mobilization

being maximum from crystalline Fe oxides bound fraction to residual fraction during initial

15-days period. Organic matter application retarded Cu transformation from organically

complexed Cu and into residual fraction and increased its content in amorphous Fe oxide

fractions. It also decreased water soluble plus exchangeable Cu markedly. Cu transformation

was not significantly influenced by P application.

Guan et al (2011) studied the effect of application of livestock manure spiked with

varying levels of Cu (0-800 mg kg-1

soil) and observed that Cu was predominantly distributed

in organically bound fraction. Acid soluble Cu played a significant role in controlling the

mobility and bioavailability of Cu. In a laboratory study with a Typic Udic Ferrisols of

China, Tu et al (2001) observed that application of 80 mg P kg-1

soil decreased the

exchangeable Cu but increased the content of specifically adsorbed Cu as well as Fe and Mn

oxides bound Cu.

CHAPTER III

MATERIAL AND METHODS

To achieve the objectives of this investigation following studies were undertaken

3.1 Pot culture studies

3.2 Laboratory studies

3.1 Pot culture studies

To study the interactive effects of P and Cu on availability of Cu to wheat

3.1.1 Collection of Soil Samples

Two bulk surface (0-15 cm) soil samples were used for the conduct of the pot

experiment. One of the soil samples was collected from Krishi Vigyan Kendra, Bahowal,

District Hoshiarpur and the other from the experimental area (A-8) of Department of Soil

Science, Punjab Agricultural University, Ludhiana. The soil samples were classified as Typic

Ustipssaments and Typic Haplustept, respectively. The important physico-chemical

characteristics of soils used in the study is given in Table 3.1.

3.1.2 Pot culture experiment was conducted with the following treatments

Phosphorus levels : 0, 25, 50, 100, 200 and 400 mg P kg-1

soil

Copper levels : 0, 5, 10, and 20 mg Cu kg-1

soil

Replications : 3

Soils : 2

Total number of pots : 6 x 4 x 3 x 2 = 144

Experimental design : Factorial CRD

The bulk soil samples were air dried under shade and ground by using a wooden

pestle and mortar to pass through 2 mm sieve. A pot culture experiment was conducted in a

screen house by growing wheat as a test crop. Each polyethylene lined plastic pot of 10 kg

capacity was filled with 8 kg of soil. Copper was applied @ 0, 5, 10 and 20 mg Cu kg-1

soil

through ethylene-di-amine-tetra acetic acid Cu disodium salt (Merck, 15.5% Cu) and P was

applied @ 0, 25, 50, 100, 200 and 400 mg P kg-1

soil through AR grade mono-calcium

phosphate in powder form in 24 possible combinations. The treatments were replicated thrice

and in all there were 144 pots. Treatments were randomized in a Factorial Complete

Randomized Design. Basal application of 120 mg N kg-1

soil was made through AR grade

Urea. Ten seeds of wheat (cv HD 2967) were sown on 10th Nov, 2013 in each pot and thinned

to five plants per pot after germination. Besides a pre sowing irrigation, pots were regularly

watered with water as and when required.

14

Table 3.1: Physico-chemical characteristics of experimental soils

Soil characteristic Soil I

(KVK Bahowal)

Soil II

(PAU Farm)

pH (1:2) 8.1 6.5

EC (1:2) (dS m-1

) 0.375 0.295

Particle size distribution

Sand (%) 81.5 72.5

Silt (%) 3.00 13.8

Clay (%) 15.5 13.7

Textural class Loamy sand Sandy loam

Calcium carbonate (%) 0.13 Nil

Organic carbon (%) 0.15 0.38

Available P (kg ha-1

) 8.51 12.65

Available K2O (kg ha-1) 200 220

Micronutrients Zn Cu Fe Mn Zn Cu Fe Mn

DTPA-extractable

(mg kg-1

soil)

1.12 0.18 3.20 2.65 2.24 1.05 51.6 4.15

Chemical pools of Cu (mg kg-1

soil)

EXCH SAD CARB MnOX AMPOX CRYOX OM Total

Loamy sand

(KVK Bahowal)

0.075 0.090 0.059 0.080 0.495 1.39 1.65 6.5

Sandy loam (PAU

Farm) 0.089 0.052 Nil 0.052 1.85 2.10 0.67 9.5

EXCH = Exchangeable; SAD = Specifically adsorbed; MnOX= Manganese oxides bound;

AMPOX = Amorphous Fe and aluminum oxides bound; CRYOX = Crystalline Fe and

aluminum oxides bound; OM = Organically bound.

3.1.3 Harvesting

The crop was harvested at maturity. The data on grain and straw yield per pot was

recorded. After harvesting soil samples were collected from each pot and also roots were

extracted. The plant samples were washed thoroughly in succession with tap water, 0.01 N

HCl and deionized water followed by air drying and oven drying at 65o C to constant weight.

The dried root and straw samples were ground in a Willey mill having stainless steel blades to

pass through 40 mesh sieve. Ground samples were stored in paper bags for chemical analysis.

The soil samples collected after the harvest of crop were air dried, processed and stored in

polyethylene bags for chemical analysis.

15

Table 3.2: Sequential extraction procedure for micronutrients cations in soil samples

Fraction Reagent Soil: Extractant

ratio

Conditions Reference

WS+

Exchangeable

0.005M

Pb (NO3)2

(pH 6.8)

1: 4 Shaking for 15

minutes

Manchanda et al

(2006)

Specifically

adsorbed

0.05M

Pb(NO3)2

(pH 6.0)

1: 4 Shaking for 2

hrs.

Iwasaki et al

(1993)

Carbonate

bound

1.6 M HNO3 5 g soil + 1.6 M*

HNO3 mix, dilute

to 20 ml.

Centrifugation

for 15 minutes

Warden and

Reisenauer

(1991)

Manganese

oxides bound

0.1M

NH2OH.HCl in

0.01 M HNO3

(pH 2.2)

1: 4 Shaking for 30

minutes

Chao (1972)

Amorphous Fe

and Al oxides

bound

0.25 M

NH2OH.HCl +

0.25 M HCl

(pH 1.3)

1: 4 Shaking for 30

minutes at 50o C

Chao and Zhou

(1983)

Crystalline Fe

and Al oxides

bound

0.25 M

NH2OH.HCl +

0.25 M HCl +

0.1 M

Ascorbic acid

(pH 1.21)

1: 4 Heating with

boiling water

(100o C) in a

beaker paced on

hot plate for 30

minutes

Manchanda et al

(2006)

Organically

bound

1 % Na4P2O7 1 : 4 Shaking for 1 hr Raja and

Iyengar (1986)

Residual Total minus

sum of all

fractions

Total HNO3: HClO4

digestion

Hesse (1971)

*According to CaCO3 content, 1 ml of 1.6 M HNO3 = 80 mg CaCO3

3.2 Laboratory studies

To study the effect of applied P on redistribution of Cu into various chemical pools,

soil samples collected after the harvest of crop at maturity in the above pot experiment were

analysed for various chemical pools of Cu as per details given in Table 3.2.

16

3.2.1 Sequential fractionation of soils for various pools of micronutrients

3.2.1.1 Exchangeable fraction (EXCH)

5 g of each soil sample was weighed in 50 ml centrifuge tube and 20 ml 0.005M

Pb(NO3)2 (pH 6.8) was added, stoppered and shaken for 15 minutes. After shaking,

centrifugation was done at 10000 g and the extract was filtered.

3.2.1.2 Specifically Adsorbed (SAD)

The soil left in the centrifuge tube, after extraction of EXCH fraction, was stirred with

glass rod and 20 ml of 0.05M Pb(NO3)2 (pH 6.0) was added and shaken for 2 hours,

centrifuged and filtered.

3.2.1.3 Carbonate fraction (CARB)

3.2.1.4 Manganese oxide bound fraction (MnOX)

The soil left in the centrifuge tube after the extraction of SAD fraction was stirred

with glass rod and was shaken with 20ml 0.1M NH2OH.HCI in 0.01M HNO3 (pH 2.2) for 30

minutes, centrifuged and filtered.

3.2.1.5 Amorphous Fe & Al oxides bound (AMPOX)

The soil left in the centrifuge tube after extraction of MnOX fraction was stirred with

glass rod and was shaken with 20 ml 0.25M NH2OH.HCl + 0.25M HCl (pH 1.3) for 30

minutes at 50°C, centrifuged and filtered

3.2.1.6 Crystalline Fe & Al oxides bound (CRYOX)

The soil left in the centrifuge tube after extraction of AMPOX fraction was stirred

with glass rod and was shaken with 20ml 0.25M NH2OH.HCl + 0.25M HCl + 0.1M ascorbic

acid (pH 1.21) and heated for 30 minutes in boiling water (100o C) in a beaker placed on hot

plate, centrifuged and filtered.

3.2.1.7 Organically bound fraction (OM)

The soil left in the centrifuge tube after extraction of CRYOX fraction was stirred

with glass rod and was shaken with 20 ml 1% Na4P2O7 for 1 hour, centrifuged and filtered.

The extracts were digested in diacid, made to known volume and filtered before feeding to

atomic absorption spectrophotometer.

3.2.1.8 Residual mineral fraction (RES)

Residual mineral fraction was obtained by subtracting the sum total of all the above

fractions from the total fraction.

3.2.1.9 Total fraction

Total fraction was determined by digesting 0.5 g soil with diacid HClO4: HNO3 (1:3)

mixture as described by Hesse (1971). Zn, Cu, Fe and Mn in all the above extracts were

determined on atomic absorption spectrophotometer.

17

3.3 Analytical Methods

3.3.1 Soil analysis

3.3.1.1 Soil reaction

1:2 soil-water suspension was used to determine soil pH by using digital pH meter.

3.3.1.2 Electrical Conductivity

Electrical conductivity of 1:2 soil water supernatant solution was measured by digital

conductivity meter after keeping soil samples overnight for settling soil particles.

3.3.1.3 Available Mn, Cu, Zn, Fe

Available Mn, Cu, Zn, Fe in soils were determined by using 0.005M DTPA-CaCl2

(pH 7.3) following the procedure given by Lindsay and Norvell (1978).

3.3.1.4 Available P

Available P in soil was determined by the method of Olsen et al (1954).

3.3.1.5 Organic Carbon

Organic Carbon was determined by rapid titration method of Walkley and Black

(1934).

3.3.1.6 Calcium carbonate

Calcium carbonate was determined by the rapid titration method of Puri (1950).

3.3.1.7 Soil texture

Soil texture was determined by using International Pipette method as described by

Piper (1950).

3.4 Plant analysis

Half a gram of the ground root/straw and 1.0 g of wheat grain sample were digested

in a di-acid mixture of distilled HNO3: HClO4 (3:1). The digested plant material was

transferred to 25 ml volumetric flask and volume was made with deionized water, filtered

through Whatman No.1 filter paper.

3.4.1 Determination of P in plant extracts

Phosphorus in the plant samples extracts was determined by vanado–molybdo–

phosphoric yellow colour method in nitric acid system as outlined by Jackson (1967).

3.4.2 Determination of micronutrients in plant extracts

Micronutrients in plant extracts were determined on atomic absorption spectrophotometer

3.4.3 Enzyme analysis

3.4.3.1 Reagents

i) 6 mM pyrogallol

Fresh solution was prepared for essay

ii) 6 mM EDTA in deionized water

iii) 0.1M Tris HCl Buffer (pH 8.2)

18

iv) Extraction

The enzyme was extracted from 100 mg of fresh leaf samples with 0.1M potassium

phosphate buffer (pH 7.5) containing 1% PVP (Polyvenyl pyrolidone), 1M EDTA and 10 mM

β-mercaptoethanol using pre-chilled pestle and motar. The extracts were centrifuged at 10000

g at 4°C for 10 minutes and supernatant was used for enzyme assay.

v) Procedure

To the spectrophotometric cuvette, 1.5 ml of buffer (0.1M Tris HCl buffer, pH 8.2),

0.5 ml of 6 mM EDTA and 0.1 ml of 6 mM pyrogallol were added. Absorption (OD) was

recorded at 420 ηm at an interval of 30 seconds up to 3 minutes. The blank was used as

reaction mixture without enzyme extract (Marklund and Marklund 1974).

vi) Calculations

Final OD – Initial OD

OD change min-1

=

Minutes

Control OD/minute – Treatment OD/minute

% Inhibition = × 100 = X%

Control OD/minute

50% Inhibition = 1 unit

X% Inhibition = 1/50 × X = Y units

Y units × Volume after centrifuging (ml)

Enzyme activity (units) =

ml of Enzyme extract taken for estimation × Weight of tissue (g)

= units min-1

g-1

FW

OD of test solution × Concentration of standard solution × Volume after centrifuge

Protein =

OD of standard solution × Volume taken for estimation × Fresh weight

Enzyme activity

Specific activity of enzyme = = units min-1

mg-1

protein

Protein (mg g-1

fresh weight)

3.5 Statistical analysis

The data generated in these studies were treated statistically in accordance with

methods outlined by Panse and Sukhatme (1967) by using computerized programmes

available in the computer centre of College of Agricultural Engineering, Punjab Agricultural

University, Ludhiana.

CHAPTER IV

RESULTS AND DISCUSSION

The results of present investigation are presented and discussed under the following

headings:-

4.1 Effect of P and Cu application on soil pH, electrical conductivity, available P and

DTPA-extractable Cu, Fe, Mn and Zn in soils after harvest

4.2 Effect of P and Cu application on transformation of Cu in soils.

4.3 Effect of P and Cu application on root dry matter yield, grain and straw yield

4.4 Effect of P and Cu application on concentration of P, Cu and Fe in root, grain and

straw

4.5 Copper to Fe concentration ratios in root, grain and straw

4.6 Upper critical levels of Cu in soil, root, grain and straw

4.7 Effect of P and Cu application on P, Cu and Fe uptake by root, grain and straw

4.1 Effect of phosphorus and Cu application on soil pH, electrical conductivity, available

P and DTPA-extractable Cu, Fe, Mn and Zn in soils after harvest

4.1.1 Soil pH

The data on soil pH as influenced by P and Cu application after the harvest of wheat

crop is presented in Table 4.1. In loamy sand alkaline calcareous soil, the pH of soil increased

significantly from 7.98 in control to 8.05 with application of 100 mg P kg-1

soil but

thereafter it decreased significantly to 7.95 and 7.91 over control with application of 200 and

400 mg P kg-1

soil, respectively (Table 4.1). However, the differences in soil pH observed

with application of 200 and 400 mg P kg-1

soil were not significant. Soil pH and Olsen P were

negatively correlated (r= -0.397) with each other (Table 4.3). Ali et al (2014) also observed a

significant negative correlation (r= -0.83) of soil pH with applied P in an alkaline calcareous

soil. In loamy sand alkaline calcareous soil, the quadratic relationship of levels of applied P

with soil pH (R2=0.343) indicated that soil pH increased up to a point when soil had about160

mg P kg-1

soil and thereafter soil pH exhibited a downward trend (Fig. 4.1a).

In sandy loam, non-calcareous nearly acidic soil, the pH decreased significantly

over control with increasing levels of applied P except at the highest level of applied P.

Its pH decreased from 6.36 in control to 6.01 and 5.92 with application of 25 and 200 mg

P kg-1

soil, respectively but thereafter it increased significantly to 6.30 with application of

400 mg P kg-1

soil over the application of 200 mg P kg-1

soil, however it remained lower

than control (Fig.4.1b). Contrary to loamy sand soil, the quadratic relationship of levels of

applied P with soil pH (R2=0.733)observed in sandy loam soil indicated that soil pH

20

decreased up to a point when soil had about160 mg P kg-1

soil and thereafter soil pH

exhibited an upward trend (Fig. 4.1b). Shuman (1988b) also observed a significant

increase in pH of piedmont and coastal plain soils from 5.4 in control to 5.9 and from 5.2

to 6.2, respectively with application of 60 mg P kg-1

soil as potassium di-hydrogen

orthophosphate.

Table 4.1: Effect of phosphorus and Cu application on pH of soils after harvest

Copper

levels

(mg kg-1

soil)

Soil pH (1: 2)

Phosphorus levels (mg kg-1

soil)

0 25 50 100 200 400 Mean

Loamy sand

0 7.95 8.12 8.07 8.13 8.07 8.01 8.06

5 8.02 8.10 7.99 8.12 7.97 7.99 8.03

10 8.01 8.03 8.09 8.12 7.96 7.96 8.03

20 7.95 7.92 8.03 7.81 7.82 7.69 7.87

Mean 7.98 8.04 8.04 8.05 7.95 7.91

CD (5%) P levels 0.073 Cu levels 0.059 P x Cu NS

Sandy loam

0 6.47 6.14 6.07 6.01 5.99 6.04 6.12

5 6.39 6.04 5.90 5.94 5.96 6.45 6.11

10 6.29 5.99 5.86 5.90 5.89 6.40 6.05

20 6.27 5.90 5.87 5.88 5.84 6.30 6.01

Mean 6.36 6.01 5.92 5.93 5.92 6.30

CD (5%) P levels 0.091 Cu levels 0.075 P x Cu 0.182

In both the soils, pH decreased with increasing levels of Cu application as Cu-EDTA.

In loamy sand, the pH decreased significantly from 8.06 in control to 7.87 when Cu was

applied @ 20 mg kg-1

soil whereas in sandy loam soil, the pH decreased significantly from

6.12 in control to 6.05 and 6.01 with application of 10 and 20 mg Cu kg-1

soil, respectively

(Table 4.1).

A significant interaction effect of P and Cu application on soil pH in sandy loam

soil revealed that at each level of applied Cu, soil pH increased significantly over control

when P was applied @ 400 mg kg-1

soil. Also, the pH decreased significantly from 6.47 in

21

control to 6.27 when no P was applied which may be due to the acidic nature of Cu-EDTA

used as a source of Cu. A maximum decrease of 0.63 unit in pH of acidic sandy loam soil

over control was observed with a combined application of 200 mg P and 20 mg Cu kg-1

soil.

Mamo and Parsons (1987) also observed a significant decrease in pH of Ethiopian

bottomland soil from 6.8 to 5.2 with application of 400 mg P kg-1

soil as mono ammonium

phosphate.

When P fertilizer is applied to soil, the dissolution and precipitation processes

become active. These processes involve both the formation of and subsequent dissolving of

phosphate precipitates. The fertilizer granules on soil surface attract moisture to them

resulting in chemical reactions which convert the soluble P into phosphoric acid and a less

soluble form of P, di-calcium phosphate.

When mono-calcium phosphate is added in soil it will react with water to form di-

calcium phosphate and phosphoric acid.

Ca(H2PO4)2 + H2O = CaH2PO4 + H3PO4

The phosphoric acid progressively releases H+ ions to the soil as pH is raised from

about 3.0 to above 7.0

H3PO4 = H+ + H2PO4

- = 2H

+ + HPO4

2- = 3H

+ + PO4

3-

In general, the third hydrogen is lost only at pH values above neutrality. However, the

two hydrogen ions will readily be lost in acid pH range. Lindsay (1979) reported that when

mono-calcium phosphate monohydrate dissolves in soils it produces a solution of pH about

1.3 in an area immediately around a fertilizer band which gradually diffuses into the soil

surrounding the band.

The initial dissolution of phosphate and slower reactions with soil components can

also alter the soil pH. If the prevailing soil conditions are acidic and sufficient amounts of Fe

and aluminium and Mn are present, the P from the granules can be transformed into low

solubility phosphates of these metals. This process can lead to the production of hydroxyl ions

due to the following reactions

FeOOH + H2PO4 = FeOH2PO4 + OH-

Al2O3.H2O + H2O + 2H2PO4- = 2Al(OH)2H2PO4 + 2OH

-

The increase in soil pH as a result of P fertilization may be due the production of

OH- ions resulting from the reaction of Fe and Al oxides with H2PO4- ions (Shuman 1988).

On the other hand if the overall pH is neutral or alkaline and adequate levels of Ca are

present the di-calcium phosphate can further be converted into tri-calcium phosphate. This

latter scenario is more important in maintain the overall concentration of P in soil solution.

22

Maintaining an overall soil pH 6-6.5 lessens the likelihood of P precipitation and enhances

P solubility.

In the present study, an initial increase in soil pH of the alkaline calcareous loamy

sand soil may be due to the dominance of the formation of di-calcium phosphate while an

increase in pH of acidic sandy loam soil at the highest level of applied P may be due to the

production of OH- ions because of the reaction of phosphate ions with the Fe and Al oxides

and hydroxides (Shuman 1988b) because this soil had more amounts of Fe and Al oxides as

compared to the alkaline calcareous soil. The decrease in pH of alkaline soil at the higher

levels of P application and that of acidic soil at lower levels of applied P may be due to the

production of H+ ions due to the dissociation of phosphoric acid produced as a result of

reaction of mono-calcium phosphate with soil moisture. Du et al (2010) while studying the

effect of monocalcium phosphate and potassium chloride on soil pH changes observed that

both the fertilizers significantly decreased soil pH after 7 and 28 days of incubation. They

further observed that in acidic red soil by application of monocalcium phosphate the decrease

in soil pH was slowed down but was prompted in calcareous soil.

4.1.2 Electrical conductivity

In both the soils, electrical conductivity increased with increasing levels of applied P

as well as Cu. In loamy sand soil it increased significantly from 0.486 dS m-1

in control to

0.498, 0.527, 0.539, 0.550 and 0.551 dS m-1

with application of 25, 50, 100, 200 and 400 mg

P kg-1

soil, respectively (Table 4.2, Fig. 4.1c, d). In sandy loam soil, electrical conductivity

increased significantly from 0.504 dS m-1

in control to 0.548, 0.570, 0.590, 0.595 and 0.612

with application of 25, 50, 100, 200 and 400 mg P kg-1

soil respectively. However, the

differences in EC observed with application of 100, 200 and 400 mg P kg-1

soil were not

significant in both the soils. The quadratic relationship of levels of applied P with soil EC

(R2=0.921 for loamy sand and 0.854 for sandy loam) indicated that soil EC increased up to a

point when soil had about 300 mg P kg-1

soil and thereafter it exhibited a downward trend in

both the soils (Fig. 4.1c, d).

Similarly, electrical conductivity increased significantly from 0.441 dS m-1

in

control to 0.495, 0.548 and 0.617 dS m-1

when Cu was applied @ 0, 5, 10 and 20 mg Cu

kg-1

soil, respectively in loamy sand soil. Whereas in sandy loam soil electrical

conductivity increased significantly from 0.473 dS m-1

in control to 0.579, 0.600 and

0.627 dS m-1

with application of 5, 10 and 20 mg Cu kg-1

soil, respectively. The

interaction effect of applied P and Cu levels on electrical conductivity was not significant

for both the soils. The increase in EC of soils may be due to addition of soluble salts of P

and Cu. Soil pH and EC were found to be negatively correlated with each other in loamy

23

Table 4.2: Effect of phosphorus and Cu application on electrical conductivity of soils

after harvest

Copper

levels

(mg kg-1

soil)

Electrical conductivity (dS m-1

)

Phosphorus levels (mg kg-1

soil)

0 25 50 100 200 400 Mean

Loamy sand

0 0.411 0.422 0.436 0.456 0.450 0.470 0.441

5 0.430 0.443 0.520 0.525 0.526 0.527 0.495

10 0.499 0.517 0.540 0.556 0.603 0.574 0.548

20 0.603 0.609 0.614 0.618 0.623 0.634 0.617

Mean 0.486 0.498 0.527 0.539 0.550 0.551

CD (5%) P levels 0.021 Cu levels 0.017 P x Cu NS

Sandy loam

0 0.374 0.440 0.453 0.520 0.510 0.543 0.473

5 0.490 0.550 0.596 0.603 0.610 0.626 0.579

10 0.550 0.600 0.600 0.606 0.623 0.624 0.600

20 0.601 0.603 0.630 0.633 0.640 0.655 0.627

Mean 0.504 0.548 0.570 0.590 0.595 0.612

CD (5%) P levels 0.038 Cu levels 0.031 P x Cu NS

Table 4.3: Linear coefficients of correlation among soil pH, EC, Olsen P and with

DTPA-extractable micronutrient cations (n=24)

pH EC Olsen-P

Loamy sand

EC -0.632**

Olsen-P -0.397 0.272

DTPA-Cu -0.672**

0.906**

0.062

DTPA-Fe -0.433* 0.323 0.998

**

DTPA-Mn -0.433* 0.322 0.998

**

DTPA-Zn -0.476* 0.382 0.992

**

Sandy loam

EC -0.215

Olsen-P 0.318 0.400

DTPA-Cu -0.086 0.726**

-0.044

DTPA-Fe 0.311 0.454* 0.997

**

DTPA-Mn 0.312 0.453* 0.997

**

DTPA-Zn 0.301 0.518**

0.986**

*Significant at 5%

**Significant at 1%

24

Fig. 4.1: Soil pH, electrical conductivity and available P in soil at harvest

y = -2E-06x2 + 0.000x + 7.975R² = 0.343

7.88

7.92

7.96

8

8.04

8.08

0 100 200 300 400

Soil

pH

Phosphorus levels (mg kg-1 soil)

y = 1E-05x2 - 0.003x + 6.201R² = 0.733

5.7

5.8

5.9

6

6.1

6.2

6.3

6.4

0 100 200 300 400

Soil

pH

Phosphorus levels (mg kg-1 soil)

y = -9E-07x2 + 0.000x + 0.491R² = 0.921

0.48

0.49

0.5

0.51

0.52

0.53

0.54

0.55

0.56

0.57

0 100 200 300 400

Ele

ctri

cal c

on

du

ctiv

ity

(dS

m-1

)

Phosphorus levels (mg kg-1 soil)

y = -1E-06x2 + 0.000x + 0.524R² = 0.854

0.5

0.52

0.54

0.56

0.58

0.6

0.62

0.64

0 100 200 300 400

Ele

ctri

cal c

on

du

ctiv

ity

(dS

m-1

)

Phosphorus levels (mg kg-1 soil)

y = 0.315x + 3.798R² = 0.997

0

20

40

60

80

100

120

140

0 100 200 300 400

Ava

ilab

le P

(m

g kg

-1so

il)

Phosphorus levels (mg kg-1 soil)

y = 0.201x + 11.63R² = 0.993

0

10

20

30

40

50

60

70

80

90

100

0 100 200 300 400

Ava

ilab

le P

(m

g kg

-1so

il)

Phosphorus levels (mg kg-1 soil)

(d) (c)

(e) (f)

(a) (b)

25

sand (r= - 0.632*) and sandy loam soil (r= -0.215) (Table 4.3). Kumar and Babel (2011)

also observed a negative coefficient of correlation between pH and EC in soils of

Rajasthan, India.

4.1.3 Available phosphorus

Availability of P in soil at harvest increased linearly with increasing levels of

applied P in loamy sand (Fig 4.1e R2=0.997) as well as sandy loam soil (Fig. 4.1f

R2=0.993). In loamy sand soil, extractability of P increased from 8.32 mg kg

-1 soil in

control to 10.43, 17.40, 33.06, 67.37 and 130.6 mg P kg-1

soil when P was applied @ 25,

50, 100, 200 and 400 mg kg-1

soil, respectively (Table 4.4). Whereas, in sandy loam soil

with the same levels of applied P, the extractability of available P increased significantly

from 12.62 mg kg-1

soil in control to 17.15, 23.88, 30.29, 47.70 and 94.62 mg kg-1

soil,

respectively. It was further observed that more amount of applied P remained in solution

form in light textured loamy sand as compared to sandy loam soil especially when P was

applied @ 200 and 400 mg P kg-1

soil. The extractability of P increased by 109, 297, 709

and1469 per cent over control in loamy sand and by 89, 140, 278 and 649 per cent only in

sandy loam soil when P was applied @ 50, 100, 200 and 400 mg kg-1

soil, respectively.

This may be due to more amounts of finer particles present in sandy loam soil (Table

3.1) as compared to loamy sand soil, thus leading to more fixation and lesser

extractability of P in sandy loam soil. The texture of soil has a bearing on the availability

of P in soils. In case of heavy soils, the fertilizer P reacts with soil colloid micelles to

form less water soluble P compounds than in sandy soils indicating that P fixation tends

to be more pronounced in heavy soils than in coarse textured soils (Olsen and Watanabe

1963). An increase in fertilizer P availability with decrease in clay content has been

reported (Sharpley et al 1984). The clay content of soils retain more P in solid phases and

lower the activity of P in soil solution thereby resulting in a decrease in availability of P

(Lakshmi et al 1987) to plants. Brar et al (1991) reported a decrease in response of wheat

to applied P with fineness of soil texture. A negative coefficient of correlation of Olsen P

with pH (r = -0.397) and positive correlation with EC (r=0.272) in loamy sand soil, and

positive correlations with both pH (r=0.318) and EC (r=0.400) in sandy loam soil were

observed (Table 4.3). An increase in the extractability of available P with increasing

levels of applied P in a silt loam soil has also been reported by Ali et al (2014).

26

Table 4.4: Effect of phosphorus and copper application on available phosphorus in soils

after harvest

Copper

levels

(mg kg-1

soil)

Available phosphorus (mg kg-1

soil)

Phosphorus levels (mg kg-1

soil)

0 25 50 100 200 400 Mean

Loamy sand

0 8.44 10.34 17.04 32.26 66.19 129.6 43.99

5 8.25 10.48 17.74 33.87 66.63 130.4 44.58

10 8.39 10.47 17.56 32.76 67.67 131.1 44.66

20 8.20 10.41 17.26 33.36 69.00 131.2 44.91

Mean 8.32 10.43 17.40 33.06 67.37 130.6

CD (5%) P levels 0.96 Cu levels NS P x Cu NS

Sandy loam

0 12.60 15.95 23.76 30.18 47.86 93.88 37.37

5 12.58 16.75 23.13 30.69 47.61 95.02 37.63

10 12.52 16.93 23.35 30.11 46.35 94.08 37.22

20 12.76 18.98 25.27 30.175 48.98 95.49 38.61

Mean 12.62 17.15 23.88 30.29 47.70 94.62

CD (5%) P levels 1.38 Cu levels NS P x Cu NS

4.1.4 DTPA-Cu

Available Cu in both the soils decreased with increasing levels of applied P except

that in loamy sand calcareous soil DTPA-Cu increased significantly over control when 400

mg P kg-1

soil was applied. In loamy sand, DTPA-Cu decreased from 3.17 mg kg-1

soil in

control to 2.84 mg kg-1

soil thus resulting in a decrease of 10.4 per cent over control with

application of 100 mg P kg-1

soil. But it increased significantly to 3.50 mg kg-1

soil when 400

mg P kg-1

soil was applied (Table 4.5). The quadratic relationship of Olsen P with DTPA-Cu

(R2=0.939) indicated that the latter decreased up to a point when soil had about 50 mg P kg

-1

soil and thereafter it exhibited an upward trend (Fig. 4.2a). In sandy loam soil, DTPA-Cu

decreased significantly from 3.65 mg kg-1

soil in control to 3.37 and 3.31 mg kg-1

soil which

represented a decrease of 7.67 and 9.31 per cent over control when P was applied @ 200 and

400 mg kg-1

soil respectively. Just like soil pH, the quadratic relationship of Olsen P with

DTPA-Cu (R2=0.975) in sandy loam soil indicated that the latter decreased up to a point when

soil had about 75 mg P kg-1

soil and thereafter it became constant (Fig. 4.2b). The decrease in

DTPA-Cu may be due to an increase in soil pH (Fig. 4.3a, b). Significant decrease in DTPA-

27

extractable Cu with varying rates of P application (0-100 mg P kg-1

soil) was observed by

Mandal and Haldar (1980) in rice soils. Goel and Duhan (2014) observed a significant

decrease in DTPA extractable Zn, Cu, Fe and Mn with application of varying levels of P (0-

37.5 kg P ha-1

) to a Typic Torripsamment of Hisar. In an alkaline calcareous soil of Pakistan,

Ali et al (2014) observed a decrease in the availability of Cu with application of P (0-150 kg

P ha-1

). Bingham and Garber (1960) reported that regardless of the source of P used excess P

(1000 kg P ha-1

) resulted in acute Cu deficiency in sour orange seedlings. They further

observed that antagonism between P and Cu was reduced in going from acid to alkaline soils.

However, Shuman (1988b) observed an increase in content of DTPA-Cu with P application

@ 60 mg p kg-1

soil. A significant negative coefficient of correlation (r= -0.672** n=24) of

pH with DTPA-Cu was observed in loamy sand soil (Fig. 4.3a, R2=0.862). It was also

negatively correlated with pH of sandy loam soil (Fig. 4.3b, R2=0.936)). However, DTPA-Cu

was significantly positively correlated with EC (r= 0.906** for loamy sand and 0.726** for

sandy loam) and poorly correlated with Olsen P in both the soils.

Application of Cu significantly increased DTPA-Cu from 0.20 mg kg-1

soil in

control to 6.69 in loamy sand soil and from 0.885 mg kg-1

soil in control to 6.29 mg kg-1

soil in sandy loam soil with the highest level of applied Cu. An increase in the

extractability of DTPA-Cu with Cu application is due to a significant decrease in soil pH

(Table 4.1) because of the acidic nature of the Cu-EDTA used as a source of Cu. The

extractability of Cu increased by 875, 1670, and 3245 per cent that of control in loamy

sand and by 200, 375 and 610 per cent only in sandy loam soil when 5, 10 and 20 mg Cu

kg-1

soil was applied respectively, thereby indicating that in light textured soils more

amounts of applied Cu remain in plant available forms as compared to medium textured

soils. Shuman (1988b) also observed an increase in the extractability of DTPA-Cu with

its application to fine as well as coarse textured soils. DTPA-Cu was significantly

positively correlated with EXCH-Cu, SAD-Cu, CARB-Cu, MnOX-Cu, AMPOX-Cu,

CRYOX-Cu, OM-Cu and RES-Cu in both the soils indicating that DTPA is not only

extracting exchangeable, adsorbed and complexed forms of Cu but also other pools are

significantly contributing to it (Table 4.17). Guan et al (2011) also reported that DTPA

may overestimate the Cu bioavailability as the DTPA solution could extract soluble as

well as a part of the stable forms. The interaction effect of P and Cu levels on DTPA-Cu

was significant. It was observed that DTPA-Cu decreased significantly when 100 mg P

kg-1

soil was applied together with 20 mg Cu kg-1

soil as compared to when only 20 mg

Cu kg-1

soil was applied in both the soils.

28

Table 4.5: Effect of phosphorus and copper application on DTPA - copper in soil after

harvest

Copper

levels

(mg kg-1

soil)

DTPA – Cu (mg kg-1

soil)

Phosphorus levels (mg kg-1

soil)

0 25 50 100 200 400 Mean

Loamy sand

0 0.19 0.18 0.18 0.18 0.22 0.24 0.20

5 1.89 1.81 2.10 1.94 1.79 2.19 1.95

10 3.45 3.38 3.52 2.80 3.26 4.80 3.54

20 7.16 6.83 6.44 6.45 6.51 6.79 6.69

Mean 3.17 3.05 3.06 2.84 2.94 3.50

CD (5%) P levels 0.21 Cu levels 0.17 P x Cu 0.42

Sandy loam

0 0.99 0.90 0.85 0.88 0.85 0.83 0.88

5 2.59 2.78 2.80 2.95 2.41 2.42 2.66

10 4.50 4.55 4.32 4.67 3.35 3.89 4.21

20 6.53 6.26 6.43 5.55 6.88 6.11 6.29

Mean 3.65 3.62 3.60 3.51 3.37 3.31

CD (5%) P levels 0.25 Cu levels 0.20 P x Cu 0.51

4.1.5 DTPA-Fe

In loamy sand soil, DTPA-Fe decreased significantly from 3.25 mg kg-1

soil in

control to 3.05, 3.08, 3.00 and 3.04 mg kg-1

soil when P was applied @ 25, 50, 100 and 200

mg kg-1

soil which represented a decrease of 6.15, 5.23, 7.69, 6.46 and 1.84 per cent over

control, respectively (Table 4.6). But when P was applied @ 400 mg kg-1

soil, DTPA-Fe

increased significantly over the application of 200 mg P kg-1

soil however it remained lower

than control. The quadratic relationship of Olsen P with DTPA-Fe (R2=0.526) indicated that

the latter decreased up to a point when soil had about 60 mg P kg-1

soil and thereafter it

exhibited an upward trend (Fig. 4.2c).

In sandy loam soil, a significant decrease in DTPA-Fe was observed only when P was

applied @ 100, 200 and 400 mg kg-1

soil. It decreased from 58.6 mg kg-1

soil in control to

29

53.6, 52.0 and 50.2 mg kg-1

soil, thereby resulting in a decrease of 8.53, 11.26 and 14.33 per

cent over control with application of 100, 200 and 400 mg P kg-1

soil, respectively. A

quadratic response of DTPA-Fe to Olsen P (R2=0.948) in soil at harvest indicated that DTPA-

Fe decreased up to a point when soil had 75 mg P kg-1

soil and thereafter it became constant

(Fig. 4.2d). The differences in the DTPA- Fe observed with application of 100, 200 and 400

mg P kg-1

soil were not significant. The decrease in DTPA-Fe may be due to the formation of

Fe-phosphates (Olsen 1972). Motalebifard et al (2013) also observed a significant decrease in

extractable Fe in a clayey soil when P was applied @ 60 mg kg-1

soil as monocalcium

phosphate. Phosphorus is universally known to form insoluble Fe phosphates and it is

expected that P fertilizers change the micronutrients concentrations in soil solution and

influence their capacity factor (Rattan and Deb, 1981). Additionally, P fertilization may

encounter transformations of Fe soluble forms to insoluble forms (Mandal et al 2000). The

reduction in availability of Fe by P application has been reported by several workers (Singh

and Dahiya 1976, Ohki 1984, Kochain 1991). Contrary to these observations an increase in

DTPA-Fe with application of 30 and 60 mg P kg-1

soil in acidic soils has also been reported

(Shuman 1988b).

Application of Cu @ 10 and 20 mg kg-1

soil resulted in a significant increase in

DTPA-Fe in both the soils. In loamy sand soil it increased from 2.95 mg kg-1

soil in

control to 3.32 and 3.16 mg kg-1

soil when Cu was applied @ 10 and 20 mg kg-1

soil,

respectively. A significant decrease in DTPA- Fe (4.8%) in calcareous loamy sand soil

was observed when Cu was applied @ 20 mg kg-1

soil as compared to application of 10

mg Cu kg-1

soil. The decrease in the availability of Fe due to application of 20 mg Cu kg-1

soil in the calcareous soil led to the development of Fe chlorosis of leaves at each level of

applied P. Michaud et al (2007) also reported that risks of excess Cu supply to wheat

(Triticum durum) in situ might be greater in calcareous soils due to interaction with Fe

nutrition.

In sandy loam soil DTPA- Fe increased from 47.9 mg kg-1

soil in control to 52.3, 57.7

and 61.7 mg kg-1

soil, thereby representing an increase of 9.18, 20.4 and 28.8 per cent over

control, respectively with application of 5, 10 and 20 mg Cu kg-1

soil. The increase in DTPA-

Fe due to applied Cu as Cu-EDTA may be due to decrease in soil pH as a result of Cu

application (Table 4.1). Soil pH was significantly negatively correlated (r= -0.433* n=24)

with DTPA-Fe in loamy sand soil (Table 4.3). In sandy loam soil, also DTPA-Fe was

negatively correlated with soil pH (Fig. 4.3d, R2=0.950)). Kumar and Babel (2011) also

observed a significant negative correlation of soil pH with available Cu, Fe, Mn and Zn in

soils of Rajasthan. It was significantly positively correlated with Olsen P (r= 0.998** for

loamy sand and 0.997** for loamy sand soil) in both soils.

30

Table 4.6: Effect of phosphorus and copper application on DTPA- Fe in soils after

harvest

Copper

levels

(mg kg-1

soil)

DTPA – Fe (mg kg-1

soil)

Phosphorus levels (mg kg-1

soil)

0 25 50 100 200 400 Mean

Loamy sand

0 3.24 2.68 3.00 2.74 2.98 3.01 2.95

5 3.22 3.70 2.68 3.02 2.50 2.86 2.99

10 3.22 2.86 3.66 3.53 3.33 3.32 3.32

20 3.34 2.94 2.98 2.73 3.38 3.60 3.16

Mean 3.25 3.05 3.08 3.00 3.04 3.19

CD (5%) P levels 0.128 Cu levels 0.104 P x Cu 0.257

Sandy loam

0 50.2 49.8 45.0 47.2 47.6 47.5 47.9

5 52.2 60.7 53.2 50.5 44.4 53.0 52.3

10 67.2 62.0 61.2 53.8 53.6 48.7 57.7

20 65.0 64.3 64.2 63.0 62.3 51.4 61.7

Mean 58.6 59.2 55.9 53.6 52.0 50.2

CD (5%) P levels 4.06 Cu levels 3.31 P x Cu 8.12

4.1.6 DTPA- Mn

In loamy sand soil DTPA-Mn decreased (3.09%) significantly from 6.79 mg kg-1

soil

in control to 5.83 mg kg-1

soil when P was applied @ 100 mg kg-1

soil but it increased to 6.73

and 6.80 mg kg-1

soil when P application level was increased to 200 and 400 mg P kg-1

soil,

respectively (Table 4.7). The decrease in availability of Mn in soil may be attributed to the

formation of insoluble Mn phosphates (Larsen 1956). Mandal and Haldar (1980) reported that

application of P @ 50 and 100 mg P kg-1

soil decreased the DTPA-Mn in rice soils due to

precipitation of Mn as insoluble Mn phosphates. In sandy loam non- calcareous soil DTPA-

Mn increased with increasing levels of applied P. It increased from 4.26 mg kg-1

soil in

control to 4.36, 4.63, 5.07 and 4.69 mg kg-1

soil which represented an increase of 2.34, 9.62,

8.68, 19.01 and 10.09 per cent over control when P was applied @ 25, 50, 100, 200 and 400

mg kg-1

soil, respectively (Table 4.7). A quadratic response of Olsen P to DTPA-Mn (R2

=

0.942) was observed only for sandy loam soil which indicated that DTPA-Mn increased up to

a point when soil had 60 mg P kg-1

soil and thereafter it followed a decreasing trend (Fig.

4.2f).These findings corroborate with those of Larsen (1964) who reported an increase in Mn

availability in an acidic soil at the highest level of applied P (600 mg P kg-1

soil) due to

formation of soluble Mn phosphate complexes but a decrease in Mn availability at lower rates

31

(40 mg P kg-1

soil) of applied P due to formation of insoluble Mn phosphate complexes.

Singh et al (2005) also observed that the availability of Mn in soil may be expected to

increase when the level of Olsen P in soil exceeds 60 mg P kg-1

soil. Ali et al (2014) observed

an increase in the availability of Fe and Mn in alkaline calcareous soils of Pakistan with P

application.

In sandy loam soil, application of 10 and 20 mg Cu kg-1 soil resulted in a significant

increase of 6.17 and 10.7 per cent in DTPA-Mn over control, respectively. It increased from 4.37

mg kg-1 soil in control to 4.64 and 4.84 mg kg

-1 soil with application of 10 and 20 mg Cu kg

-1 soil,

respectively. The increase in DTPA-Mn due to copper application may be due to decrease in soil

pH (Table 4.1). Soil pH was significantly negatively correlated (r= -0.433* n=24) with DTPA-Mn

in loamy sand soil (Fig. 4.3e, R2=0.853). It was also negatively correlated with DTPA-Mn in

sandy loam soil (Fig. 4.3f, R2=0.799). Olsen P was significantly positively correlated with DTPA-

Mn (r=0.998** for loamy sand and 0.997** for sandy loam) in both soils.

Table 4.7: Effect of phosphorus and copper application on DTPA - Mn in soils after harvest

Copper

levels

(mg kg-1

soil)

DTPA Mn (mg kg-1

soil)

Phosphorus levels (mg kg-1

soil)

0 25 50 100 200 400 Mean

Loamy sand

0 6.69 6.64 6.16 5.50 6.82 6.96 6.46

5 6.73 6.86 6.05 6.13 6.60 6.86 6.54

10 6.66 6.72 6.80 5.40 6.35 6.45 6.40

20 7.10 6.08 6.97 6.31 7.17 6.92 6.76

Mean 6.79 6.58 6.50 5.83 6.73 6.80

CD (5%) P levels 0.45 Cu levels NS P x Cu NS

Sandy loam

0 3.95 3.94 4.21 4.43 4.94 4.77 4.37

5 4.14 4.32 4.91 4.24 5.17 4.83 4.60

10 4.30 4.37 4.48 5.21 5.00 4.51 4.64

20 4.67 4.82 5.08 4.62 5.20 4.66 4.84

Mean 4.26 4.36 4.67 4.63 5.07 4.69

CD (5%) P levels 0.32 Cu levels 0.26 P x Cu NS

4.1.7 DTPA- Zn

Application of P resulted in a significant decrease in DTPA-Zn in both the soils. In

loamy sand calcareous soil, it decreased significantly from 1.24 mg kg-1

soil in control to 1.05,

1.03, 1.02, 1.07 and 1.00 mg kg-1

soil when P was applied @ 25, 50, 100, 200 and 400 mg kg-1

soil thereby representing a decrease of 15.32, 16.93, 17.74, 13.7 and 19.35 per cent over

32

control, respectively (Table 4.8). However, the differences in DTPA-Zn observed with graded

levels of applied P were not significant. Similarly, DTPA-Zn decreased from 2.29 mg kg-1

soil

in control to 2.06, 2.09, 2.03 and 2.12 mg kg-1

soil representing a decrease of 10.0, 9.0, 11.3 and

7.42 per cent over control when P was applied @ 50, 100, 200 and 400 mg kg-1

soil,

respectively in sandy loam soil. But again the differences in DTPA- Zn with various levels of

applied P were not significant. A quadratic response of Olsen P to DTPA- Zn (R2

= 0.774) was

observed only for sandy loam soil which indicated that DTPA- Zn decreased up to a level when

soil had about 60 mg P kg-1

soil (Fig. 4.2h). Phosphorus bears a strong influence on Zn reactions

and plays a critical role in its solubility in soils (Alloway 2009). Excessive applications of P

fertilizers increase the intensity of Zn deficiency in soils especially in soils low in available Zn

(Norvell et al 1987, Shuman 1988b). However, Pasricha et al (1987) observed that applying P

to four soils (acidic, neutral, alkaline, and calcareous) did not decrease Zn intensity in the soil

solution. Motalebifard et al (2013) observed a significant decrease in extractable Zn in a clayey

soil when P was applied @ 60 mg kg-1

soil as monocalcium phosphate. Similarly, Manchanda et

al (2012) also observed a significant decrease of 29% in DTPA- Zn in a Typic Haplustept with

application of 400 mg P kg-1

soil.

Table 4.8: Effect of phosphorus and copper application on DTPA - Zn in soils after harvest

Copper

levels

(mg kg-1

DTPA Zn (mg kg-1

soil)

Phosphorus levels (mg kg-1

soil)

0 25 50 100 200 400 Mean

soil) Loamy sand

0 1.02 0.90 1.02 1.07 0.93 0.93 0.98

5 1.08 1.09 0.95 0.91 0.97 1.00 1.00

10 1.35 0.96 1.09 0.86 1.18 1.02 1.08

20 1.51 1.26 1.08 1.25 1.21 1.06 1.23

Mean 1.24 1.05 1.03 1.02 1.07 1.00

CD (5%) P levels 0.072 Cu levels 0.059 P x Cu 0.145

Sandy loam

0 2.11 2.12 2.00 2.02 1.94 1.83 2.00

5 2.16 2.21 1.87 1.93 1.87 1.96 2.00

10 2.40 2.57 2.19 2.15 2.01 2.30 2.27

20 2.51 2.29 2.20 2.25 2.31 2.40 2.33

Mean 2.29 2.30 2.06 2.09 2.03 2.12

CD (5%) P levels 0.124 Cu levels 0.101 P x Cu NS

33

Fig. 4.2: Effect of Olsen P in soil on DTPA-extractable micronutrient cations at harvest

y = 4E-05x2 - 0.005x + 3.184R² = 0.526

2.95

3

3.05

3.1

3.15

3.2

3.25

3.3

0 25 50 75 100 125 150

DTP

A-F

e (

mg

kg-1

soil)

Olsen P (mg kg-1 soil)

y = 0.002x2 - 0.360x + 63.36R² = 0.948

48

50

52

54

56

58

60

0 25 50 75 100DTP

A-F

e (

mg

kg-1

soil)

Olsen P (mg kg-1 soil)

y = 9E-05x2 - 0.010x + 6.643R² = 0.231

5.5

6

6.5

7

0 50 100 150

DTP

A-M

n (

mg

kg-1

soil)

Olsen P (mg kg-1 soil)

y = -0.000x2 + 0.044x + 3.740R² = 0.942

4

4.25

4.5

4.75

5

5.25

0 50 100

DTP

A-M

n (

mg

kg-1

soil)

Olsen P (mg kg-1 soil)

y = 1E-05x2 - 0.002x + 1.133R² = 0.265

0.95

1.05

1.15

1.25

0 25 50 75 100 125 150

DTP

A-Z

n(m

g kg

-1so

il)

Olsen P (mg kg-1 soil)

y = 0.000x2 - 0.016x + 2.475R² = 0.774

1.75

2

2.25

2.5

0 25 50 75 100

DTP

A--

Zn (

mg

kg-1

soil)

Olsen P (mg kg-1 soil)

(a) (b)

(c) (d)

(e) (f)

(g) (h)

34

Fig. 4.3: Effect of soil pH on DTPA-extractable micronutrient cations in soil at harvest

y = -42.88x + 263.9R² = 0.936

0

1

2

3

4

5

6

7

6 6.05 6.1 6.15

DTP

A-C

u (

mg

kg-1

soil)

Soil pH

Sandy loam (b)

y = -0.603x + 7.934R² = 0.093

2.90

3.00

3.10

3.20

3.30

3.40

7.8 7.9 8 8.1

DTP

A-F

e (

mg

kg-1

soil)

Soil pH

Loamy sand(c)

y = -113.6x + 745.2R² = 0.950

40

45

50

55

60

65

6 6.05 6.1 6.15

DTP

A F

e (

mg

kg-1

soil)

Soil pH

Sandy loam (d)

y = -1.688x + 20.04R² = 0.853

6.30

6.40

6.50

6.60

6.70

6.80

7.8 7.9 8 8.1

DTP

A-M

n (

mg

kg-1

soil)

Soil pH

Loamy sand (e)

y = -3.322x + 24.78R² = 0.799

4.3

4.4

4.5

4.6

4.7

4.8

4.9

6 6.05 6.1 6.15

DTP

A-M

n (

mg

kg-1

soil)

Soil pH

Sandy loam (f)

y = -1.255x + 11.11R² = 0.908

0.80

0.90

1.00

1.10

1.20

1.30

7.8 7.9 8 8.1

DTP

A-Z

n (

mg

kg-1

soil)

Soil pH

Loamy sand (g)

y = -3.306x + 22.22R² = 0.961

1.90

2.00

2.10

2.20

2.30

2.40

6 6.05 6.1 6.15

DTP

A-Z

n (

mg

kg-1

soil)

Soil pH

Sandy loam (h)

35

Application of Cu @ 10 and 20 mg kg-1

soil significantly increased DTPA-Zn by 10.2

and 25.5 per cent in loamy sand and by 13.5 and 16.5 per cent in sandy loam soil over control,

respectively. It increased from 0.98 mg kg-1

soil in control to 1.08 and 1.23 mg kg-1

soil in

loamy sand, and from 2.0 mg kg-1

soil in control to 2.27 and 2.33 mg kg-1

soil in sandy loam

soil with application of 10 and 20 mg Cu kg-1

soil, respectively which may be due to decrease

in soil pH due to Cu application (Table 4.1). DTPA- Zn was significantly negatively

correlated with soil pH (r= -0.476* n=24) in loamy sand soil (Fig. 4.3g, R2=0.908). In sandy

loam also DTPA- Zn was negatively correlated with soil pH (Fig. 4.3h, R2=0.961). It was

significantly positively correlated with Olsen P (r= 0.992** for loamy sand and 0.986** for

sandy loam soil) in both soils.

4.2 Effect of phosphorus and copper application on transformation of copper in soils.

The data on the effect of P and Cu application on various chemical pools of Cu in

soils are presented and discussed as under.

4.2.1 Exchangeable copper (EXCH-Cu)

Exchangeable Cu in both the soils decreased significantly over control with

increasing levels of applied P. In loamy sand calcareous soil it decreased from 0.301 mg kg-1

soil in control to 0.276, 0.248, 0.246, 0.212 and 0.224 mg kg-1

soil, respectively (Table 4.9,

Fig. 4.4a). In sandy loam soil, the exchangeable Cu decreased from 0.282 mg kg-1

soil in

control to 0.243, 0.230, 0.227, 0.221 and 0.254 mg kg-1

soil, with the application of 25,

50, 100, 200 and 400 mg kg-1

soil, respectively (Fig. 4.4b). Exchangeable Cu decreased

by 8.3, 17.6, 18.2, 29.5 and 25.5 per cent in loamy sand soil, and by 13.8, 18.4, 19.5, 21.6

and 9.9 per cent over control in sandy loam soil with increasing levels of applied P,

respectively. A non-significant increase in exchangeable Cu was observed in both the

soils when P was applied @ 400 mg kg-1

soil over the application of 200 mg P kg-1

soil.

In a laboratory study with a Typic Udic Ferrisols of China, Tu et al (2001) observed that

application of 80 mg P kg-1

soil decreased the exchangeable Cu but increased the content

of specifically adsorbed Cu.

The decrease in exchangeable Cu may be due to an increase in soil pH (Table 3.1). It

was significantly negatively correlated with soil pH in loamy sand soil (r= -0.656*) and

poorly correlated in sandy loam soil (r= 0.053) (Table 4.17). Several workers have reported a

decrease in EXCH-Cu in soils with increase in soil pH (Rupa and Shukla 1998, McLaren and

Crawford 1973). McLaren et al (1983) reported an increase in Cu sorption with increase in

pH and ascribed it to its greater association with soil oxides. Hickey and Kittrick (1984)

indicated that Cu had less mobility and bioavailability than Zn and it resided mostly in non-

available forms.

36

Table 4.9: Effect of phosphorus and copper application on exchangeable copper in

soils after harvest

Copper

levels

(mg kg-1

soil)

Exchangeable- Cu (mg kg-1

soil)

Phosphorus levels (mg kg-1

soil)

0 25 50 100 200 400 Mean

Loamy sand

0 0.071 0.058 0.055 0.072 0.056 0.052 0.060

5 0.071 0.066 0.098 0.056 0.045 0.050 0.064

10 0.220 0.268 0.120 0.162 0.118 0.104 0.165

20 0.845 0.713 0.718 0.694 0.631 0.691 0.715

Mean 0.301 0.276 0.248 0.246 0.212 0.224

CD (5%) P levels 0.034 Cu levels 0.028 P x Cu 0.069

Sandy loam

0 0.083 0.062 0.063 0.064 0.066 0.043 0.063

5 0.090 0.070 0.065 0.106 0.108 0.166 0.101

10 0.190 0.160 0.153 0.138 0.166 0.280 0.181

20 0.767 0.680 0.641 0.600 0.546 0.526 0.627

Mean 0.282 0.243 0.230 0.227 0.221 0.254

CD (5%) P levels 0.041 Cu levels 0.033 P x Cu 0.082

Application of Cu resulted in a significant increase in exchangeable Cu in both the

soils. It increased significantly from 0.060 mg kg-1 soil in control to 0.165 and 0.715 mg kg

-1

soil in loamy sand soil (Table 4.9, Fig. 4.5a) and from 0.063 mg kg-1

soil in control to 0.181 and

0.627 mg kg-1

soil in sandy loam soil with application of 10 and 20 mg Cu kg-1 soil, respectively

(Fig. 4.5b). Exchangeable Cu increased by 175 and 1091per cent in loamy sand soil, and by 187

and 895 per cent in sandy loam soil over control with application of 10 and 20 mg Cu kg-1

soil,

respectively. In both the soils, a greater increase in exchangeable Cu was observed when it was

applied @ 20 mg kg-1

soil. Exchangeable Cu was significantly positively correlated with SAD-

Cu, CARB-Cu, MnOX-Cu, AMPOX-Cu, CRYOX-Cu, OM-Cu and RES-Cu in both the soils

(Table 4.17). Soil EC was significantly positively correlated with exchangeable Cu (r=0.746**

for loamy sand and 0.624** for sandy loam) in both the soils.

4.2.2. Specifically adsorbed copper (SAD-Cu)

Metals held on soil surfaces by covalent type bonding (Sposito 1984) to oxide or

organic functional groups is termed as specifically adsorbed or inner sphere complexed

metals. In loamy sand soil, specifically adsorbed Cu decreased significantly over control

37

when P was applied @ 100, 200 and 400 mg P kg-1

soil (Table 4.10, Fig. 4.4a). It decreased

from 0.277 mg kg-1

soil in control to 0.217, 0.230 and 0.211 mg kg-1

soil with application of

100, 200 and 400 mg P kg-1

soil, respectively. Whereas in sandy loam soil a significant

decrease in SAD-Cu was observed only when 100 mg P kg-1

soil was applied and thereafter it

tended to increase when 200 and 400 mg P kg-1

soil was applied (Fig. 4.4b). It decreased

significantly from 0.163 mg kg-1

soil in control to 0.130 mg kg-1

soil with application of 100

mg P kg-1

soil and therefore it increased to 0.181 and 0.184 with application of 200 and 400

mg P kg-1

soil, respectively. In loamy sand soil, SAD-Cu increased by 3.9 and 2.16 per cent

over control with application of 25 and 50 mg P kg-1

soil but in sandy loam soil application of

200 and 400 mg P kg-1

soil increased the SAD-Cu by 11.04 and 12.88 per cent, respectively

over control. The content of Cu in SAD-Cu was higher in loamy sand soil as compared to

sandy loam soil. Tu et al (2001) also observed that application of 80 mg P kg-1

soil increased

the content of specifically adsorbed Cu.

Table 4.10: Effect of phosphorus and copper application on specifically adsorbed copper

in soils after harvest

Copper

levels

(mg kg-1

soil)

Specifically adsorbed- Cu (mg kg-1

soil)

Phosphorus levels (mg kg-1

soil)

0 25 50 100 200 400 Mean

Loamy sand

0 0.087 0.085 0.078 0.079 0.077 0.074 0.080

5 0.090 0.095 0.077 0.086 0.098 0.099 0.091

10 0.138 0.149 0.153 0.174 0.196 0.192 0.167

20 0.793 0.821 0.826 0.529 0.550 0.478 0.664

Mean 0.277 0.288 0.283 0.217 0.230 0.211

CD (5%) P levels 0.035 Cu levels 0.028 P x Cu0.070

Sandy loam

0 0.048 0.044 0.042 0.048 0.065 0.064 0.052

5 0.080 0.090 0.090 0.090 0.095 0.086 0.088

10 0.184 0.196 0.169 0.134 0.140 0.170 0.165

20 0.340 0.314 0.305 0.250 0.424 0.418 0.342

Mean 0.163 0.161 0.152 0.130 0.181 0.184

CD (5%) P levels 0.026 Cu levels 0.021 P x Cu 0.052

38

Specifically adsorbed Cu in both the soils increased significantly over control with

application of 10 and 20 mg Cu kg-1

soil. In loamy sand soil it increased from 0.080 mg kg-1

soil in control to 0.167 and 0.664 mg kg-1

soil (Table 4.10, Fig. 4.5a and in sandy loam soil it

increased from 0.052 mg kg-1

soil in control to 0.165 and 0.342 mg kg-1

soil with application

of 10 and 20 mg Cu kg-1

soil, respectively (Fig. 4.5b). Specifically adsorbed Cu increased by

108 and 730 per cent in loamy sand and by 217 and 557 per cent in sandy loam soil over

control with application of 10 and 20 mg Cu kg-1

soil, respectively.

The increase in SAD-Cu may be due to greater adsorption of Cu by soil colloids.

High molecular weight organic colloids diminish as soil pH increases to expose surfaces

where metals can be adsorbed (Geering and Hodgson 1969). A significant interaction

effect of P and Cu levels on SAD- Cu was observed in both the soils. When Cu was

applied @ 20 mg kg-1

soil, in loamy sand soil SAD-Cu decreased significantly from 0.793

mg kg-1

soil in control to 0.550 and 0.478 mg kg-1

soil but in sandy loam soil it increased

from 0.340 mg kg-1

soil in control to 0.424 and 0.418 mg kg-1

soil with application of 200

and 400 mg kg-1

soil, respectively (Table 4.10). Specifically adsorbed Cu was

significantly negatively correlated with soil pH (r=-0.562*) and positively with soil EC

(r=0.746**) in loamy sand soil. Specifically adsorbed Cu was significantly positively

correlated with CARB-Cu, MnOX-Cu, AMPOX-Cu, CRYOX-Cu, OM-Cu and RES-Cu in

both the soils (Table 4.17).

4.2.3 Carbonate bound copper (CARB-Cu)

In loamy sand calcareous soil, carbonate bound Cu increased significantly from 0.425

mg kg-1

soil in control to 0.562, 0.578, 0.579 and 0.574 mg kg-1

soil with application of 50,

100, 200 and 400 mg P kg-1

soil which represented an increase of 32.2, 36.0, 36.2 and 35.05

per cent over control, respectively (Table 4.11, Fig. 4.4a). However, the differences in CARB-

Cu at these rates of applied P were not significant. Application of Cu resulted in a significant

increase in CARB-Cu. It increased from 0.096 mg kg-1

soil in control to 0.231, 0.481 and

1.312 mg kg-1

soil, respectively when Cu was applied @ 5, 10 and 20 mg kg-1

soil (Fig. 4.5a)

and this increase was about 140, 401 and 1266 per cent over control, respectively. Cu is

usually high in the carbonate fraction in alkaline calcareous soils and in Fe oxides fraction in

acid soils (Hickey and Kittrick 1984, Sims 1986). The interaction effect was not significant.

Carbonate bound Cu was not determined in sandy loam soil as it was non-calcareous soil. In

loamy sand soil it was significantly negatively correlated with soil pH (r= -0.692**) and

positively with soil EC (r=0.887**) (Table 4.17). Carbonate bound Cu was significantly

positively correlated with MnOX-Cu, AMPOX-Cu, CRYOX-Cu, OM-Cu and RES-Cu in

loamy sand calcareous soil (Table 4.17).

39

Table 4.11: Effect of phosphorus and copper application on carbonate bound copper in

soil after harvest

Copper

levels

(mg kg-1

Carbonate bound – Cu (mg kg-1

soil)

Phosphorus levels (mg kg-1

soil)

0 25 50 100 200 400 Mean

soil) Loamy sand

0 0.054 0.092 0.086 0.092 0.108 0.144 0.096

5 0.161 0.180 0.256 0.262 0.240 0.286 0.231

10 0.328 0.405 0.436 0.438 0.601 0.678 0.481

20 1.158 1.169 1.469 1.522 1.368 1.188 1.312

Mean 0.425 0.461 0.562 0.578 0.579 0.574

CD (5%) P levels 0.107 Cu levels 0.088 P x Cu NS

4.2.4 Manganese oxides bound copper (MnOX-Cu)

Application of 25 and 50 mg P kg-1

soil significantly increased the MnOX-Cu by 19.0

and 21.6 per cent over control in loamy sand soil. It increased from 0.236 mg kg-1

soil in

control to 0.281 and 0.287 mg kg-1

soil, respectively (Table 4.12, Fig 4.4a). This increase in

MnOX-Cu may be due to an increase in soil pH (Table 4.1) because the adsorption of metals

by oxides increases with increase in soil pH. But it decreased (41.5%) significantly to 0.138

mg kg-1

soil when application of P increased to 400 mg kg-1

soil due to decrease in soil pH at

this level (Table 4.1). In sandy loam acidic soil, it increased (33.57%) from 0.140 mg kg-1

soil

in control to 0.187 mg kg-1

soil with application of 400 mg P kg-1

soil (Fig. 4.4b) due to an

increase in soil pH.

An increase in Fe and Mn oxides bound Cu in an acidic soil of China due to P

application was observed by Tu et al (2001). In soils rich in Fe oxides P may enhance the

specific adsorption of heavy metals on Fe and Al hydroxides (Xie and MacKenzie 1988).

Copper application significantly increased MnOX-Cu in both the soils. It

increased from 0.069 mg kg-1

soil in control to 0.585 mg kg-1

soil in loamy sand soil (747

per cent) (Fig, 4.5a) and from 0.066 mg kg-1

soil in control to 0.295 mg kg-1

soil in sandy

loam soil (346 per cent) when Cu was applied @ 20 mg kg-1

soil (Fig. 4.5b).

Comparatively higher amounts of MnOx-Cu were observed in loamy sand than sandy

loam soil.

40

Table 4.12: Effect of phosphorus and copper application on manganese oxides bound

copper in soil after harvest

Copper

levels

Manganese oxide bound – Cu (mg kg-1

soil)

Phosphorus levels (mg kg-1

soil)

(mg kg-1

soil)

0 25 50 100 200 400 Mean

Loamy sand

0 0.072 0.071 0.076 0.071 0.064 0.062 0.069

5 0.072 0.088 0.086 0.128 0.098 0.080 0.092

10 0.206 0.286 0.192 0.125 0.184 0.146 0.190

20 0.596 0.681 0.794 0.732 0.448 0.264 0.585

Mean 0.236 0.281 0.287 0.264 0.198 0.138

CD (5%) P levels 0.039 Cu levels 0.032 P x Cu 0.077

Sandy loam

0 0.050 0.056 0.068 0.068 0.066 0.090 0.066

5 0.081 0.085 0.097 0.138 0.088 0.088 0.096

10 0.162 0.152 0.126 0.128 0.154 0.198 0.153

20 0.266 0.289 0.280 0.232 0.328 0.374 0.295

Mean 0.140 0.145 0.143 0.141 0.159 0.187

CD (5%) P levels 0.027 Cu levels 0.022 P x Cu 0.055

A significant interaction effect of P and Cu application on MnOX-Cu revealed that

when 20 mg Cu kg-1

soil was applied, its content decreased significantly from 0.596 mg kg-1

soil in control to 0.448 and 0.264 mg kg-1

soil in loamy sand soil, and increased significantly

from 0.266 mg kg-1

soil in control to 0.328 and 0.374 mg kg-1

soil in sandy loam soil when

200 and 400 mg P kg-1

soil was applied, respectively. It was significantly negatively

correlated with soil pH (r= -0.476* for loamy sand soil) and positively with soil EC

(r=0.727** for loamy sand and 0.670* for sandy loam soil). In calcareous soils of India,

Singh et al (1988) observed a negative correlation of soil pH with Mn oxides bound Cu while

Singh (2005) observed significant positive correlation of soil pH with MnOX-Cu. Manganese

oxides bound Cu was significantly positively correlated with AMPOX-Cu, CRYOX-Cu, OM-

Cu and RES-Cu in both the soils (Table 4.17).

4.2.5 Amorphous oxides bound copper (AMPOX-Cu)

In loamy sand alkaline calcareous soil, amorphous oxides bound Cu increased

significantly over control with graded levels of applied P. Its content increased significantly

from 3.44 mg kg-1

soil in control to 3.84, 3.81, 3.78, 3.78 and 3.52 mg kg-1

soil with

41

application of 25, 50, 100, 200 and 400 mg P kg-1

soil (Table 4.13, Fig. 4.4a) which

represented an increase of 11.62, 10.75, 9.88, 9.88 and 2.32 per cent respectively over control.

However, it decreased significantly by 6.8% with application of 400 mg kg-1

soil compared

with application of 200 mg kg-1

soil. The differences in the content of amorphous oxides

bound Cu observed with application of 25, 50, 100 and 200 mg kg-1

soil were not significant.

Shuman (1988b) also observed a significant increase in Cu held by amorphous oxides by P

application in coarse as well as fine textured soils.

Table 4.13: Effect of phosphorus and copper application on amorphous oxides bound

copper in soils after harvest

Copper

levels

(mg kg-1

Amorphous oxides bound – Cu (mg kg-1

soil)

Phosphorus levels (mg kg-1

soil)

0 25 50 100 200 400 Mean

soil) Loamy sand

0 0.48 0.52 0.52 0.53 0.56 0.59 0.53

5 2.32 2.46 2.61 2.25 2.23 2.49 2.39

10 3.90 5.04 4.79 5.42 5.04 4.58 4.80

20 7.03 7.35 7.31 6.93 7.29 6.40 7.05

Mean 3.44 3.84 3.81 3.78 3.78 3.52

CD (5%) P levels 0.172 Cu levels 0.140 P x Cu 0.345

Sandy loam

0 1.77 1.78 1.72 1.78 1.76 1.70 1.75

5 3.79 4.01 4.00 4.94 3.79 4.13 4.11

10 6.50 5.95 5.62 5.07 5.22 5.36 5.62

20 9.89 9.75 8.55 7.83 7.69 8.10 8.63

Mean 5.49 5.37 4.97 4.90 4.62 4.82

CD (5%) P levels 0.50 Cu levels 0.41 P x Cu 1.00

The mechanism of adsorption of metals on oxides due to P application includes that P

can act as a bridge between the soil oxides surfaces and the sorbed heavy metals thus leading

to an increase in adsorption of these metals on oxides and following chemical species may be

formed (Xie and MacKenzie 1988, Bolland et al 1977).

Fe-OH Fe-OH O

O + H2PO4- + M

2+ = O P + 2H2O

Fe-OH Fe-OH O-M+

42

In contrast to loamy sand soil, AMPOX-Cu decreased significantly by 2.18, 9.47, 10.79,

15.84 and 12.2 per cent over control with graded levels of applied in sandy loam acidic soil. It

decreased significantly from 5.49 mg kg-1 soil in control to 5.37, 4.97, 4.90, 4.62 and 4.82 mg kg

-1

soil with application of 25, 50, 100, 200 and 400 mg kg-1 soil, respectively (Fig. 4.4b).

Copper application significantly increased AMPOX-Cu from 0.534 mg kg-1 soil in

control to 2.39, 4.80 and 7.05 mg kg-1 soil in loamy sand soil (Fig. 4.5a) and from 1.75 mg kg

-1

soil in control to 4.11, 5.62 and 8.63 mg kg-1 soil in sandy loam with application of 5, 10 and 20

mg Cu kg-1 soil, respectively (Fig. 4.5b). Thus, the content of Cu in amorphous oxides increased

by 347, 798 and 1220 per cent in loamy sand and by 134, 221 and 393 per cent in sandy loam over

control with application of 5, 10 and 20 mg Cu kg-1 soil, respectively. The absolute amounts of Cu

in this fraction were observed to be more in medium textured sandy loam soil as compared to light

textured loamy sand soil. The amorphous Fe and Al oxides have high specific surface area and

hence posses very high adsorption capacity for Cu (Okazaki et al 1986). Studies on Cu adsorption

after selective extraction showed that amorphous Fe oxides are more important than organic

matter in adsorption (Cavallaro and McBride 1984) and that oxide forms in general and organic

coatings have a high affinity for metals (Lion et al 1982).

A significant interaction effect of applied P and Cu levels on AMPOX-Cu in light

textured soil revealed that when Cu was applied @ 10 mg kg-1

soil, its content increased

significantly at each level of applied P compared to when no P was applied. But in medium

textured sandy loam soil when Cu was applied @ 20 mg kg-1

soil, its content decreased

significantly when P was applied @ 50, 100, 200 and 400 mg kg-1

soil as compared to when

no P was applied. It was significantly negatively correlated with soil pH (r=-0.568* for loamy

sand) but positively with soil EC (r=0.914** for loamy sand and 0.709** for sandy loam)

(Table 4.17). A negative coefficient of correlation of soil pH with amorphous oxides bound

Cu in calcareous soils of India was observed by Singh et al (1988). Amorphous oxides bound

Cu was significantly positively correlated with CRYOX-Cu, OM-Cu and RES-Cu in both the

soils (Table 4.17).

4.2.6 Crystalline oxides bound copper (CRYOX-Cu)

The data on the effect of P and Cu application on crystalline oxides bound Cu is

presented in Table 4.14. In loamy sand soil it increased significantly from 1.95 mg kg-1

soil in

control to 2.16 mg kg-1

soil thereby resulting in an increase of 10.76 per cent over control

with application of 25 mg P kg-1

soil. However, its content decreased significantly by 1.3,

11.5, 15.27 and 17.12 per cent with application of 50, 100, 200 and 400 mg P kg-1

soil,

respectively compared to application of 25 mg P kg-1

soil (Fig 4.4a).

In medium textured sandy loam soil, the content of Cu in crystalline oxides decreased

significantly from 3.08 mg kg-1 soil in control to 2.61, 2.60, 2.43, 2.32 and 2.40 mg kg

-1 soil with

application of 25, 50, 100, 200 and 400 mg P kg-1 soil which represented a decrease of 15.2, 15.5,

43

21.1, 24.6 and 22.0 per cent, respectively over control (Fig. 4.4b). Shuman (1988b) observed a

significant decrease in the content of crystalline oxides bound Cu in fine textured and an increase

in coarse textured acidic soils with P application @ 60 mg P kg-1 soil.

Table 4.14: Effect of phosphorus and copper application on crystalline oxides bound

copper in soils after harvest

Copper

levels

(mg kg-1

soil)

Crystalline oxides bound – Cu (mg kg-1

soil)

Phosphorus levels (mg kg-1

soil)

0 25 50 100 200 400 Mean

Loamy sand

0 1.30 1.27 1.38 1.34 1.26 1.28 1.30

5 1.62 1.97 2.26 1.73 1.20 1.40 1.70

10 2.28 2.37 2.20 1.77 2.36 1.69 2.11

20 2.60 3.02 2.66 2.80 2.49 2.78 2.73

Mean 1.95 2.16 2.13 1.91 1.83 1.79

CD (5%) P levels 0.128 Cu levels 0.105 P x Cu 0.257

Sandy loam

0 1.91 1.51 1.37 1.32 1.26 1.26 1.44

5 1.74 2.71 1.87 1.92 2.09 1.29 1.93

10 3.43 2.92 3.04 2.51 3.02 2.73 2.94

20 5.26 3.32 4.11 3.96 2.92 4.34 3.98

Mean 3.08 2.61 2.60 2.43 2.32 2.40

CD (5%) P levels 0.342 Cu levels 0.279 P x Cu 0.684

Application of Cu significantly increased the content of CRYOX-Cu at each level of

applied Cu in both the soils. Its content increased from 1.30 mg kg-1 soil in control to 1.70, 2.11

and 2.73 mg kg-1 soil in loamy sand (Fig. 4.5a) and from 1.44 mg kg

-1 soil in control to 1.93, 2.94

and 3.98 mg kg-1 soil in sandy loam soil, with application of 5, 10 and 20 mg Cu kg

-1 soil,

respectively (Fig. 4.5b). The content of Cu in this fraction increased by about 110 and 176 per cent

over control in loamy sand and sandy loam soil, respectively with the highest level of applied Cu.

A significant interaction effect of P and Cu levels on CRYOX-Cu revealed that at each level of

applied P, the content of Cu in crystalline oxides increased significantly over control when Cu was

applied @ 20 mg kg-1 soil in both the soils. Overall the content of Cu in crystalline oxides in

medium textured soil was more as compared to light textured soil. Crystalline oxides bound Cu

was significantly negatively correlated with soil pH (r=-0.570* for loamy sand) and positively

with soil EC (r=0.793** for loamy sand and 0.554* for sandy loam). Singh et al (1988) also

observed a negative coefficient of correlation of soil pH with crystalline oxides bound Cu in

calcareous soils of India.

44

4.2.7 Organically bound copper (OM-Cu)

In loamy sand soil, organically bound Cu increased (5.95%) significantly from 1.68

mg kg-1

soil in control to 1.78 mg kg-1

soil when P was applied @ 100-400 mg kg-1

soil (Table

4.15, Fig. 4.4a). Similarly, in sandy loam soil, its content increased significantly from 0.72

mg kg-1

soil in control to 0.94 and 1.36 mg kg-1

soil when P was applied @ 25 and 400 mg kg-

1 soil thus resulting in an increase of 30.5 and 88.8 per cent over control, respectively (Fig.

4.4b). The differences in OM-Cu observed with application of 50, 100 and 200 mg P kg-1

soil

were not significant. Organically bound Cu increased significantly from 1.65 mg kg-1

soil in

control to a maximum of 1.9 mg kg-1

soil in loamy sand (Table 4.15, Fig. 4.5a) and from 0.75

mg kg-1

soil in control to 1.24 mg kg-1

soil in sandy loam resulting in an increase of 15 and

65.3 per cent over control with application of 20 mg Cu kg-1

soil, respectively (Table 4.15,

Fig. 4.5b).

Table 4.15: Effect of phosphorus and copper application organically bound copper in

soils after harvest

Copper

levels (mg

kg-1

soil)

Organically bound – Cu (mg kg-1

soil)

Phosphorus levels (mg kg-1

soil)

0 25 50 100 200 400 Mean

Loamy sand

0 1.60 1.61 1.61 1.62 1.67 1.78 1.65

5 1.67 1.66 1.62 1.78 1.71 1.70 1.69

10 1.60 1.71 1.64 1.81 1.85 1.75 1.72

20 1.86 1.74 2.15 1.91 1.88 1.90 1.90

Mean 1.68 1.68 1.75 1.78 1.78 1.78

CD (5%) P levels 0.087 Cu levels 0.074 P x Cu 0.175

Sandy loam

0 0.61 0.61 0.74 0.78 0.83 0.89 0.75

5 0.74 0.79 0.92 0.79 0.82 1.51 0.93

10 0.74 0.80 0.94 0.95 0.92 1.53 0.98

20 0.77 1.57 1.01 1.12 1.42 1.54 1.24

Mean 0.72 0.94 0.90 0.91 0.99 1.36

CD (5%) P levels 0.072 Cu levels 0.059 P x Cu 0.144

45

Fig. 4.4: Copper content in various chemical pools as influenced by applied P

0

1

2

3

4

5

6

7

0 25 50 100 200 400

Cu

(m

g k

g-1

soil

)

Levels of applied P (mg kg-1 soil)

EXCH-Cu

SAD-Cu

CARB-Cu

MnOX-Cu

AMPOX-Cu

CRYOX-Cu

OM-Cu

RES-Cu

0

3

6

9

0 25 50 100 200 400

Cu

(m

g k

g-1

soil

)

Levels of applied P (mg kg-1 soil)

EXCH-Cu

SAD-Cu

MnOX-Cu

AMPOX-Cu

CRYOX-Cu

OM-Cu

RES-Cu

Loamy sand

Sandy loam

(a)

(b)

46

Fig. 4.5: Copper content in various chemical pools as influenced by applied Cu

0

2

4

6

8

10

12

0 5 10 20

Cu

(m

g k

g-1

soil

)

Levels of applied Cu (mg kg-1 soil)

EXCH-Cu

SAD-Cu

CARB-Cu

MnOX-Cu

AMPOX-Cu

CRYOX-Cu

OM-Cu

RES-Cu

0

2

4

6

8

10

12

14

0 5 10 20

Cu

(m

g k

g-1

soil

)

Levels of applied Cu (mg kg-1 soil)

EXCH-Cu

SAD-Cu

MnOX-Cu

AMPOX-Cu

CRYOX-Cu

OM-Cu

RES-Cu

Loamy sand

Sandy loam

(a)

(b)

47

A significant interaction effect of P and Cu levels in both the soils revealed that

application of 20 mg Cu kg-1

soil significantly increased the contents of OM-Cu over

control at each level of applied P. Though the content of organic matter was higher in

sandy loam soil as compared to sandy loam, the content of Cu in organic matter fraction

was higher in loamy sand soil compared to sandy loam soil. Organically bound Cu was

poorly but positively correlated with soil pH. However, it was significantly positively

correlated with soil EC (r=0.528* for loamy sand and 0.659* for sandy loam) in both the

soils. While studying the effect of graded levels of applied P on transformation of native

Cu in a near neural Typic Haplustept, Singh (2005) also observed a positive correlation of

soil pH with OM-Cu. Among the heavy metals, Cu is known to possess the highest

affinity for organic matter (Hickey and Kittrick 1984). The results of the present study

also indicated that added as well as native Cu dissolved from other solid phases of Cu due

to P application tended to associate with the soil organic matter.

4.2.8 Residual copper (RES-Cu)

P application significantly decreased the content of Cu in residual mineral fraction

over control in loamy sand soil except when it was applied @ 400 mg kg-1

soil where its

content increased significantly over that observed with application of 200 mg kg-1

soil (Table

4.16). Residual Cu decreased by 9.71, 10.43, 6.23 and 3.7 per cent over control with

application of 25, 50, 100 and 200 mg P kg-1

soil in loamy sand (Fig. 4.4a). Shuman (1988b)

reported that P additions shifted Cu from residual to exchangeable fractions thus making it

more plant available. In sandy loam soil, the content of Cu in residual fraction increased

significantly by 10.7, 13.6, 16.8 and 8.25 per cent over control when P was applied @ 50,

100, 200 and 400 mg kg-1

soil, though it decreased significantly by 7.3 per cent with

application of 400 mg P kg-1

soil when compared to 200 mg P kg-1

soil (Fig. 4.4b).

Application of Cu significantly increased the content of residual Cu when it was

applied @ 5, 10 and 20 mg kg-1

soil in both the soils. Its content increased by 325 per cent in

loamy sand (Fig. 4.5a) and by 160 per cent in sandy loam (Fig. 4.5b) over control with the

highest level of applied Cu. A significant interaction effect of P and Cu level on Res-Cu

revealed that in loamy sand soil the content of Cu in residual fraction increased significantly

when P was applied @ 400 mg kg-1

soil compared to an application of 200 mg P kg-1

soil. But

in sandy loam soil, the content of Cu in residual fraction decreased significantly by

application of 400mg P kg-1

soil compared to when it was applied @ 200 mg kg-1

soil. This

indicated that in medium textured acidic soils high rate of applied P may lead to dissolution of

residual mineral fraction of Cu which may be redistributed among different pools of Cu. But

48

in light textured alkaline calcareous soils applied Cu may enter the residual mineral fraction

when high rates of P are applied thus rendering it unavailable to the plants.

Table 4.16: Effect of phosphorus and copper application on residual copper in soil after

harvest

Copper

levels

(mg kg-1

soil)

Residual Cu (mg kg-1

soil)

Phosphorus levels (mg kg-1

soil)

0 25 50 100 200 400

Loamy sand

0 2.83 2.79 2.69 2.70 2.70 2.50

5 5.49 4.97 4.48 5.20 5.86 5.38

10 7.77 6.18 6.99 6.66 6.13 7.35

20 11.53 11.00 10.56 11.30 11.87 12.75

Mean 6.90 6.23 6.18 6.47 6.64 6.99

CD (5%) P levels 0.30 Cu levels 0.25 P x Cu 0.61

Sandy loam

0 5.01 5.42 5.48 5.43 5.44 5.45

5 7.96 6.68 7.44 6.50 7.49 7.21

10 8.28 9.34 9.45 10.56 9.87 9.24

20 12.18 13.56 14.67 15.50 16.29 14.31

Mean 8.36 8.75 9.26 9.50 9.77 9.05

CD (5%) P levels 0.704 Cu levels 0.57 P x Cu 1.40

Linear coefficients of correlation among various chemicals pools of Cu (Table 4.17)

indicated that most of these pools were significantly and positively correlated amongst

themselves thereby suggesting the existence of a dynamic equilibrium among different forms.

Thus, the depletion in the concentration of one pool either due to plant uptake or its

transformation may be replenished by other pools.

49

Table 4.17: Linear coefficients of correlation among various pools of copper and soil

properties (n=24)

Loamy sand soil

EXCH-

Cu

SAD-Cu CARB-

Cu

MnOX-

Cu

AMPOX-

Cu

CRYOX-

Cu

OM-Cu

SAD-Cu 0.969**

CARB-Cu 0.935**

0.913**

MnOX-Cu 0.917**

0.950**

0.912**

AMPOX-Cu 0.847**

0.826**

0.912**

0.827**

CRYOX-Cu 0.833**

0.802**

0.835**

0.805**

0.901**

OM-Cu 0.871**

0.852**

0.922**

0.857**

0.995**

0.935**

RES-Cu 0.898**

0.847**

0.934**

0.804**

0.944**

0.856**

0.589**

pH -0.656**

-0.562**

-0.692**

-0.476* -0.568

** -0.570

** 0.287

EC 0.764**

0.746**

0.887**

0.727**

0.914**

0.793**

0.528**

Olsen P -0.076 -0.106 0.083 -0.208 -0.013 -0.178 -0.224

DTPA-Cu 0.903**

0.884**

0.946**

0.847**

0.965**

0.890**

0.655**

Sandy loam soil

SAD-Cu 0.928**

MnOX-Cu 0.899**

0.921**

AMPOX-Cu 0.904**

0.956**

- 0.897**

CRYOX-Cu 0.837**

0.907**

- 0.817**

0.903**

OM-Cu 0.905**

0.963**

- 0.897**

0.992**

0.951**

RES-Cu 0.888**

0.893**

- 0.921**

0.896**

0.802**

0.619**

pH 0.053 0.003 - -0.017 -0.006 0.044 0.161

EC 0.536**

0.624**

- 0.670**

0.709**

0.554**

0.659**

Olsen P 0.010 -0.020 - 0.195 -0.061 -0.129 0.612**

DTPA-Cu 0.863**

0.939**

0.907**

0.972**

0.880**

0.546**

*Significant at 5%

**Significant at 1%

4.2.9 Per cent recovery of applied Cu @ 20 mg kg-1

soil in different chemical pools

When Cu is applied to soil though external sources many chemical reactions like

adsorption, precipitation and complexation take place. The percent recovery of added Cu @

20 mg kg-1

soil at different levels of applied P in a particular fraction was calculated by

taking the difference of the content of that fraction in Cu treated pot and the corresponding

no Cu pots (Table 4.18). Soil Cu is largely found in unavailable forms and it was observed

that in loamy sand soil only 3.27 and 2.93 per cent and in sandy loam soil 2.81 and 1.44 per

cent of applied Cu @ 20 mg kg-1

soil entered into exchangeable and specifically adsorbed

fractions, respectively which are considered to be the most plant available forms. A major

50

portion (43-45%) of the added Cu entered into residual mineral fraction followed by

amorphous oxides (32-34%) and crystalline oxides (7.1-12.7%) in both the soils. It is due to

the fact that Cu has a strong tendency to associate with the crystalline structures of the

minerals and organic ligands (Fuentes et al 2004, Nemati et al 2009). Guan et al (2011)

also reported that about 51 and 45 per cent of applied Cu accumulated in residual and oxide

forms of Cu in a Chinese soil. Shuman (1988b) also reported that added Cu was evident in

exchangeable, organic and amorphous oxides fraction in light textured soils. About 6.08 per

cent of added Cu also entered in to carbonate bound fraction in alkaline calcareous loamy

sand soil because metals can also be associated with soil carbonates (Shuman 1991). Only

1.29 and 2.47 per cent of added Cu entered into OM-Cu in loamy sand and sandy loam

soils, respectively. Mean values of per cent recovery of applied Cu @ 20 mg kg-1

soil in

various pools of Cu presented in Table 4.18 were used to study the distribution and

movement of Cu in to its various chemical pools in the presence and absence of applied P.

The data indicated that in loamy sand soil, with P application plant available forms like

EXCH-Cu and SAD-Cu decreased but CARB-Cu increased and most of the applied Cu

redistributed into the various chemical pools and a very little of it got in to RES-Cu (Fig.

4.6a,b). In sandy loam soil, applied P decreased EXCH-Cu, AMPOX-Cu and CRYOX-Cu

but increased the Cu content in OM-Cu and RES-Cu (Fig. 4.6c, d).

Table 4.18 Percent recovery of applied Cu @ 20 mg kg-1

soil in different fractions of soil

Loamy sand soil

Levels of P EXCH-

Cu

SAD-

Cu

CARB-

Cu

MnOX-

Cu

AMPOX-

Cu

CRYOX-

Cu

OM-

Cu

RES-

Cu

0 3.87 3.53 5.52 2.62 32.71 6.50 1.30 42.80

25 3.27 3.68 5.38 3.05 34.16 8.75 0.65 40.80

50 3.31 3.74 6.91 3.59 33.94 6.40 2.70 38.90

100 3.11 2.25 7.15 3.30 32.02 7.30 1.45 42.30

200 2.87 2.36 6.30 1.92 33.60 6.15 1.05 47.20

400 3.19 2.02 5.22 1.01 29.02 7.50 0.60 49.80

Mean 3.27 2.93 6.08 2.58 32.50 7.10 1.29 43.63

Sandy loam soil

0 3.42 1.46 ND 1.08 40.60 16.80 0.80 35.80

25 3.09 1.35 ND 1.16 39.80 9.05 4.80 40.70

50 2.89 1.31 ND 1.06 34.10 13.70 1.35 45.90

100 2.68 1.01 ND 0.82 30.20 13.20 1.70 50.30

200 2.40 1.79 ND 1.31 29.60 8.30 2.95 54.20

400 2.41 1.77 ND 1.42 32.00 15.40 3.25 44.70

Mean 2.81 1.44 ND 1.14 34.40 12.70 2.47 45.30

ND-not determined

51

4.2.10 Distribution of native Cu in different chemical pools

Distribution of native Cu in different fractions as a percent of total Cu as influenced

by P fertilization are presented in Table 4.19 and Fig. 4.7. A major portion of Cu resided in

residual mineral fraction in both the soils. Mean RES-Cu was 41.5 and 56.5 per cent in loamy

sand and sandy loam soil, respectively. In loamy sand soil the crystalline Fe and Al oxides

bound Cu decreased from 20.0% to 19.6% as the level of P application increased from 0 to

400 mg P kg-1

soil but that of OM-Cu increased from 25.8% to 27.3% and that of AMPOX-

Cu increased from 7.5% to 9.16% as the level of P application increased from 0 to 400 mg P

kg-1

soil. The data indicated that only 0.93% and 1.23% of total Cu was present in EXCH-Cu

and SAD-Cu fractions, respectively, which are considered to be the most plant available

forms. The residual mineral fraction decreased from 43.5 % to 38.4% as the level of P

application increased from 0 to 400 mg P kg-1

soil thereby indicating that addition of P tended

to redistribute the native Cu among the other pools of Cu. So, application of P moved Cu

from residual mineral pool to OM-Cu, AMPOX-Cu and CARB-Cu in alkaline calcareous soil.

Table 4.19: Percent distribution of native Cu in different chemical pools

Per cent of Total copper

Loamy sand

Levels of P

(mg kg-1

soil)

EXC –

Cu

SAD-

Cu

CARB-

Cu

MnOX-

Cu

AMPOX-

Cu

CRYOX-

Cu

OM-

Cu

RES-

Cu

0 1.09 1.33 0.83 1.10 7.50 20.00 25.80 43.50

25` 0.89 1.30 1.41 1.09 7.96 19.50 25.80 42.90

50 0.84 1.20 1.32 1.16 8.01 21.20 26.90 41.30

100 1.10 1.21 1.41 1.09 8.09 20.60 27.30 41.50

200 0.86 1.18 1.66 0.98 8.58 19.30 27.30 41.50

400 0.80 1.13 2.21 0.95 9.16 19.60 27.30 38.40

Mean 0.93 1.23 1.47 1.06 8.22 20.10 26.70 41.50

Sandy loam

0 0.74 0.50 - 0.52 18.60 20.10 6.42 52.70

25 0.61 0.46 - 0.58 18.70 15.80 6.42 57.00

50 0.57 0.44 - 0.71 18.10 14.40 7.78 57.70

100 0.75 0.50 - 0.71 18.70 13.80 8.21 57.10

200 0.58 0.68 - 0.69 18.50 13.20 8.73 57.30

400 0.54 0.67 - 0.52 17.80 13.20 9.36 57.40

Mean 0.63 0.54 - 0.62 18.40 15.10 7.82 56.50

52

In sandy loam soil, the RES-Cu increased from 52.7% to 57.3% as the level of P

application increased from 0 to 400 mg P kg-1

soil. Organically bound Cu increased from

6.42% to 9.36% and CRYOX-Cu decreased from 20.1% to 14.8%. Only 0.63% and 0.54% of

total Cu was present in EXCH-Cu and SAD-Cu fractions, respectively, and these contents

were quite lower as compared to that in loamy sand soil indicating that light textured soils

may contain more amounts of plant available forms of Cu.

Mean values of per cent distribution of native Cu in various pools of Cu presented in

Table 4.19 were used to study the distribution and movement of Cu in to its various chemical

pools in the presence and absence of applied P. The data indicated that in loamy sand soil

application of P moved Cu from RES-Cu to AMPOX-Cu and CARB-Cu (Fig. 4.7a, b) and in

sandy loam soil application of P moved Cu from EXCH-Cu, SAD-Cu, and CRYOX-Cu forms

to OM-Cu and RES-Cu (Fig. 4.7c, d).

4.3 Effect of phosphorus and copper application on root dry matter yield, grain and

straw yield

4.3.1 Visual observations

In the absence of applied Cu, growth of the crop improved significantly with graded

levels of applied P in loamy sand (Plate1) as well as sandy loam (Plate 2) soil. However,

when each level of P was applied in combination with 20 mg Cu kg-1

soil as Cu-EDTA in

loamy sand soil, severe Fe chlorosis of leaves was observed and the growth of the crop was

reduced drastically (Plate 3) but no such symptoms of Fe chlorosis were observed in case of

sandy loam soil (Plate 4). The growth impressions of various combinations of applied P and

Cu levels in calcareous loamy sand (Plate 5) and sandy loam soil (Plate 6) are also exhibited.

4.3.2 Root dry matter yield (RDMY)

The data on dry matter yield of root (RDMY) as influenced by P and Cu application

is presented in Table 4.20. Application of P significantly increased the mean RDMY with

increasing levels of its application over control in both the soils. In loamy sand soil, the

mean RDMY increased significantly from 0.69 g pot-1

in control to a maximum of 1.34 g

pot-1

when P was applied @ 200 mg kg-1

soil. However, in sandy loam soil mean RDMY

increased significantly from 1.27 g pot-1

in control to a maximum 2.09 g pot-1

with the

highest level of applied Cu. In loamy sand soil, RDMY increased by 34.3, 57.9, 79.7, 94.2

and 85.5 per cent whereas in sandy loam soil it increased by 28.3, 26.7, 44.8, 55.1 and 64.5

per cent over control with application of 25, 50, 100, 200 and 400 mg P kg-1

soil,

respectively.

53

Fig. 4.6: Per cent distribution of Cu applied @ 20 mg kg-1

soil in different pools in

loamy sand (a, b) and sandy loam (c, d) soils as influenced by P application

3.87

3.535.52

2.62

32.7

6.5

1.3

42.8

EXCH

SAD

CARB

MnOX

AMPOX

CRYOX

OM

RES

-P -P

(a)

3.152.81 6.19

2.57

32.5

7.22

1.29

43.8

EXCH

SAD

CARB

MnOX

AMPOX

CRYOX

OM

RES

+P

(b)

3.42 1.46

1.08

40.6

16.80.8

35.8

EXCH

SAD

MnOX

AMPOX

CRYOX

OM

RES

-P

(c)

2.6941.446

1.154

33.14

11.93

2.81

47.16

EXCH

SAD

MnOX

AMPOX

CRYOX

OM

RES

+P

(d)

54

Fig. 4.7: Per cent distribution of native Cu in different pools in loamy sand (a, b) and

sandy loam (c, d) soils as influenced by P application

1.091.33

0.83

1.1

7.5

20

25.8

43.5

EXCH

SAD

CARB

MnOX

AMPOX

CRYOX

OM

RES

-P

(a)

0.898

1.2041.602

1.054

8.36

20.04

26.92

41.12

EXCH

SAD

CARB

MnOX

AMPOX

CRYOX

OM

RES

+P

(b)

0.74 0.50.52

18.6

20.1

6.42

52.7

EXCH

SAD

MnOX

AMPOX

CRYOX

OM

RES

(c)

-P

0.61

0.55

0.64

18.36

14.08

8.1

57.3

EXCH

SAD

MnOX

AMPOX

CRYOX

OM

RES

+P

(d)

Plate 1: Growth of wheat as influenced by graded levels of applied P in the absence of applied

copper in loamy sand

Plate 2: Growth of wheat as influenced by graded levels of applied P in the absence of applied

copper in sandy loam

56

Plate 3: Growth of wheat as influenced by graded levels of applied P in combination with 20

mg Cu kg-1

soil in loamy sand

Plate 4: Growth of wheat as influenced by graded levels of applied P in combination with 20

mg Cu kg-1

soil in sandy loam

57

Plate 5: Growth of wheat as influenced by various combinations of applied P and Cu in loamy

sand

58

Plate 6: Growth of wheat as influenced by various combinations of applied P and Cu in sandy loam

55

Table 4.20: Effect of phosphorus and copper application on root dry matter yield of

wheat

Copper

levels

(mg kg-1

soil)

Dry matter yield of root (g pot-1

)

Phosphorus levels (mg kg-1

soil)

0 25 50 100 200 400 Mean

Loamy sand

0 0.79 1.16 1.23 1.52 1.65 1.54 1.31

5 0.71 1.06 1.32 1.46 1.53 1.56 1.27

10 0.67 0.87 1.14 1.36 1.58 1.46 1.18

20 0.58 0.61 0.69 0.64 0.62 0.56 0.62

Mean 0.69 0.92 1.09 1.24 1.34 1.28

CD (5%) P levels 0.109 Cu levels 0.089 P x Cu 0.219

Sandy loam

0 1.36 1.66 1.68 1.75 0.90 2.01 1.73

5 1.31 1.84 1.61 1.81 1.98 2.21 1.79

10 1.27 1.56 1.59 1.87 2.07 2.12 1.75

20 1.13 1.46 1.57 1.93 1.93 2.04 1.67

Mean 1.27 1.63 1.61 1.84 1.97 2.09

CD (5%) P levels 0.106 Cu levels NS P x Cu NS

The quadratic response of relative dry matter yield of root to Olsen P indicated that in

loamy sand (Fig. 4.8a, R2=0.836) and sandy loam (Fig. 4.8b, R

2=0.907) soils, maximum root

yield was produced when soil contained about 90 and 75 mg P kg-1

soil, respectively and

thereafter the yield declined. Phosphorus is known to play a significant role in the root

development of plants (Fageria 2009, Fageria and Moreira 2011, Fageria et al 2013). Singh et

al (2005) observed a significant increase of 121 per cent in dry matter yield of wheat roots

with application of 400 mg P kg-1

soil.

In loamy sand soil, Cu application @ 10 and 20 mg kg-1

soil significantly decreased

the mean RDMY by 9.92 and 52.7 per cent over control. It decreased from 1.31 g pot-1

in

control to 0.61 g pot -1

when Cu was applied @ 20 mg kg-1

soil (Table 4.20). Guan et al

(2011) reported that root biomass of wheat in a Chinese Mollisol having 30% clay increased

only up to a level of 200 mg Cu kg-1

soil and thereafter it decreased up to a level of 800 mg

Cu kg-1

soil.

56

In sandy loam soil, the effect of Cu application on RDMY was not significant. The

quadratic response of relative dry matter yield of root to DTPA-Cu indicated that in loamy

sand (Fig. 4.9a, R2=0.998) the root yield decreased steeply when soil had >2 mg DTPA-Cu

kg-1

soil whereas in sandy loam (Fig. 4.9b, R2=0.953) soil maximum root yield was produced

when soil contained about 2.5 mg DTPA-Cu kg-1

soil and thereafter the yield declined. A

significant negative interaction of P and Cu levels on RDMY was observed in loamy sand soil

at each level of applied P. The RDMY decreased significantly when Cu was applied @ 20 mg

kg-1

soil. In the absence of applied Cu mean RDMY in loamy sand soil was 0.795, 1.16, 1.23,

1.52, 1.65 and 1.54 g pot-1

when P was applied @ 0, 25, 50, 100, 200 and 400 mg kg-1

soil,

respectively which decreased to 0.58, 0.61, 0.69, 0.64, 0.62 and 0.56 g pot-1

when 20 mg Cu

kg-1

soil was applied along with graded levels of P, respectively. This type of negative

interaction of P and Cu levels on RDMY was not observed for medium textured soil. The

results indicated that in light textured soils higher levels of applied Cu may prove toxic for the

growth of roots as more amounts of applied Cu remain in available form as compared to

medium textured soils as indicated by the higher content of DTPA-Cu in loamy sand soil

(Table 4.4). Cu in excess interferes with plant’s capacity to absorb and /or translocate other

nutrients, inhibits root hair growth, root elongation and adversely affects the permeability of

root cell membrane (Fageria 2014, Kopittke and Menizes 2006, Sheldon and Menizes 2005).

4.3.3 Grain yield

The data on the grain yield of wheat as influenced by graded levels of applied P and

Cu is presented in Table 4.21. Growth of the crop was improved with graded levels of

applied P in both the soils (Plate1 and 2). In loamy sand alkaline calcareous soil, the mean

grain yield of wheat increased significantly from 5.77 g pot-1

in control to 11.86, 12.58, 13.82,

14.15 and 14.50 g pot-1

resulting in an increase of 105, 118, 139, 145 and 151 per cent over

control with application of 25, 50, 100, 200 and 400 mg P kg-1

soil, respectively. In sandy

loam soil, grain yield increased significantly from 14.68 g pot-1

in control to 18.19, 18.09,

19.30, 19.76 and 23.09 g pot-1

producing an increase of 24, 23, 31.4, 34.6 and 57.2 per cent

over control with graded levels of applied P, respectively.

The quadratic response of relative grain yield to Olsen P indicated that in loamy sand

(Fig. 4.8c, R2=0.554) and sandy loam (Fig. 4.8d, R

2=0.863) soils, maximum grain yield was

produced when soil contained about 90 and 75 mg P kg-1

soil, respectively and thereafter the

yield declined. Singh et al (2005) observed a significant increase in dry matter yield of root

and shoot of wheat with graded levels of applied P (0-400 mg P kg-1

soil) in a Typic

Haplustept. Prasad et al (1983) observed a significant increase in grain yield of wheat with 60

kg P2O5 ha-1

. In a green house study, Shukla and Singh (1979) observed a significant increase

in grain yield of wheat up to 50 mg P kg-1

soil and thereafter it decreased considerably with

250 mg P kg-1

soil in a P deficient soil (4.2 mg P kg-1

soil). During a two year study, Zhang et

57

al (2012) observed that the wheat grain yield increased with increasing levels of applied P (0-

400 kg ha-1

) in a P deficient soil.

Table 4.21 Effect of phosphorus and copper application on grain yield of wheat

Copper

levels

(mg kg-1

soil)

Grain yield (g pot-1

)

Phosphorus levels (mg kg-1

soil)

0 25 50 100 200 400 Mean

Loamy sand

0 10.23 16.82 16.32 20.66 20.75 20.90 17.61

5 7.02 16.40 16.98 17.06 17.76 18.53 15.62

10 5.58 12.96 15.80 15.89 16.71 17.31 14.04

20 0.25 1.26 1.23 1.66 1.37 1.25 1.17

Mean 5.77 11.86 12.58 13.82 14.15 14.50

CD (5%) P levels 1.09 Cu levels 0.89 P x Cu 2.19

Sandy loam

0 15.23 18.50 19.28 19.30 19.90 21.28 18.91

5 15.06 20.80 18.79 18.93 21.47 26.55 20.26

10 14.56 16.91 17.46 18.95 19.28 23.18 18.39

20 13.86 16.56 16.85 20.04 18.38 21.35 17.84

Mean 14.68 18.19 18.09 19.30 19.76 23.09

CD (5%) P levels 1.07 Cu levels 0.87 P x Cu 2.14

The mean grain yield decreased significantly from 17.61 g pot-1 in control to 15.62,

14.04 and 1.17 g pot-1 thereby representing a decrease of 11.3, 20.2 and 93.3 per cent over

control as Cu was applied @ 5, 10 and 20 mg kg-1 soil, respectively in loamy sand soil which

was deficient in DTPA-Cu (0.18 mg kg-1

soil). But in medium textured soil which was sufficient

in DTPA-Cu (1.05 mg kg-1

soil) the mean grain yield increased significantly from 18.91 g pot-1

in control to 20.26 g pot-1

resulting in an increase of 7.31 per cent over control when 5 mg Cu

kg-1 soil was applied but thereafter with application of 10 and 20 mg Cu kg

-1 soil the grain yield

decreased significantly by 2.74 and 5.65 per cent over control and by 9.2 and 11.9 per cent over

application of 5 mg kg-1 soil, respectively. Arshad et al (2011) reported a significant increase of

35.2 per cent in grain plus straw yield of wheat with 4 mg Cu kg-1

soil in three soils of variable

58

texture. The authors further observed that biomass production increased as the clay content in

soil increased. The quadratic response of relative grain yield to DTPA-Cu indicated that in

loamy sand (Fig. 4.9c, R2=0.993) the grain yield decreased steeply when soil had >2 mg DTPA-

Cu kg-1

soil whereas in sandy loam (Fig. 4.9d, R2=0.621) soil grain yield was produced when

soil contained about 2.5 mg DTPA-Cu kg-1

soil and thereafter the yield declined. Shukla and

Singh (1979) observed that a maximum grain yield of wheat was produced when 50 mg P was

combined with 5 mg Cu kg-1

soil but grain yield declined with application of 10, 20 and 50 mg

Cu kg-1

soil as Cu sulphate. Morteza et al (1991) reported a decline in grain yield of wheat with

application of 33.6 and 67.3 kg Cu ha-1

over no Cu application. Sharma and Singh (2012)

observed a significant increase in wheat grain yield only up to a level of 5 kg Cu ha-1

and

thereafter the yield declined at 10 kg Cu ha-1. Similarly, Chhibba et al (1994) also observed a

significant increase in wheat grain yield up to al level of 5 mg Cu kg-1 soil.

The growth of the crop in loamy sand soil was drastically reduced as compared to

control when 20 mg Cu kg-1

soil was applied. Severe deficiency symptoms of Fe chlorosis

were observed at each level of applied P in combination with 20 mg Cu kg-1

soil (Plate 3

and 5). Chhibba et al (1994) observed the symptoms of Fe chlorosis in wheat leaves when

40 mg Cu kg-1

soil was applied in a Typic Ustipsamment. Interestingly, in the absence of

applied Cu no symptoms of Fe deficiency were observed even with the highest level of

applied P (400 mg kg-1

soil, Plate 1).The results clearly indicated that Fe chlorosis was

observed due to excessive Cu application and not due to P application even though the soil

was deficient in available Fe (DTPA-Fe 3.24 kg-1

soil). The antagonism between P and Fe is

well documented but in present study it was not observed so for as the Fe chlorosis was

concerned. Brown et al (1955) also reported that application of P up to a level of 800 kg P

ha-1

did not influence the absorption or utilization of Fe to the extent that could develop

chlorosis unless Cu was also present in the growth medium. Copper and P were more

effective in producing Fe chlorosis if applied together compared to when applied separately.

In a pot experiment with rice, Brown et al (1954) observed that rice yield decreased with

increasing levels of P (0-720 mg kg-1

soil) and Cu (0-180 mg kg-1

soil) and caused the

plants to develop chlorosis. However, the crop growth was good at all levels of applied P in

the absence of applied Cu. Michaud et al (2007) also observed Fe deficiency symptoms in

durum wheat cultivated in Cu contaminated former vineyard soils. They further reported

that the risks of Cu phytotoxicity to wheat (Triticum durum) in situ might be greater in

calcareous soils due to interaction with Fe nutrition. Similar were the observations of

Chaignon et al (2011).

59

Fig. 4.8: Relationship of Olsen P in soil at harvest with relative root, grain and straw yield

y = -0.007x2 + 1.272x + 53.32R² = 0.836

50

60

70

80

90

100

110

0 25 50 75 100 125 150

Re

lati

ve r

oo

t d

ry m

atte

r yi

eld

(%

)

Olsen P (mg kg-1 soil)

y = -0.01x2 + 1.48x + 49.16R² = 0.907

50

60

70

80

90

100

110

0 25 50 75 100

Re

lati

ve r

oo

t d

ry m

atte

r yi

eld

(%

)

Olsen P (mg kg-1 soil)

y = -0.006x2 + 1.195x + 56.08R² = 0.554

30

40

50

60

70

80

90

100

110

0 25 50 75 100 125 150

Re

lati

ve g

rain

yie

ld (

%)

Olsen P (mg kg-1 soil)

y = -0.004x2 + 0.788x + 60.69R² = 0.863

50

60

70

80

90

100

0 25 50 75 100

Re

lati

ve g

rain

yie

ld (

%)

Olsen P (mg kg-1 soil)

y = -0.007x2 + 1.313x + 53.46R² = 0.662

50

60

70

80

90

100

110

120

0 25 50 75 100 125 150

Re

lati

ve s

traw

yie

ld (

%)

Olsen P (mg kg-1 soil)

y = -0.006x2 + 1.062x + 55.63R² = 0.905

50

60

70

80

90

100

110

0 25 50 75 100

Re

lati

ve s

traw

yie

ld (

%)

Olsen P (mg kg-1 soil)

(a) (b)

(c) (d)

(e) (f)

60

Fig. 4.9: Relationship of DTPA-Cu in soil at harvest with relative root, grain and straw yield

y = -1.577x2 + 2.737x + 99.02R² = 0.998

40

50

60

70

80

90

100

110

0 1 2 3 4 5 6 7

Re

lati

ve r

oo

t d

ry m

atte

r yi

eld

(%

)

DTPA-Cu (mg kg-1 soil)

y = -0.561x2 + 3.333x + 94.37R² = 0.953

90

95

100

0.5 1.5 2.5 3.5 4.5 5.5 6.5Re

alti

ve r

oo

t d

ry m

atte

r yi

eld

(%

)

DTPA-Cu (mg kg-1 soil)

y = -2.344x2 + 2.108x + 98.07R² = 0.993

0

20

40

60

80

100

0 1 2 3 4 5 6 7

Re

lati

ve g

rain

yie

ld (

%)

DTPA-Cu (mg kg-1 soil)

y = -0.632x2 + 3.163x + 92.23R² = 0.621

85

90

95

100

0.5 1.5 2.5 3.5 4.5 5.5 6.5

Re

lati

ve g

rain

yie

ld (

%)

DTPA-Cu (mg kg-1 soil)

y = -2.286x2 + 2.772x + 98.28R² = 0.995

0

20

40

60

80

100

0 1 2 3 4 5 6 7

Re

lati

ve s

traw

yie

ld (

%)

DTPA-Cu (mg kg-1 soil)

y = -0.500x2 + 1.960x + 92.31R² = 0.424

75

80

85

90

95

100

0.5 1.5 2.5 3.5 4.5 5.5 6.5

Re

lati

ve s

traw

yie

ld (

%)

DTPA-Cu (mg kg-1 soil)

(a) (b)

(c) (d)

(e) (f)

61

Though the antagonistic effect of higher rates of Cu application on grain yield of wheat

was also observed in sandy loam soil but no Fe deficiency symptoms were observed (Plate 4 and 6

) because this soil contained very high amounts of available Fe (DTPA-Fe 50.2 mg kg-1 soil). The

loamy sand alkaline calcareous soil was deficient in available Cu (DTPA-Cu 0.18 mg kg-1 soil),

even then no significant response of the crop to even the lowest level of applied Cu (5 mg kg-1

soil) was observed. Had Cu been applied at some rates lower than 5 mg kg-1 soil response of

wheat to its application could have been observed. A significant increase of 63 per cent in grain

yield of wheat was observed by Kumar et al (2009) with 1.5 mg Cu kg-1 soil.

A significant negative interaction effect of P and Cu levels on grain yield of wheat in

this soil was observed and the grain yield decreased significantly with increasing levels of

applied Cu at every level of applied P. The results indicated that in light textured soils the Cu

requirements of wheat will be much lower than that in heavy textured soils because in light

textured soils more amounts of applied Cu will remain in solution form (Table 4.5.) which

may prove toxic to the plants either directly or indirectly by effecting the availability and the

absorption of Fe from root to the above ground portion.

A significant interaction effect of P and Cu levels on grain yield of wheat also observed

for medium textured sandy loam soil. In the absence of applied P, mean grain yield decreased

over control at any level of applied Cu but this decrease was not significant. However, when P

was applied @ 25 mg kg-1

soil, the grain yield increased significantly from 18.50 g pot-1 in

control to 20.8 g pot-1 when Cu was also applied @ 5 mg kg

-1 soil but thereafter the grain yield

decreased significantly to 16.91 and 16.51 g pot-1

when Cu was applied @ 10 and 20 mg kg-1

soil, respectively. But when P was applied @ 50, 100 and 200 mg kg-1 soil the grain yield either

decreased or increased with Cu application but this decrease or increase was not significant over

control. However, when P was applied @ 400 mg kg-1

soil, the mean grain yield increased

significantly from 21.28 g pot-1

in control to 26.55 g pot-1 when Cu was also applied @ 5 mg kg

-

1 soil. But the yield decreased significantly to 23.18 and 21.35 g pot

-1 with further application of

Cu @ 10 and 20 mg kg-1

soil, respectively. However, this decrease in yield with application of

20 mg Cu kg-1

soil was not significant over control. The results showed that high levels of Cu in

soils might be interfering with Fe nutrition of the crops but the adverse effect is more

pronounced if the soil is light textured and deficient in available Fe.

4.3.4 Chlorophyll content of leaves

The data on SPAD values of the leaves an indicator of chlorophyll content at 65 days

after sowing is presented in Table 4.22. Mean values of SPAD in loamy sand soil decreased

by 3.74, 18.4 and 63.5 per cent over control with increasing levels of applied Cu. Its value

significantly decreased from 45.8 in control to 42.8, 35.8 and 17.6 with application of 5, 10

and 20 mg Cu kg-1

soil, respectively.

62

Severe deficiency symptoms of Fe chlorosis were also observed at each level of

applied P when 20 mg Cu kg-1

soil was applied (Plate 3 and 5), thereby resulting in a

significant decrease in root (Table 4.20), grain (Table 4.21) and straw (Table 4.24) yield.

DTPA-Cu in soil at harvest was negatively correlated with SPAD values in loamy sand (Fig.

4.10a, R2= 0.957) as well as sandy loam soil (Fig. 4.10b, R

2=0.990). A significant interaction

effect of P and Cu levels on SPAD values in loamy sand soil revealed that chlorophyll content

of leaves deceased significantly with Cu application @ 10 and 20 mg Cu kg-1

soil at each

level of applied P. Sahu et al (1988) also observed severe Fe chlorosis and a significant

reduction in chlorophyll content of garden pea leaves with P and Cu application. In sandy

loam soil (DTPA- Fe 50.2 mg kg-1

soil) the effect of P and Cu levels and their interaction on

chlorophyll content was not significant, crop growth was normal and no symptoms of Fe

deficiency were observed. In sandy loam, the SPAD values decreased non significantly by

2.45, 3.71 and 5.9 per cent over control with increasing levels of applied Cu, respectively.

The results indicated that if soil is deficient in available Fe, high levels of applied Cu may

lead to Fe chlorosis.

Table 4.22: Effect of phosphorus and copper application on SPAD values at 65 days

after sowing

Copper

levels

(mg kg-1

soil)

SPAD values

Phosphorus levels (mg kg-1

soil)

0 25 50 100 200 400 Mean

Loamy sand

0 43.10 47.50 47.90 42.80 47.10 46.30 45.80

5 39.90 42.60 43.20 40.20 43.50 47.60 42.80

10 33.60 34.50 41.30 38.20 32.60 35.00 35.80

20 19.00 14.40 26.80 15.30 15.90 14.40 17.60

Mean 33.90 34.70 39.80 34.10 34.80 35.80

CD (5%) P levels 2.76 Cu levels 2.25 P x Cu 5.53

Sandy loam

0 46.96 48.13 51.70 49.83 49.06 49.83 49.25

5 46.16 48.30 48.83 49.53 48.63 46.80 48.04

10 44.86 48.90 48.33 49.60 46.60 46.23 47.42

20 42.86 48.13 46.90 45.80 45.26 49.10 46.34

Mean 45.21 48.36 48.94 48.69 47.39 47.99

CD (5%) P levels NS Cu levels NS P x Cu NS

63

Fig. 4.10: Relationship of DTPA-Cu in soil at harvest with relative chlorophyll content

in leaves at 65 days after sowing

4.3.5 Specific activity of superoxide dismutase in leaves

The data on the specific activity of total superoxide dismutase (SOD) in leaves at 65

days growth for selected treatments is presented in Table 4.23. Mean specific activity of SOD

significantly increased (87 per cent) from 3.72 units min-1

mg-1

protein to 6.98 units min-1

mg-

1 protein as the level of applied P increased from 25 mg P kg

-1 soil to 400 mg P kg

-1 soil.

Similarly, the specific activity of SOD increased (84 per cent) significantly from 3.68 units

min-1

mg-1

protein in no Cu treatment to 6.80 units min-1

mg-1

protein when 20 mg Cu kg-1

soil

was applied, the level at which the severe chlorosis of leaves was observed (Plate 3 and 5). In

many studies it has been confirmed that excess Cu could promote and stimulate the generation

of reactive oxygen species (ROS) leading to an increase in the activities of antioxidant

enzymes as a defecse system (Miller et al 2004; Fariduddin et al 2009; Verma et al 2011).

The results of present study are in harmony with these reported findings because application

of Cu @ 20 mg kg-1

soil caused a significant increase in activity of SOD which can be

considered as a circumstantial evidence of enhanced ROS production. The higher activity of

SOD at higher concentrations of Cu gives an indication of ability of wheat to cope with ROS.

Thus, it may be concluded that the increase recorded in the activities of SOD may be

attributed to the adaptive defence system of wheat plants against the toxic effects of Cu.

Under green house conditions, Azooz et al (2012) also observed that when Cu was applied in

excess of 10 mM up to100 mM, the activity of SOD along with catalase, peroxidase and

ascorbate peroxidase increased in wheat at 30 days growth. An increase in SOD activity in

leaves of Jatropha (Jatropha curcas L.) seedlings under Cu stress up to 400 µmol was

reported by Gao et al (2008) in a pot experiment. Similarly, Karimi et al (2012) observed that

during the exposure of Astagalus neo-mobayenii plants to excess Cu helped in increasing the

SOD activity compared to control.

y = -4.488x + 49.39R² = 0.957

10

15

20

25

30

35

40

45

50

55

0 2 4 6 8

Spad

val

ue

s

DTPA-Cu (mg kg-1 soil)

y = -0.526x + 49.61R² = 0.990

46

46.5

47

47.5

48

48.5

49

49.5

0 2 4 6 8

Spad

val

ue

s

DTPA-Cu (mg kg-1 soil)

(a) (b)

64

The interaction effect of P and Cu levels on specific activity of SOD indicated that it

increased by 92 and 135 per cent over no Cu when 5 and 20 mg Cu kg-1

soil was applied in

combination with 400 mg P kg-1

soil, respectively. Further, at any level of applied Cu the

specific activity of SOD was more pronounced when combined with application of 400 mg P

kg-1

soil. The specific activity of SOD increased by 119 and 117 per cent when 5 and 20 mg

Cu kg-1

soil was applied together with 400 mg P kg-1

soil, respectively.

Table 4.23: Specific activity of total super oxide dismutase in leaves at 65 days after

sowing as influenced by P and Cu application in selected treatments

Copper levels (mg kg-1

soil)

Specific activity of total Super Oxide Dismutase (units min-1

mg-1

protein)

Phosphorus levels (mg kg-1

soil)

0 25 400 Mean

5 3.39 3.97 3.68

20 3.48 7.64 5.56

Mean 4.28 9.33 6.80

CD (5%) P levels 1.23 Cu levels 1.50 P x Cu 2.13

4.3.6 Direct and indirect effect of various pools of copper towards grain yield of wheat

Path coefficient analysis of grain yield of wheat against various pools of Cu revealed

that in loamy sand soil DTPA-Cu, CARB-Cu and AMPOX-Cu produced significantly direct

effects on grain yield of wheat (Table 4.24). About 21.9, 45.5 and 18.6 percent variation in

grain yield was directly controlled by DTPA-Cu, CARB-Cu and AMPOX-Cu, respectively.

Also, DTPA-Cu controlled a variation of 40.8 and 17.3 per cent in grain yield indirectly

through CARB-Cu and AMPOX-Cu, respectively. Exchangeable Cu and SAD-Cu, the most

plant available pools controlled a variation of 17.8 and 17.0 per cent in grain yield indirectly

through DTPA-Cu. An indirect variation of 39.8 and 13.3 per cent, in grain yield was

controlled by EXCH-Cu through CARB-Cu and AMPOX-Cu and 37.9 and 12.7 per cent by

SAD-Cu through CARB-Cu and AMPOX-Cu, respectively. The results indicated that

importance of DTPA-Cu, EXCH-Cu, SAD-Cu and AMPOX-Cu in controlling the grain yield

of wheat in loamy sand calcareous soils.

However, in sandy loam nearly acidic soil only CRYOX-Cu produced significantly

positive effect on gain yield and controlled a direct variation of 76.2 per cent in grain yield

(Table 4.24). Organically bound and residual Cu also controlled a direct variation of 1.4 and

11.6 per cent of grain yield. An indirect variation of 59.1, 42.1 and 10.0 per cent by DTPA-

Cu, 63.4, 42.5 and 9.1 per cent by EXCH-Cu, 62.7, 40.4 and 9.3 per cent by SAD-Cu, 50.8,

60.6 and 9.8 per cent by MnOX-Cu, 62.2, 40.5 and 9.3 per cent by AMPOX-Cu was

65

controlled through CRYOX-Cu, OM-Cu and RES-Cu, respectively. Thus, in sandy loam

acidic soil oxides and organic matter forms of Cu played an important role either directly or

indirectly in effecting the grain yield.

Table 4.24: Direct and indirect effect of various pools of copper towards grain yield of

wheat

Loamy sand

DTPA-

Cu

EXCH-

Cu

SAD-

Cu

CARB-

Cu

MnOX-

Cu

AMPOX-

Cu

CRYOX-

Cu

OM-

Cu

RES-

Cu

DTPA-Cu 0.468 -0.574 0.166 0.639 -0.362 0.417 -0.154 -0.103 -1.31

EXCH-Cu 0.422 -0.635 0.182 0.631 -0.392 0.366 -0.144 -0.115 -1.12

SAD-Cu 0.413 -0.616 0.188 0.616 -0.406 0.357 -0.139 -0.123 -1.14

CARB-Cu 0.442 -0.594 0.172 0.675 -0.390 0.394 -0.145 -0.111 -1.25

MnOX-Cu 0.396 -0.582 0.179 0.616 -0.427 0.357 -0.140 -0.126 -1.08

AMPOX-Cu 0.451 -0.538 0.155 0.616 -0.353 0.432 -0.156 -0.096 -1.27

CRYST-Cu 0.416 -0.529 0.151 0.563 -0.344 0.389 -0.174 -0.098 -1.15

OM-Cu 0.306 -0.465 0.148 0.478 -0.343 0.265 -0.108 -0.157 -0.794

RES-Cu 0.458 -0.570 0.159 0.630 -0.343 0.408 -0.149 -0.092 -1.34

Sandy loam

DTPA-Cu -0.718 -0.282 -0.487 - -0.207 -0.312 0.769 0.649 0.320

EXCH-Cu -0.619 -0.327 -0.480 - -0.205 -0.290 0.731 0.652 0.303

SAD-Cu -0.674 -0.303 -0.517 - -0.210 -0.307 0.792 0.636 0.305

MnOX-Cu -0.651 -0.294 -0.476 - -0.228 -0.288 0.713 0.779 0.314

AMPOX-Cu -0.698 -0.295 -0.494 - -0.205 -0.321 0.789 0.637 0.305

CRYOX-Cu -0.632 -0.273 -0.469 - -0.186 -0.290 0.873 0.335 0.273

OM-Cu -0.392 -0.179 -0.276 - -0.149 -0.172 0.246 0.118 0.211

RES-Cu -0.673 -0.290 -0.462 - -0.210 -0.288 0.700 0.736 0.341

Bold figures represent direct effects of chemical pools

4.3.7 Straw yield

The data on the effect of P and Cu application on straw yield is presented in Table

4.25. In both the soils straw yield increased significantly over control with graded levels of

applied P. In loamy sand soil, straw yield increased from 8.17 g pot-1

in control to a maximum

of 19.40 g pot-1

and in medium textured soil straw yield increased from 20.14 g pot-1

in

control to a maximum of 32.32 g pot-1

with the highest level of 400 mg P kg-1

soil. A

significant increase of 86.4, 95.3, 130.4, 137.0 and 137.4 per cent in loamy sand and 23.9,

25.7, 35.1, 42.5, and 60.4 per cent in sandy loam soil over control was observed with

application of 25, 50, 100, 200 and 400 mg P kg-1

soil, respectively. The quadratic response of

relative straw yield to Olsen P indicated that in loamy sand (Fig. 4.8e, R2=0.662) and sandy

66

loam (Fig. 4.8f, R2=0.905) soils, maximum straw yield was produced when soil contained

about 90 and 75 mg P kg-1

soil, respectively and thereafter the yield declined. Shukla and

Singh (1979) observed a significant increase in straw yield of wheat up to 50 mg P kg-1

soil.

Similar were the findings of Singh and Swarup (1982).

Table 4.25: Effect of phosphorus and copper application on straw yield of wheat

Copper

levels

(mg kg-1

soil)

Straw yield (g pot-1

)

Phosphorus levels (mg kg-1

soil)

0 25 50 100 200 400 Mean

Loamy sand

0 13.66 20.17 19.90 24.66 27.94 28.49 22.47

5 9.77 20.93 21.65 23.08 23.82 23.98 20.54

10 8.04 16.82 19.44 23.01 21.78 21.89 18.49

20 1.23 3.02 2.87 4.59 3.94 3.25 3.15

Mean 8.17 15.23 15.96 18.83 19.37 19.40

CD (5%) P levels 1.54 Cu levels 1.26 P x Cu 3.09

Sandy loam

0 23.06 24.59 26.61 27.38 29.00 29.32 26.66

5 20.30 30.51 27.41 26.06 32.26 37.75 29.05

10 18.80 22.08 23.75 26.78 27.58 31.06 25.01

20 18.41 22.68 23.53 28.67 26.03 31.15 25.08

Mean 20.14 24.96 25.32 27.22 28.71 32.32

CD (5%) P levels 1.78 Cu levels 1.45 P x Cu 3.56

Copper application significantly decreased the straw yield from 22.47 g pot-1

in

control to 20.54, 18.49 and 3.15 g pot-1

thus representing a decrease of 8.58, 17.7 and 85.9 per

cent over control with application of 5, 10 and 20 mg Cu kg-1

soil, respectively in light

textured soil. Application of 5 mg Cu kg-1

soil significantly increased the mean straw yield

from 26.66 g pot-1

in control to 29.05 g pot-1

in sandy loam soil but Cu application @ 10 and

20 mg kg-1

soil significantly decreased the straw yield by 6.18 and 5.92 per cent over control,

and by 13.9 and 13.6 per cent over the application of 5 mg Cu kg-1

soil, respectively. The

quadratic response of relative straw yield to DTPA-Cu indicated that in loamy sand (Fig. 4.9e,

R2=0.995) the straw yield decreased steeply when soil had >2 mg DTPA-Cu kg

-1 soil whereas

in sandy loam (Fig. 4.9f, R2=0.424) soil maximum straw yield was produced when soil

contained about 2.5 mg DTPA-Cu kg-1

soil and thereafter the yield declined. Under

greenhouse conditions in a sandy loam soil, Gupta and MacLeod (1970) observed a

significant increase in grain and straw yield of wheat, barley and oats to an application of 0.5

mg Cu kg-1

soil.

67

The data on linear coefficients of correlation of chemical pools of Cu with yield of

root, grain and straw revealed that in loamy sand calcareous soil, root dry matter yield, grain

and straw yield was significantly negatively correlated with the various pools of Cu including

DTPA-Cu (Table 4.26). However, in sandy loam only organically bound Cu was significantly

positively correlated with the root (r=0.470*) and grain (r=0.476

*) yield. Also DTPA-Zn, Fe

and Mn were significantly positively correlated with root, grain and straw yield.

Table 4.26: Linear coefficients of correlation of root, grain and straw yield of wheat

with various pools of copper (n=24)

Root dry matter yield Grain yield Straw yield

Loamy sand soil

EXCH-Cu -0.761**

-0.895**

-0.899**

SAD-Cu -0.696**

-0.852**

-0.862**

CARB-Cu -0.628**

-0.816**

-0.802**

MnOX-Cu -0.668**

-0.812**

-0.817**

AMPOX-Cu -0.609**

-0.762**

-0.732**

CRYOX-Cu -0.693**

-0.780**

-0.783**

OM-Cu -0.559**

-0.671**

-0.766**

RES-Cu -0.687**

-0.848**

-0.825**

DTPA-Cu -0.662**

-0.823**

-0.809**

DTPA-Zn 0.333 0.159 0.196

DTPA-Mn 0.326 0.150 0.188

DTPA-Fe 0.339 0.166 0.204

Sandy loam soil

EXCH-Cu -0.180 -0.236 -0.311

SAD-Cu -0.193 -0.279 -0.373

MnOX-Cu 0.002 -0.132 -0.222

AMPOX-Cu -0.210 -0.282 -0.405**

CRYOX-Cu -0.280 -0.370 -0.403

OM-Cu 0.470**

0.476**

0.327

RES-Cu 0.012 -0.146 -0.257

DTPA-Cu -0.155 -0.268 -0.374

DTPA-Zn 0.766**

0.759**

0.693**

DTPA-Mn 0.769**

0.767**

0.687**

DTPA-Fe 0.771**

0.763**

0.698**

*Significant at 5%,

**Significant at 1%

68

A significant interaction effect of P and Cu application on straw yield was observed

in both the soils. In light textured soil, in the absence of applied P, straw yield decreased

significantly over control at each level of Cu application. Also at each level of applied P, the

straw yield decreased significantly over control when Cu was applied @ 20 mg kg-1

soil.

However, in case of sandy loam soil, a significant decrease in straw yield over control

with application of 10 and 20 mg Cu kg-1

soil was observed only up to a level of 50 mg P kg-1

soil. Thereafter, the increase or decrease in straw yield with Cu was not significant at any

level of applied P except that a maximum straw yield of 37.75 g pot-1

was produced when 5

mg Cu kg-1

soil was applied along with 400 mg P kg-1

soil.

4.4 Effect of phosphorus and copper application on concentration of P, Cu and Fe in

root, grain and straw

4.4.1 Phosphorus concentration in root

In loamy sand soil, P concentration in root increased significantly over control with

graded levels of applied P. It increased from 0.031% in control to a maximum of 0.1% which

represented an increase of 222 per cent over control with application of 400 mg P kg-1

soil

(Table 4.27). But in sandy loam soil a significant increase of 24.09 and 28.91per cent over

control was observed only when P was applied @ 200 and 400 mg P kg-1

soil, respectively. It

increased from 0.084% in control to 0.108% with application of 400 mg P kg-1

soil. Mamo

and Parsons (1987) observed an increase of about 33 per cent P in roots of teff plants with

application of 500 mg P kg-1

soil. A significant increase of 57.5 per cent in root P

concentration of wheat with application of 400 mg P kg-1

soil was reported by Singh et al

(2005).

The quadratic response of root P concentration to Olsen P indicated that in loamy

sand (Fig. 4.11a, R2=0.885) and sandy loam (Fig. 4.11b, R

2=0.887) soils, maximum root P

concentration was observed when soil contained about 90 and 90 mg P kg-1

soil, respectively

and thereafter it declined. A significant positive coefficient of correlation of dry matter yield

of root with root P concentration in loamy sand (r=0.597**

) and sandy loam (r=0.622**

) soil

was observed (Table 4.34).

In light textured soil, Cu application significantly increased root P concentration from

0.070% in control to 0.082% with application of 10 mg Cu kg-1

soil but it decreased to

0.075% with application of 20 mg Cu kg-1

soil over the application of 10 mg Cu kg-1

soil.

Chhibba et al (1994) observed a significant decrease in P concentration of flag leaves of

wheat with Cu application. Similar were the findings of Adriano et al (1971) for maize

seedlings. In a near neutral soil (pH 6.9), Morteza et al (1991) observed that P concentration

in wheat leaves increased from 0.37 per cent in control (P0Cu0) to 0.50 per cent with

combined application of 3360 kg P ha-1

as mono-calcium phosphate and 67.3 kg Cu ha-1

as

Cu sulphate. They further observed that at any level of applied Cu, concentration of P in

69

wheat leaf increased as the Cu application increased from 0 to 67.3 kg ha-1

.The effect of Cu

application on root concentration in sandy loam soil was not significant. The quadratic

response of root P concentration to DTPA-Cu indicated that in loamy sand (Fig. 4.12a,

R2=0.999) root P concentration decreased steeply when soil had >3.6 mg DTPA-Cu kg

-1 soil

whereas in sandy loam (Fig. 4.12b, R2=0.857) root P concentration decreased linearly with

increase in DTPA-Cu.

Table 4.27: Effect of phosphorus and copper application on phosphorus concentration

in root

Copper

levels

(mg kg-1

soil)

Root P concentration (%)

Phosphorus levels (mg kg-1

soil)

0 25 50 100 200 400 Mean

Loamy sand

0 0.030 0.034 0.058 0.084 0.107 0.107 0.070

5 0.033 0.046 0.085 0.084 0.103 0.120 0.079

10 0.030 0.055 0.081 0.108 0.121 0.097 0.082

20 0.030 0.055 0.078 0.102 0.105 0.077 0.075

Mean 0.031 0.048 0.076 0.095 0.109 0.100

CD (5%) P levels 0.006 Cu levels 0.005 P x Cu NS

Sandy loam

0 0.076 0.088 0.086 0.077 0.124 0.110 0.093

5 0.091 0.078 0.088 0.082 0.105 0.105 0.091

10 0.087 0.086 0.083 0.090 0.096 0.105 0.091

20 0.080 0.085 0.082 0.093 0.089 0.111 0.090

Mean 0.083 0.084 0.085 0.085 0.103 0.107

CD (5%) P levels 0.006 Cu levels NS P x Cu NS

4.4.2 Phosphorus concentration in grain

The data on effect of P and Cu levels on P concentration in wheat grain is presented

in Table 4.28. In loamy sand, grain P concentration increased significantly from 0.074 per

cent in control to 0.109, 0.126, 0.157 and 0.148 per cent which represented an increase of

12.1, 47.2, 70.2, 112 and 100 per cent over control when P was applied @ 50, 100, 200 and

400 mg kg-1

soil, respectively. The decrease in grain P concentration observed with

application of 400 mg P kg-1

soil over that of 200 mg P kg-1

soil was not significant. In sandy

loam soil, mean grain P concentration increased significantly from 0.134 per cent in control to

0.151, 0.154, 0.156, 0.181 and 0.213 per cent resulting in an increase of 12.6, 14.1, 16.4, 35.0

70

and 58.9 per cent over control with application of 25, 50, 100, 200 and 400 mg P kg-1

soil,

respectively. The differences in grain P concentration observed with application of 25, 50 and

100 mg P kg-1

soil were not significant. A positive coefficient of correlation of grain yield

with grain P concentration in loamy sand (r=0.388) and sandy loam (r=0.792**

) soil was

observed (Table 4.34). Shukla and Singh (1979) observed a significant increase in P

concentration in grain and straw of wheat with increasing levels of applied P (0-250 mg kg-1

soil).

The quadratic response of gain P concentration to Olsen P indicated that in loamy

sand (Fig. 4.11c, R2=0.970) P concentration in grain increased to the maximum up to a level

of 90 mg P kg-1

soil and thereafter it declined. In sandy loam (Fig. 4.11d, R2=0.978) grain P

concentration increased with increasing Olsen P.

Table 4.28: Effect of phosphorus and copper application on phosphorus concentration

in grain

Copper

levels

(mg kg-1

soil)

Grain P concentration (%)

Phosphorus levels (mg kg-1

soil)

0 25 50 100 200 400 Mean

Loamy sand

0 0.068 0.069 0.106 0.130 0.145 0.180 0.116

5 0.078 0.084 0.096 0.128 0.160 0.135 0.114

10 0.072 0.088 0.114 0.125 0.143 0.127 0.112

20 0.078 0.091 0.118 0.120 0.180 0.150 0.123

Mean 0.074 0.083 0.109 0.126 0.157 0.148

CD (5%) P levels 0.011 Cu levels NS P x Cu0.022

Sandy loam

0 0.128 0.144 0.154 0.156 0.199 0.211 0.165

5 0.135 0.161 0.163 0.156 0.173 0.218 0.168

10 0.142 0.149 0.139 0.143 0.166 0.196 0.156

20 0.139 0.150 0.157 0.169 0.186 0.229 0.172

Mean 0.134 0.151 0.153 0.156 0.181 0.213

CD (5%) P levels 0.013 Cu levels NS P x Cu NS

The effect of Cu levels on grain P concentration was not significant in both the soils.

In loamy sand soil a significant interaction effect of P and Cu levels on grain P concentration

indicated that at each level of applied Cu, the grain P concentration increased significantly

with application of 50, 100, 200 and 400 mg P kg-1

soil over control. The quadratic response

71

of gain P concentration to DTPA-Cu indicated that in loamy sand (Fig. 4.12c, R2=0.963) P

concentration in grain decreased up to a level of 3 mg DTPA-Cu kg-1

soil and thereafter it

followed an increasing trend. In sandy loam (Fig. 4.12d, R2=0.980) grain P concentration

increased with increasing DTPA-Cu.

4.4.3 Phosphorus concentration in straw

Concentration of P in straw increased over control with increasing levels of P

application (Table 4.29). In loamy sand soil, P concentration in straw increased significantly

from 0.050 per cent in control to 0.062, 0.085, 0.102 and 0.104 per cent which represented an

increase of 24.0, 70.0, 104 and 108 per cent over control when P was applied @ 50, 100, 200

and 400 mg kg-1

soil, respectively. In sandy loam soil, it increased significantly from 0.069

per cent in control to 0.084, 0.083, 0.104 and 0.146 percent thereby resulting in an increase of

21.7, 20.2, 50.7 and 111 per cent over control with application of 50, 100, 200 and 400 mg P

kg-1

soil, respectively. A positive coefficient of correlation of straw yield with straw P

concentration in loamy sand (r=0.264) and sandy loam (r=0.716**

) soil was observed (Table

4.34). Shukla and Singh (1979) observed that P concentration in grain and straw of wheat

increased with Cu application up to 5 mg kg-1

soil but then decreased up to 50 mg Cu kg-1

soil.

Table 4.29: Effect of phosphorus and copper application on phosphorus concentration

in straw

Copper

levels

(mg kg-1

soil)

Straw P Concentration (%)

Phosphorus levels (mg kg-1

soil)

0 25 50 100 200 400 Mean

Loamy sand

0 0.037 0.034 0.054 0.065 0.106 0.128 0.071

5 0.039 0.047 0.071 0.087 0.084 0.114 0.074

10 0.050 0.051 0.067 0.085 0.117 0.096 0.078

20 0.074 0.067 0.056 0.103 0.102 0.079 0.080

Mean 0.050 0.050 0.062 0.085 0.102 0.104

CD (5%) P levels 0.012 Cu levels NS P x Cu NS

Sandy loam

0 0.061 0.070 0.077 0.079 0.101 0.144 0.088

5 0.063 0.072 0.071 0.080 0.107 0.149 0.091

10 0.088 0.074 0.103 0.086 0.105 0.151 0.101

20 0.062 0.065 0.085 0.088 0.102 0.141 0.091

Mean 0.069 0.071 0.084 0.083 0.104 0.146

CD (5%) P levels 0.007 Cu levels NS P x Cu NS

72

Fig. 4.11: Relationship of Olsen P in soil at harvest with P concentration in root (a, b),

grain (c, d) and straw (e, f)

y = -1E-05x2 + 0.002x + 0.027R² = 0.885

0.00

0.02

0.04

0.06

0.08

0.10

0.12

0.14

0 25 50 75 100 125 150

Ro

ot

P c

on

cen

trat

ion

(%

)

Olsen P (mg kg-1 soil)

y = -4E-06x2 + 0.000x + 0.071R² = 0.887

0.07

0.08

0.09

0.10

0.11

0 25 50 75 100

Ro

ot

P c

on

cen

trat

ion

(%

)

Olsen P (mg kg-1 soil)

y = -1E-05x2 + 0.002x + 0.062R² = 0.970

0.06

0.08

0.10

0.12

0.14

0.16

0.18

0 25 50 75 100 125 150

Gra

in P

co

nce

ntr

atio

n (

%)

Olsen P (mg kg-1 soil)

y = -6E-06x2 + 0.001x + 0.119R² = 0.978

0.10

0.15

0.20

0.25

0 25 50 75 100

Gra

in P

co

nce

nta

tio

n (

%)

Olsen P (mg kg-1 soil)

y = -8E-06x2 + 0.001x + 0.037R² = 0.985

0.04

0.06

0.08

0.10

0.12

0 25 50 75 100 125 150

Stra

w P

co

nce

ntr

atio

n s

traw

(%

)

Olsen P (mg kg-1 soil)

y = 0.000x + 0.057R² = 0.991

0.05

0.07

0.09

0.11

0.13

0.15

0.17

0 25 50 75 100

Stra

w P

co

nce

ntr

atio

n in

(%

)

Olsen P (mg kg-1 soil)

(a) (b)

(c) (d)

(e) (f)

73

Fig. 4.12: Relationship of DTPA-Cu in soil at harvest with P concentration in root (a, b),

grain (c, d) and straw (e, f)

y = -0.000x2 + 0.007x + 0.068R² = 0.999

0.068

0.070

0.072

0.074

0.076

0.078

0.080

0.082

0.084

0 1 2 3 4 5 6 7

Ro

ot

P c

on

cen

trat

ion

(%

)

DTPA-Cu (mg kg-1 soil)

y = -0.000x + 0.093R² = 0.857

0.089

0.090

0.091

0.092

0.093

0 1 2 3 4 5 6 7

Ro

ot

P c

on

cen

trat

ion

(%

)

DTPA-Cu (mg kg-1 soil)

y = 0.000x2 - 0.003x + 0.117R² = 0.963

0.11

0.11

0.11

0.12

0.12

0.12

0.12

0.12

0 1 2 3 4 5 6 7

Gra

in P

co

nce

ntr

atio

n (

%)

DTPA-Cu (mg kg-1 soil)

y = -0.000x2 + 0.002x + 0.070R² = 0.980

0.07

0.07

0.07

0.08

0.08

0.08

0.08

0 1 2 3 4 5 6 7

Gra

in P

co

nce

ntr

atio

n (

%)

DTPA-Cu (mg kg-1 soil)

y = -0.000x2 + 0.002x + 0.070R² = 0.980

0.07

0.07

0.07

0.08

0.08

0.08

0.08

0 1 2 3 4 5 6 7

Stra

w P

co

nce

ntr

atio

n (

%)

DTPA-Cu (mg kg-1 soil)

y = -0.001x2 + 0.008x + 0.080R² = 0.642

0.084

0.088

0.092

0.096

0.100

0.104

0 1 2 3 4 5 6 7

Stra

w P

co

nce

ntr

atio

n (

%)

DTPA-Cu (mg kg-1 soil)

(a) (b)

(c) (d)

(e) (f)

74

The effect of Cu levels as well as interactive effect of P and Cu on P concentration in

straw was not significant in both the soils. The quadratic response of straw P concentration to

Olsen P indicated that in loamy sand (Fig. 4.11e, R2=0.985) P concentration in straw

increased to the maximum up to a level of 90 mg P kg-1

soil and thereafter it declined. In

sandy loam (Fig. 4.11f, R2=0.991) straw P concentration increased with increasing Olsen P.

Phosphorus concentration in straw increased with increase in DTPA-Cu in loamy sand (Fig.

4.12e, R2=0.980). In sandy loam, straw P concentration increased with DTPA-Cu in soil and

reached to the maximum up to a level of 4 mg DTPA-Cu kg-1

soil and then it declined (Fig.

4.12f, R2= 0.642).

4.4.4 Copper concentration in root

In loamy sand soil, Cu concentration of root increased significantly from 304 µg g-1

dry matter to 481 and 507 µg g-1

dry matter when P was applied @ 200 and 400 mg kg-1

soil

resulting in an increase of 58.2 and 66.9 per cent, respectively over control. Mamo and

Parsons (1987) observed an increase of 34 per cent in root Cu concentration of teff plants with

application of 500 mg P kg-1

soil. These authors further observed that in shoot the

concentration of Cu decreased up to a level of 300 mg P kg-1

soil but increased when P level

was raised to 500 mg kg-1

soil. In sandy loam soil Cu concentration in root decreased

significantly by 14.15, 24.04, 33.03, 36.8, and 42.9 per cent over control with application of

25, 50, 100, 200 and 400 mg P kg-1

soil, respectively. It decreased from 67.8 µg g-1

in control

to 38.75 µg g-1

with application of 400 mg P kg-1

soil (Table 4.30). A significant negative

coefficient of correlation of root dry matter yield with root Cu concentration in loamy sand

(r=-0.408*) and sandy loam (r=-0.493

*) soil was observed (Table 4.34). Guan et al (2011)

studied the effect of Cu enriched livestock manure applications to wheat. They concluded that

although the quantities of Cu in wheat were very low compared to its accumulation in soil, the

Cu concentration in roots increased from 12 to 533 mg kg-1

and that in aerial parts was in a

narrow range from 12.1 to 32.7 mg kg-1

indicating that roots were more sensitive to Cu

toxicity. Michaud et al (2007) reported a range of 128-705 µg Cu g-1

in dry matter of wheat

roots of plants which exhibited Fe chlorosis.

The quadratic response of root Cu concentration to Olsen P indicated that in loamy

sand (Fig. 4.13a, R2=0.937) Cu concentration in root increased to the maximum up to a level

of 125 mg P kg-1

soil whereas in sandy loam (Fig. 4.13b, R2=0.924) the root Cu concentration

decreased to the maximum up to a level of 75 mg P kg-1

soil and thereafter it followed and

upward trend.

Root Cu concentration increased significantly from 32.78 µg g-1

in control to 274,

516 and 702 µg g-1

in loamy sand soil and from 29.4 µg g-1

in control to 33.4, 53.4 and 86.6

µg g-1

dry matter in sandy loam soil with application of 5, 10 and 20 mg Cu kg-1

soil,

respectively. In loamy sand, Cu accumulation in root increased by about 739, 1480 and

75

2046 per cent and in sandy loam increased by 13, 81 and 194 per cent over control with

application of 5, 10 and 20 mg Cu kg-1

soil, respectively. A linear positive relationship of

root Cu concentration with DTPA-Cu for loamy sand (Fig. 4.14a, R2=0.948) as well as for

sandy loam (Fig. 4.14b, R2=0.915) soil was observed. In light textured soil, a maximum

root accumulation of 906 µg Cu g-1

dry matter was observed with a combined application of

400 mg P and 20 mg Cu kg-1

soil. A significant interaction effect of P and Cu levels on root

Cu concentration for sandy loam soil was observed. It was revealed that in the absence of

applied Cu, the root Cu concentration decreased significantly from 36.26 µg g-1

dry matter

in control to 24.9, 23.5 and 22.2 µg g-1

when P was applied @ 100, 200 and 400 mg kg-1

soil.

Table 4.30: Effect of phosphorus and copper application on copper concentration of root

Copper

levels

(mg kg-1

soil)

Root Cu concentration (µg g-1

)

Phosphorus levels (mg kg-1

soil)

0 25 50 100 200 400 Mean

Loamy sand

0 37.5 32.0 32.0 32.2 32.6 30.2 32.7

5 268 268 290 264 2705 286 274

10 280 350 404 411 847 807 516

20 631 656 593 651 774 906 702

Mean 304 326 329 339 481 507

CD (5%) P levels 37.12 Cu levels 30.31 P x Cu 74.25

Sandy loam

0 36.2 35.5 34.1 24.9 23.5 22.2 29.4

5 40.7 39.7 35.9 32.7 30.4 20.8 33.4

10 68.60 63.1 52.2 47.6 45.2 43.6 53.4

20 125 94.5 84.0 76.5 70.8 68.3 86.6

Mean 67.8 58.2 51.5 45.4 42.5 38.7

CD (5%) P levels 3.01 Cu levels 2.45 P x Cu 6.01

4.4.5 Copper concentration in grain

The data on the effect of P and Cu levels on grain Cu concentration is presented in

Table 4.31. In loamy sand soil, the mean grain Cu concentration increased significantly from

7.61 µg g-1

in control to 9.95, 10.9 and 11.4 µg g-1

thereby resulting in an increase of 30.7,

44.2 and 50.8 per cent over control when P was applied @ 100, 200 and 400 mg kg-1

soil,

respectively. Whereas in sandy loam soil, grain Cu concentration significantly decreased from

76

8.34 µg g-1

in control to 7.66, 7.45, 7.40 and 7.39 µg g-1

thereby producing a decrease of 8.1,

10.6, 11.2 and 11.3 per cent over control when P was applied @ 50, 100, 200 and 400 mg kg-1

soil, respectively. However, the differences in grain Cu concentration observed with

application of 50, 100, 200 and 400 mg kg-1

soil were not significant. A negative coefficient

of correlation of grain yield with grain Cu concentration in loamy sand (r=-0.547**

) and sandy

loam (r=0.312) soil was observed (Table 4.34). Zhang et al (2012) observed that Cu

concentration in wheat grain but not in straw was significantly decreased by P applications up

to 100 mg kg-1 soil and thereafter remained constant.

The quadratic response of gain Cu concentration to Olsen P in loamy sand (Fig.

4.13c, R2=0.958) indicated that concentration of Cu in grain increased up to a level of 90 mg

P kg-1

soil and thereafter it declined. In sandy loam (Fig. 4.13d, R2=0.851) grain Cu

concentration decreased up to a level of 60 mg P kg-1

soil and thereafter it followed an upward

trend.

Grain Cu concentration increased significantly with application of graded levels of

Cu in both the soils. The content of Cu in grain produced in loamy sand soil was higher than

that produced in sandy loam soil. Mean grain Cu concentration increased from 4.61 µg g-1

in

control to 8.2, 10.8 and 13.2 µg g-1

in loamy sand soil and from 6.09 µg g-1

in control to 7.23,

7.90 and 9.84 µg g-1

in sandy loam soil with application of 5, 10 and 20 mg Cu kg-1

soil,

respectively. The increase in grain Cu content over control with application of 5, 10 and 20

mg Cu kg-1

soil was of the order of 77.8, 134 and 187 per cent in loamy sand and 18.7, 29.7

and 61.5 per cent in sandy loam soil, respectively. A positive linear relationship of grain Cu

concentration with DTPA-Cu in loamy sand (Fig. 4.14c, R2=0.941) as well as sandy loam

(Fig. 4.14d, R2=0.977) soil was observed.

The interaction effect of P and Cu levels on grain Cu in loamy sand calcareous soil

revealed that at each level of applied P its concentration increased with increasing levels of

applied Cu. A concentration of 10.58, 10.87, 11.25, 15.83, 14.95 and 16.0 µg g-1

was

observed when 20 mg Cu kg-1

soil was applied along with 0, 25, 50, 100, 200 and 400 mg P

kg-1

soil, respectively. Though severe Fe chlorosis of leaves was observed with application of

20 mg Cu kg-1

soil, yet the concentration of Cu in wheat grain did not approach the threshold

value for toxicity (20 mg kg-1

) (Guan et al 2011). In sandy loam soil, significant interaction

effect on grain Cu concentration indicated that in the absence of applied Cu, the grain Cu

concentration decreased significantly when P was applied @ 100, 200 and 400 mg P kg-1

soil.

However, the differences in grain Cu concentration at these levels of applied P were not

significant.

77

Table 4.31: Effect of phosphorus and copper application on copper concentration in

grain

Copper

levels

(mg kg-1

Grain Cu concentration (µg g-1

)

Phosphorus levels (mg kg-1

soil)

0 25 50 100 200 400 Mean

soil) Loamy sand

0 4.37 4.00 4.08 4.62 5.12 5.50 4.61

5 6.75 6.58 6.75 8.50 10.50 10.12 8.20

10 8.75 9.00 8.62 10.87 13.37 14.30 10.82

20 10.58 10.87 11.25 15.83 14.95 16.00 13.24

Mean 7.61 7.61 7.67 9.95 10.98 11.48

CD (5%) P levels 0.52 Cu levels 0.43 P x Cu 0.91

Sandy loam

0 7.50 7.37 6.58 5.33 5.08 4.68 6.09

5 7.50 7.75 7.25 6.75 7.16 7.00 7.23

10 7.87 7.91 7.83 7.66 8.12 8.00 7.90

20 10.50 10.37 9.00 10.08 9.25 9.87 9.84

Mean 8.34 8.35 7.66 7.45 7.40 7.39

CD (5%) P levels 0.43 Cu levels 0.35 P x Cu 0.87

4.4.6 Direct and indirect effects of various pools of copper towards copper concentration

of wheat grain

In loamy sand calcareous soil CARB-Cu played a significant role in effecting the Cu

concentration of wheat grain. Organically bound Cu controlled an indirect variation of 74.8

per cent in grain Cu concentration through CARB-Cu (Table 4.32). A direct variation of 6.1

and 30.9 per cent in grain Cu concentration was controlled by AMPOX-Cu and RES-Cu,

respectively. All the different pools of Cu together controlled an indirect variation of 44.4 per

cent in grain Cu concentration through AMPOX-Cu.

Like that on grain yield, the oxides and residual Cu played an important role in

controlling grain Cu concentration in sandy loam soil. A direct variation of 18.3, 15.8 and

20.7 per cent in grain Cu concentration was controlled by AMPOX-Cu, CRYOX-Cu and

RES-Cu, respectively. An indirect variation of 17.3, 12.2 and 18.3 per cent by DTPA-Cu,

14.9, 11.0 and 16.4 per cent by EXCH-Cu, 16.7, 13.0 and 16.5 per cent by SAD-Cu, 14.7,

10.5 and 17.6 per cent by MnOX-Cu, 18.3, 12.9 and 16.7 per cent by OM-Cu towards grain

Cu concentration was controlled through AMPOX-Cu, CRYOX-Cu and RES-Cu,

respectively (Table 4.32).

78

Table 4.32: Direct and indirect effects of various pools of copper towards copper

concentration of grain

Loamy sand soil

DTPA-

Cu

EXCH-

Cu

SAD-

Cu

CARB-

Cu

MnOX-

Cu

AMPOX-

Cu

CRYOX-

Cu

OM-

Cu

RES-

Cu

DTPA-Cu 0.159 -0.717 -0.085 1.15 -0.403 0.238 -0.100 0.052 0.544

EXCH-Cu 0.144 -0.794 -0.093 1.14 -0.436 0.209 -0.094 0.058 0.499

SAD-Cu 0.140 -0.769 -0.096 1.11 -0.451 0.204 -0.090 0.062 0.471

CARB-Cu 0.154 -0.743 -0.087 1.22 -0.434 0.225 -0.094 0.056 0.519

MnOX-Cu 0.135 -0.728 -0.091 1.11 -0.475 0.204 -0.091 0.063 0.447

AMPOX-Cu 0.153 -0.672 -0.079 1.11 -0.393 0.247 -0.101 0.048 0.525

CRYOX-Cu 0.141 -0.661 -0.077 1.01 -0.383 0.222 -0.113 0.049 0.476

OM-Cu 0.104 -0.581 -0.075 0.865 -0.382 0.222 -0.113 0.049 0.476

RES-Cu 0.156 -0.713 -0.081 1.11 -0.382 0.233 -0.096 0.046 0.556

Sandy loam soil

DTPA-Cu -0.315 0.060 0.251 - -0.361 0.416 0.350 0.044 0.428

EXCH-Cu -0.272 0.069 0.248 - -0.358 0.387 0.333 0.044 0.405

SAD-Cu -0.296 0.064 0.267 - -0.367 0.409 0.361 0.043 0.407

MnOX-Cu -0.286 0.062 0.246 - -0.398 0.384 0.325 0.052 0.420

AMPOX-Cu -0.307 0.063 0.255 - -0.357 0.428 0.360 0.043 0.409

CRYOX-Cu -0.278 0.058 0.242 - -0.325 0.386 0.398 0.022 0.366

OM-Cu -0.172 0.038 0.143 - -0.261 0.229 0.112 0.080 0.282

RES-Cu -0.296 0.061 0.239 - -0.367 0.383 0.319 0.049 0.456

Bold figures represent direct effects

4.4.7 Copper concentration in straw

The data on effect of P and Cu levels on Cu concentration in straw is presented in

Table 4.33. In both the soils Cu concentration in straw decreased significantly over control

with increasing levels of applied P. The magnitude of Cu concentration in straw was very

high in loamy sand soil as compared to sandy loam soil.

Straw Cu concentration decreased significantly from 38.00 µg g-1

in control to

32.77, 30.05, 26.45, 26.52 and 21.29 µg g-1

in loamy sand soil and from 10.07 µg g-1

in

control to 8.02, 7.23, 5.73, 5.29 and 4.51 µg g-1

in sandy loam soil when P was applied @ 25,

50, 100, 200 and 400 mg kg-1

soil, respectively. The corresponding values of per cent

decrease in straw Cu concentration over control with these levels of applied P were 13.7,

79

20.9, 30.3, 30.2 and 43.9 for loamy sand and 20.3, 28.2, 43.4, 47.4 and 55.2 for sandy loam

soil, respectively. A significant negative coefficient of correlation of straw yield with straw

Cu concentration in loamy sand (r=-0.848**

) and sandy loam (r=0.674**

) soil was observed

(Table 4.34). Shukla and Singh (1979) observed a significant decrease in Cu concentration in

grain and straw of wheat with increasing levels of applied P (0-250 mg kg-1

soil). In loamy

sand (Fig. 4.13e, R2=0.806) and sandy loam (Fig. 4.13f, R

2=0.915) soils, straw Cu

concentration decreased to the maximum level at 100 and 70 mg P kg-1

soil, respectively and

thereafter it either remained constant or declined.

The concentration of Cu in straw was much lower as compared to roots. This is due to

the reason that wheat plants are capable of regulating the and hence restricting the

translocation of acquired Cu in to aerial parts resulting in much larger concentration in roots

than in straw (Chaignon et al 2011).

Table 4.33: Effect of phosphorus and copper application on copper concentration in

straw

Copper

levels

(mg kg-1

soil)

Straw Cu concentration (µg g-1

)

Phosphorus levels (mg kg-1

soil)

0 25 50 100 200 400 Mean

Loamy sand

0 5.75 5.57 4.68 5.25 6.85 5.63 5.62

5 11.80 11.77 12.46 13.05 12.40 13.00 12.41

10 33.20 25.26 27.70 23.52 26.70 20.11 26.08

20 101.20 88.46 75.37 64.00 60.15 46.43 72.61

Mean 38.00 32.77 30.05 26.45 26.52 21.29

CD (5%) P levels 2.87 Cu levels 2.35 P x Cu 5.75

Sandy loam

0 6.06 5.92 4.51 3.46 3.46 3.18 4.43

5 8.32 7.60 7.52 4.47 4.30 3.91 6.02

10 8.50 7.57 6.75 4.90 4.90 4.88 6.25

20 17.41 11.01 10.15 10.11 8.51 6.06 10.54

Mean 10.07 8.02 7.23 5.73 5.29 4.51

CD (5%) P levels 0.88 Cu levels 0.71 P x Cu 1.76

Copper application significantly increased the Cu concentration in wheat straw in

both the soils. In loamy sand soil its concentration increased from 5.62 µg g-1

in control to

12.41, 26.08 and 72.61 µg g-1

and in sandy loam soil it increased from 4.43 µg g-1

in control

80

to 6.02, 6.25 and 10.54 µg g-1

when Cu was applied @ 5, 10 and 20 mg kg-1

soil, respectively.

The corresponding values of per cent increase in straw Cu concentration over control with

these levels of applied Cu were 120, 364, and 1191 for loamy sand and 35.8, 41.1 and 137 for

sandy loam soil, respectively. Straw Cu concentration in loamy sand (Fig. 4.14e, R2= 1) as

well as in sandy loam (Fig. 4.14f, R2= 0.953) increased with increase in DTPA-Cu. An

increase in Cu concentration in grain and straw of wheat with Cu application ranging from 4-

8 mg kg-1

soil were observed by Arshad et al (2011).

A significant interaction effect of P and Cu levels on Cu concentration in straw

revealed that at each level of applied P, it increased significantly over control when Cu was

applied @ 20 mg kg-1

soil in loamy sand soil. The content of Cu in wheat straw was 101.2,

88.46 and 75.37 µg g-1

and reached above the reported toxic level of 75 µg Cu g-1

in wheat

shoot (Reuter and Robinson 1997) when 0, 25 and 50 mg P kg-1

soil was combined with 20

mg Cu kg-1

soil. However, when 100, 200 and 400 mg P kg-1

soil was applied with 20 mg Cu

kg-1

soil the straw Cu concentration decreased to 64.0, 60.15 and 46.43 µg g-1

, respectively

thereby indicating that higher doses of applied tended to reduce the toxic effect of applied Cu.

In the absence of applied Cu in sandy loam soil, the Cu concentration in straw decreased

significantly over control when P was applied @100, 200 and 400 mg kg-1

soil. It decreased

from 6.06 µg g-1

in control to 3.18 µg g-1

with application of 400 mg P kg-1

soil.

Table 4.34: Linear coefficients of correlation of P, Cu and Fe concentration in root,

grain and straw with crop yield

Plant parameter Loamy sand Sandy loam

Root dry matter yield

P concentration 0.597**

0.622

Cu concentration -0.408* -0.493

*

Fe concentration -0.024 0.991**

Grain yield

P concentration 0.388 0.792**

Cu concentration -0.547**

-0.312

Fe concentration 0.336 -0.761**

Straw yield

P concentration 0.264 0.716**

Cu concentration -0.848**

-0.674**

Fe concentration 0.428* 0.540

**

*Significant at 5%

**Significant at 1%

81

Fig. 4.13: Relationship of Olsen P in soil at harvest with Cu concentration in root (a, b),

grain (c, d) and straw (e, f)

y = -0.014x2 + 3.721x + 271.4R² = 0.937

275

325

375

425

475

525

0 25 50 75 100 125 150

Ro

ot

Cu

co

ncn

etr

atio

n (

µg

g-1)

Olsen P (mg kg-1 soil)

y = 0.008x2 - 1.251x + 78.48R² = 0.924

30

35

40

45

50

55

60

65

70

0 25 50 75 100

Ro

ot

Cu

co

ncn

etr

atio

n (

µg

g-1)

Olsen P (mg kg-1 soil)

y = -0.000x2 + 0.100x + 6.623R² = 0.958

7.00

7.50

8.00

8.50

9.00

9.50

10.00

10.50

11.00

11.50

12.00

0 25 50 75 100 125 150

Gra

in C

u c

on

cen

trat

ion

(µ

g g-1

)

Olsen P (mg kg-1 soil)

y = 0.000x2 - 0.058x + 8.994R² = 0.851

7.00

7.20

7.40

7.60

7.80

8.00

8.20

8.40

0 25 50 75 100

Gra

in C

u c

on

cen

trat

ion

(µ

g g-1

)

Olsen P (mg kg-1 soil)

y = 0.001x2 - 0.251x + 36.22R² = 0.806

20

25

30

35

40

0 25 50 75 100 125 150

Stra

w C

u c

on

cen

trat

ion

(µ

g g-1

)

Olsen P (mg kg-1 soil)

y = 0.001x2 - 0.232x + 11.98R² = 0.915

3

5

7

9

11

0 25 50 75 100

Stra

w C

u c

on

cen

trat

ion

(µ

g g-1

)

Olsen P (mg kg-1 soil)

(a) (b)

(c) (d)

(e)

(f)

82

Fig. 4.14: Relationship of DTPA-Cu in soil at harvest with Cu concentration in root (a,

b), grain (c, d) and straw (e, f)

y = 102.7x + 63.45R² = 0.948

0

100

200

300

400

500

600

700

800

0 1 2 3 4 5 6 7

Ro

ot

Cu

co

nce

ntr

ario

n (

µg

g-1)

DTPA-Cu (mg kg-1 soil)

y = 10.88x + 12.48R² = 0.915

20

40

60

80

100

0 1 2 3 4 5 6 7

Ro

ot

Cu

co

nce

ntr

ario

n (

µg

g-1)

DTPA-Cu (mg kg-1 soil)

y = 1.300x + 5.191R² = 0.941

4

6

8

10

12

14

0 1 2 3 4 5 6 7

Gra

in C

u c

on

cen

trar

ion

(µ

g g-1

)

DTPA-Cu (mg kg-1 soil)

y = 0.676x + 5.388R² = 0.977

5

6

7

8

9

10

0 1 2 3 4 5 6 7

Gra

in C

u c

on

cen

trar

ion

(µ

g g-1

)

DTPA-Cu (mg kg-1 soil)

y = 1.340x2 + 1.095x + 5.303R² = 1

0

10

20

30

40

50

60

70

80

0 1 2 3 4 5 6 7

Stra

w C

u c

on

cen

trar

ion

(µ

g g-1

)

DTPA-Cu (mg kg-1 soil)

y = 0.184x2 - 0.267x + 4.736R² = 0.953

4

5

6

7

8

9

10

11

0 1 2 3 4 5 6 7

Stra

w C

u c

on

cen

trar

ion

(µ

g g-1

)

DTPA-Cu (mg kg-1 soil)

(a) (b)

(c) (d)

(e) (f)

83

4.4.8 Iron concentration in root

Data on root Fe concentration is presented in Table 4.35. In loamy sand alkaline

calcareous soil concentration of Fe in root decreased significantly from 2254 µg g-1

dry

matter in control to 2095, 2062, 2060 and 2080 µg g-1

dry matter which represented a

decrease of 7.05, 8.5, 8.6 and 7.7 per cent over control when P was applied @ 50, 100, 200

and 400 mg kg-1

soil, respectively. However, the differences in root Fe concentration

observed with application of 100, 200 and 400 mg P kg-1

soil were not significant. Michaud et

al (2007) reported a range of 1078-7166 µg Fe g-1

in dry matter of wheat roots grown in Cu

contaminated former vineyard soils. In medium textured sandy loam soil, significant decrease

of 10.3 per cent in root Fe concentration over control was observed only when P was applied

@ 400 mg kg-1

soil. A positive coefficient of correlation of root yield with root Fe

concentration in sandy loam (r=0.991**

) soil was observed (Table 4.34). Mamo and Parsons

(1987) observed a significant decrease in root and shoot Fe concentration of teff plants raised

in a Scottish soil with the highest level of applied P at 500 mg kg-1

soil.

Table 4.35: Effect of phosphorus and copper application on iron concentration in root

Copper

levels

(mg kg-1

soil)

Root Fe concentration (µg g-1

)

Phosphorus levels (mg kg-1

soil)

0 25 50 100 200 400 Mean

Loamy sand

0 2406 2243 2213 2136 2144 2076 2203

5 2301 2240 2205 2042 2077 2012 2146

10 2237 2191 2144 2043 2075 2109 2133

20 2075 2048 1820 2030 1947 2122 2007

Mean 2254 2181 2095 2062 2060 2080

CD (5%) P levels 118.2 Cu levels 96.5 P x Cu NS

Sandy loam

0 2847 2741 2602 2636 2641 2513 2663

5 2812 2695 2582 2852 2748 2310 2666

10 2572 2549 2755 2484 2580 2268 2534

20 2411 2525 2506 2359 2416 2453 2445

Mean 2661 2627 2611 2582 2596 2386

CD (5%) P levels 154.9 Cu levels 126.5 P x Cu NS

The quadratic response of root Fe concentration to Olsen P indicated that in loamy

sand (Fig. 4.15a, R2=0.691) Fe concentration in root decreased to the maximum up to a level

84

of 80 mg P kg-1

soil and thereafter it followed an upward trend whereas in sandy loam (Fig.

4.15b, R2=0.953) the root Fe concentration decreased with increasing levels of Olsen P.

Similarly, Cu application @ 20 mg kg-1

soil resulted in a significant decrease of 8.89

and 8.18 per cent in root Fe concentration over control in loamy sand and sandy loam soil,

respectively. A linear negative relationship of root Fe concentration with DTPA-Cu for loamy

sand (Fig. 4.16a, R2=0.962) as well as for sandy loam (Fig. 4.16b, R

2=0.890) soil was observed.

A significant reduction in Fe concentration of flag leaves of wheat by Cu application up to 320

mg kg-1 soil has been reported (Chhibba et al 1994). The interaction effect of P and Cu levels on

root Fe concentration was not significant in both the soils.

4.4.9 Iron concentration in grain

The data on Fe concentration in grain is presented in Table 4.36. Mean grain Fe

concentration increased significantly from 27.7 µg g-1

in control to 32.39, 41.61 and 41.4 µg

g-1

when P was applied @ 100, 200 and 400 mg kg-1

soil in loamy sand soil. The increase in

grain Fe content due to P application at 100, 200 and 400 mg kg-1

soil, respectively was 16.1,

49.6 and 48.9 per cent over control. However, the differences in grain Fe concentration

observed with 200 and 400 mg kg-1

soil were not significant. Zhang et al (2012) observed that

Fe concentration in wheat grain increased significantly with P application of more than 200

kg ha-1

but this increase was not observed for straw.

In sandy loam soil, the concentration of Fe in grain decreased significantly from 62.6

µg g-1

in control to 55.37, 50.48, 48.38, 49.36 and 48.16µg g-1

which represented a decrease

of 11.6, 19.4, 22.8, 21.2 and 23.1 per cent over control when P was applied @ 25, 50, 100,

200 and 400 mg kg-1

soil, respectively. The differences in grain Fe concentration observed

with application of 100, 200 and 400 mg P kg-1

soil were not significant. A negative

coefficient of correlation of grain yield with grain Fe concentration in sandy loam (r=-0.761**

)

soil was observed (Table 4.34). The quadratic response of gain Fe concentration to Olsen P in

loamy sand (Fig. 4.15c, R2=0.973) indicated that concentration of Fe in grain increased up to

a level of 90 mg P kg-1

soil and thereafter it declined. In sandy loam (Fig. 4.15d, R2=0.742)

grain Fe concentration decreased up to a level of 64 mg P kg-1

soil and thereafter it followed

an upward trend.

In loamy sand soil, grain Fe concentration decreased significantly from 36.65 µg g-1

in control to 32.16, 32.38 and 32.65 µg g-1

thereby resulting in a decrease of 12.2, 11.7 and

10.9 per cent over control when Cu was applied @ 5, 10 and 20 mg kg-1

soil, respectively.

However, the differences in grain Fe concentration observed with application of 10 and 20 mg

Cu kg-1

soil were not significant. The quadratic response of gain Fe concentration to DTPA-

Cu in loamy sand (Fig. 4.16c, R2=0.887) indicated that concentration of Fe in grain decreased

up to a level of 4 mg DTPA-Cu kg-1

soil and thereafter it increased. In sandy loam (Fig. 4.16d,

R2=0.616) grain Fe concentration increased with increasing DTPA-Cu content.

85

Table 4.36: Effect of phosphorus and copper application on iron concentration in grain

Copper

levels

(mg kg-1

soil)

Grain Fe concentration (µg g-1

)

Phosphorus levels (mg kg-1

soil)

0 25 50 100 200 400 Mean

Loamy sand

0 27.0 27.3 33.6 43.4 44.3 44.2 36.6

5 30.6 29.1 28.2 26.1 33.3 45.5 32.1

10 26.0 27.6 27.3 29.8 46.0 37.5 32.3

20 27.5 28.5 28.6 30.1 42.7 38.3 32.6

Mean 27.8 28.1 29.4 32.3 41.6 41.4

CD (5%) P levels 3.05 Cu levels 2.49 P x Cu NS

Sandy loam

0 61.0 52.1 51.5 48.8 45.4 50.3 51.5

5 60.5 51.6 48.0 44.5 48.6 48.2 50.3

10 66.2 59.0 48.8 49.2 51.3 47.5 53.7

20 62.8 58.7 53.5 50.8 52.0 46.5 54.1

Mean 62.6 55.3 50.4 48.3 49.3 48.1

CD (5%) P levels 3.77 Cu levels NS P x Cu NS

4..4.10 Iron concentration in straw

Concentration of Fe in straw increased significantly over control in both the soils with

the application of 200 and 400 mg kg-1

soil (Table 4.37). In loamy sand soil, it increased

significantly from187 µg g-1

in control to 237 and 228 µg g-1

and in sandy loam it increased

from 258.08 µg g-1

in control to 331 and 367 µg g-1

with application of 200 and 400 mg P kg-1

soil, respectively. The corresponding values of per cent increase in straw Fe concentration

over control with these levels of applied P were 26.4 and 21.9 for loamy sand and 28.4 and

42.5 for sandy loam soil, respectively. A positive coefficient of correlation of straw yield with

straw Fe concentration in loamy sand (r=0.428*) and sandy loam (r=-0.540**

) soil was

observed (Table 4.34). The quadratic response of straw Fe concentration to Olsen P indicated

that in loamy sand (Fig. 4.15e, R2=0.922) Fe concentration in straw increased to the

maximum up to a level of 90 mg P kg-1

soil and thereafter it declined. In sandy loam (Fig.

4.15f, R2=0.854) straw Fe concentration increased with increasing Olsen P. Straw Fe

concentration decreased with increase in DTPA-Cu in loamy sand (Fig. 4.16e, R2=0.992) but

increased in sandy loam (Fig. 4.16f, R2= 0.785).

In sandy loam soil the effect of Cu levels on Fe concentration in straw was not

significant but in loamy sand soil Fe concentration in straw decreased significantly from 220

86

µg g-1

in control to 195 and 194 µg g-1

when Cu was applied @ 10 and 20 mg kg-1

soil thereby

resulting in a decrease of 11.1 and 11.7 per cent over control, respectively. The interaction

effect of P and Cu levels on Fe concentration in straw was not significant in both the soils.

The data on linear coefficients of correlation of chemical pools of Cu with Cu

concentration in root, grain and straw revealed that in both the soils, root Cu, grain Cu and

straw Cu concentration was significantly positively correlated with the various pools of Cu

including DTPA-Cu (Table 4.38) except that organically bound Cu was poorly correlated

with root and straw Cu concentration in sandy loam. Root Fe concentration was significantly

negatively correlated with various pools of Cu in both soils. But grain and straw Fe

concentration with various pools of Cu was negatively correlated in loamy sand and

positively in sandy loam except for the relation between OM-Cu and straw Cu content.

Phosphorus concentration in root, grain and straw was significantly positively correlated with

DTPA-Zn, Mn and Fe in both soils (Table 4.38). Among the various pools of Cu only

organically bound Cu was significantly positively correlated with P concentration in root

(r=0.470*), grain (r=0.610

**) and straw (=0.574

**) in sandy loam.

Table 4.37 Effect of phosphorus and copper application on iron concentration in straw

Copper

levels

(mg kg-1

soil)

Straw Fe concentration (µg g-1

)

Phosphorus levels (mg kg-1

soil)

0 25 50 100 200 400 Mean

Loamy sand

0 198.6 197.5 196.8 217.6 251.0 259.6 220.2

5 189.6 190.5 176.8 197.8 251.6 234.0 206.7

10 180.6 177.0 178.1 165.3 254.8 218.0 195.6

20 182.0 180.0 181.3 224.5 192.4 204.2 194.1

Mean 187.7 186.2 183.2 201.3 237.4 228.9

CD (5%) P levels 22.7 Cu levels 18.5 P x Cu NS

Sandy loam

0 234.0 213.1 220.1 234.1 327.5 363.3 265.3

5 224.3 279.3 297.5 225.8 338.5 385.6 291.8

10 304.0 235.5 278.3 225.1 307.6 353.6 284.0

20 270.0 293.0 251.0 302.1 352.5 368.8 306.2

Mean 258.1 255.2 261.7 246.8 331.5 367.8

CD (5%) P levels 38.3 Cu levels NS P x Cu NS

87

Fig. 4.15: Relationship of Olsen P in soil at harvest with Fe concentration in root (a, b),

grain (c, d) and straw (e, f)

y = 0.031x2 - 5.271x + 2234.R² = 0.691

1950

2000

2050

2100

2150

2200

2250

2300

0 25 50 75 100 125 150

Ro

ot

Fe c

on

cen

trat

ion

(µ

g g-1

)

Olsen P (mg kg-1 soil)

y = -0.022x2 - 0.590x + 2649.R² = 0.953

2350

2400

2450

2500

2550

2600

2650

2700

0 25 50 75 100

Ro

ot

Fe c

on

cen

trat

ion

(µ

g g-1

)

Olsen P (mg kg-1 soil)

y = -0.001x2 + 0.354x + 24.19R² = 0.973

25

30

35

40

45

0 25 50 75 100 125 150

Gra

in F

e c

on

cen

trat

ion

((µ

g g-1

)

Olsen P (mg kg-1 soil)

y = 0.005x2 - 0.684x + 66.72R² = 0.742

40

45

50

55

60

65

0 25 50 75 100

Gra

in F

e c

on

cen

trat

ion

((µ

g g-1

)

Olsen P (mg kg-1 soil)

y = -0.007x2 + 1.389x + 169.6R² = 0.922

170

180

190

200

210

220

230

240

0 25 50 75 100 125 150

Stra

w F

e c

on

cen

trat

ion

((µ

g g-1

)

Olsen P (mg kg-1 soil)

y = 1.524x + 229.3R² = 0.854

225

250

275

300

325

350

375

0 25 50 75 100

Stra

w F

e c

on

cen

trat

ion

((µ

g g-1

)

Olsen P (mg kg-1 soil)

(a) (b)

(c) (d)

(e) (f)

88

Fig. 4.16: Relationship of DTPA-Cu in soil at harvest with Fe concentration in root (a, b), grain

(c, d) and straw (e, f)

y = -29.39x + 2213.R² = 0.962

1975

2025

2075

2125

2175

2225

0 1 2 3 4 5 6 7

Ro

ot

Fe c

on

cen

trat

ion

(µ

g g-1

)

DTPA-Cu (mg kg-1 soil)

y = -44.11x + 2731.R² = 0.890

2400

2450

2500

2550

2600

2650

2700

0 1 2 3 4 5 6 7

Ro

ot

Fe c

on

cen

trat

ion

(µ

g g-1

)

DTPA-Cu (mg kg-1 soil)

y = 0.274x2 - 2.438x + 36.75R² = 0.887

31

32

33

34

35

36

37

0 1 2 3 4 5 6 7

Gra

in F

e c

on

cen

trat

ion

(µ

g g-1

)

DTPA-Cu (mg kg-1 soil)

y = 0.618x + 50.23R² = 0.616

50.0

51.0

52.0

53.0

54.0

55.0

0 1 2 3 4 5 6 7

Gra

in F

e c

on

cen

trat

ion

(µ

g g-1

)

DTPA-Cu (mg kg-1 soil)

y = 0.972x2 - 10.82x + 222.8R² = 0.992

190

195

200

205

210

215

220

225

0 1 2 3 4 5 6 7

Star

w F

e c

on

cen

trat

ion

(µ

g g-1

)

DTPA-Cu (mg kg-1 soil)

y = 6.544x + 263.8R² = 0.785

260

270

280

290

300

310

0 1 2 3 4 5 6 7

Stra

w F

e c

on

cen

trat

ion

(µ

g g-1

)

DTPA-Cu (mg kg-1 soil)

(a) (b)

(c) (d)

(e) (f)

89

Table 4.38: Linear coefficients of correlation of P, Cu and Fe concentration in root, grain and straw with chemical pools of Cu and DTPA-

extractable micronutrient cations

Phosphorus concentration Copper concentration Iron concentration

Root Grain Straw Root Grain Straw Root Grain Straw

Loamy sand

EXCH-Cu -0.113 -0.154 0.043 0.682**

0.635**

0.953**

-0.520**

-0.160 -0.312

SAD-Cu -0.127 -0.174 0.034 0.665**

0.585**

0.974**

-0.559**

-0.169 -0.268

CARB-Cu 0.138 0.015 0.224 0.826**

0.814**

0.899**

-0.667**

-0.042 -0.194

MnOX-Cu -0.056 -0.182 0.038 0.608**

0.578**

0.932**

-0.603**

-0.247 -0.278

AMPOX-Cu 0.096 -0.084 0.155 0.885**

0.841**

0.876**

-0.595**

-0.166 -0.375

CRYOX-Cu -0.085 -0.229 0.005 0.780**

0.675**

0.835**

-0.397 -0.267 -0.438

OM-Cu -0.214 -0.232 -0.168 0.483* 0.418

* 0.708

** -0.424

* -0.346 -0.283

RES-Cu 0.044 -0.041 0.154 0.867**

0.858**

0.880**

-0.560**

-0.122 -0.299

DTPA-Cu 0.029 -0.092 0.147 0.890**

0.843**

0.911**

-0.567**

-0.147 -0.321

DTPA-Zn 0.663**

0.767**

0.753**

0.356 0.477* -0.074 -0.384 0.731

** 0.610

**

DTPA-Mn 0.662**

0.765**

0.753**

0.364 0.485* -0.065 -0.390 0.728

** 0.605

**

DTPA-Fe 0.664**

0.769**

0.753**

0.348 0.470* -0.082 -0.380 0.733

** 0.613

**

Sandy loam

EXCH-Cu -0.180 0.035 -0.022 0.899**

0.857**

0.767**

-0.560**

0.261 0.214

SAD-Cu -0.193 0.026 -0.010 0.921**

0.891**

0.775**

-0.516**

0.302 0.267

MnOX-Cu 0.002 0.229 0.176 0.787**

0.807**

0.563**

-0.560**

0.108 0.341

AMPOX-Cu -0.210 -0.034 -0.031 0.905**

0.895**

0.760**

-0.493* 0.299 0.194

CRYOX-Cu -0.280 -0.100 -0.074 0.911**

0.872**

0.781**

-0.408* 0.359 0.106

OM-Cu 0.470* 0.610

** 0.574

** 0.261 0.453

* 0.067 -0.706

** -0.265 0.669

**

RES-Cu 0.012 0.080 0.077 0.781**

0.847**

0.598**

-0.564**

0.114 0.258

DTPA-Cu -0.155 -0.025 0.000 0.863**

0.873**

0.699**

-0.510* 0.249 0.190

DTPA-Zn 0.730**

0.948**

0.957**

-0.196 -0.074 -0.386 -0.601**

-0.525**

0.822**

DTPA-Mn 0.727**

0.947**

0.956**

-0.184 -0.062 -0.376 -0.607**

-0.521**

0.824**

DTPA-Fe 0.732**

0.949**

0.957**

-0.206 -0.084 -0.394 -0.595**

-0.528**

0.821**

*Significant at 5%

**Significant at 1%

90

4.5 Copper to iron concentration ratios in root, grain and straw

4.5.1 Root Cu: Fe ratio

In root the ratio of Cu to Fe concentration increased significantly from 0.141in

control to a maximum of 0.242 in alkaline calcareous loamy sand while that in acidic sandy

loam it decreased from 0.026 in control to a maximum of 0.016 with an application of 400 mg

P kg-1

soil (Table 4.39). However, with Cu application the concentration ratio of Cu: Fe in

root increased with increasing levels of applied Cu in both the soils. The magnitude of this

increase was more pronounced in Fe deficient (DTPA-Fe 3.2 mg kg-1

soil) loamy sand

calcareous soil than sandy loam which was highly adequate in available Fe (DTPA-Fe 51.6

mg kg-1

soil). In loamy sand, root Cu: Fe concentration ratio increased significantly from

0.014 in control to 0.128, 0.245, and 0.350 thereby representing an increase of 814, 1650 and

24 per cent over control with an application of 5, 10 and 20 mg Cu kg-1

soil, respectively. The

corresponding values of this increase for sandy loam soil were only 9, 90 and 218 per cent

over control, respectively. These findings are in conformity with those of Michaud et al

(2007) who reported that root Fe concentration in durum wheat tended to decrease with

increase in root Cu as well as soil Cu concentration particularly in calcareous soils. A

comprehensive plot (n=48) of Fe and Cu concentration in root (Fig. 4.17a) for both soils taken

together showed that as the Cu accumulation in root increased the concentration of Fe

decreased. The concentration of Fe in root decreased steeply up to a level of 250 µg Cu g-1

in

root dry matter and thereafter the rate of decrease was slowed down. Kumar et al (2009)

observed a significant reduction of 10.3 per cent in Fe concentration of wheat leaves with

application of 2.5 mg Cu kg-1

soil. Brar and Sekhon (1978) reported that excess Cu inhibited

the translocation of Fe from stem to leaves in wheat. Iron deficiency in Cu contaminated soil

resulted in an elevated acquisition of Cu by wheat possibly due to an increased release of

phytosiderophores and a major proportion of acquired Cu accumulated in roots (Chaignon et

al 2002). Waters and Armbrust (2013) also reported that Cu and Fe nutrition interact to

increase or decrease Fe and/or Cu accumulation in leaves and Fe uptake processes. They

further observed that high leaf Cu concentration lowered leaf Fe concentration and under Fe

deficient conditions the concentration of Cu in leaves is increased. Chhibba et al (1994)

observed a significant decrease in Fe concentration in flag leaves of wheat with Cu

application at rates as high as 160 mg kg-1

soil.

A significant interaction effect of P and Cu levels on root Cu: Fe ratios in calcareous

soil indicated that at any level of applied P, this ratio increased significantly with each level of

applied Cu over the previous level. But in sandy loam acidic soil, the root Cu: Fe ratio

increased significantly over control only when 10 and 20 mg Cu kg-1

soil was combined with

25 and 50 mg P kg-1

soil but it decreased significantly over control when combined with 100,

200 and 400 mg P kg-1

soil, respectively.

91

It has been discussed in the previous section that in alkaline calcareous soil severe Fe

chlorosis of leaves was observed when 20 mg Cu kg-1

soil was combined with any level of

applied P. The root Cu: Fe ratio in this soil varied between 0.014 to 0.015 only when no Cu

was applied, however when 20 mg Cu kg-1

soil was combined with graded levels of P (P0Cu20,

P25Cu20, P50Cu20, P100Cu20, P200Cu20 and P400Cu20) the root Cu: Fe ratio increased and varied

between 0.304 to 0.429 which was about 2170 to 2860 per cent of control. While assessing

the in situ Cu uptake and phytotoxicity for durum wheat cultivated in Cu contaminated former

vineyard soils Michaud et al (2007) also observed that the Cu: Fe concentration ratio in roots

of wheat plants that exhibited severe Fe chlorosis varied from 0.129 to 0.519. In sandy loam

soil, in which no Fe chlorosis was observed with any combination of P and Cu the root Cu: Fe

ratio with the above mentioned combinations varied from 0.028 to 0.052 only. Overall, in

sandy loam soil the root Cu: Fe ratio varied from 0.011 to 0.093 and in calcareous soil it

varied from 0.014 to 0.429. The results of this study indicated that Fe chlorosis of wheat

leaves is expected when Cu: Fe concentration ratio in root is more than 0.30 especially in

calcareous soil.

Table 4.39: Effect of phosphorus and copper application on Cu: Fe concentration ratios

in root

Copper

levels

(mg kg-1

soil)

Root Cu: Fe ratio

Phosphorus levels (mg kg-1

soil)

0 25 50 100 200 400 Mean

Loamy sand

0 0.015 0.014 0.014 0.015 0.015 0.014 0.014

5 0.116 0.119 0.132 0.129 0.130 0.142 0.128

10 0.127 0.161 0.189 0.201 0.412 0.383 0.245

20 0.304 0.320 0.327 0.321 0.398 0.429 0.350

Mean 0.141 0.153 0.165 0.166 0.238 0.242

CD (5%) P levels 0.021 Cu levels 0.017 P x Cu 0.042

Sandy loam

0 0.013 0.013 0.013 0.093 0.090 0.090 0.011

5 0.014 0.014 0.013 0.011 0.011 0.093 0.012

10 0.026 0.025 0.019 0.019 0.018 0.019 0.021

20 0.052 0.038 0.033 0.032 0.029 0.028 0.035

Mean 0.026 0.022 0.019 0.018 0.018 0.016 0.016

CD (5%) P levels 0.002 Cu levels 0.001 P x Cu 0.004

92

Fig.4.17: Relationship of Fe concentration in root and straw with Cu concentration in

root (a) and straw (b) irrespective of the soil (n=48)

4.5.2 Grain Cu: Fe ratio

In loamy sand soil, the effect of P application on Cu: Fe ratio in grain was not

significant and in sandy loam soil it increased significantly from 0.133 in control to 0.150

when 25 mg P kg-1

soil was applied and thereafter it remained constant (Table 4.40).

Application of Cu significantly increased the Cu: Fe ratio in grain in both the soils. It

increased from 0.129 in control to 0.260, 0.340 and 0.409 in loamy sand calcareous soil and

from 0.118 in control to 0.145, 0.150 and 0.184 in sandy loam with application of 5, 10 and

20 mg Cu kg-1

soil, respectively. The grain Cu: Fe ratio increased by about 101, 163 and 217

per cent over control in loamy sand and by about 22.8, 27.1 and 42.6 per cent over control in

sandy loam with graded levels of applied Cu, respectively. Again the magnitude of increase in

grain Cu: Fe ratio was more pronounced in loamy sand calcareous soil as compared to sandy

loam. A significant interaction of P and Cu levels revealed that grain Cu: Fe ratio increased

significantly over control when 20 mg Cu kg-1

soil was combined with any level of applied P

in both the soils.

The grain Cu: Fe ratio in calcareous soil varied from 0.116 to 0.162 when no Cu

was applied. However, when 20 mg Cu kg-1

soil was combined with graded levels of P

(P0Cu20, P25Cu20, P50Cu20, P100Cu20, P200Cu20 and P400Cu20) the grain Cu: Fe ratio increased

and varied from 0.384 to 0.525. In sandy loam soil, in which no Fe chlorosis was

observed with any combination of P and Cu the grain Cu: Fe ratio with the above

mentioned combinations of P and Cu varied from 0.166 to 0.214. Overall, in sandy loam

soil the grain Cu: Fe ratio varied from 0.094 to 0.214 and in calcareous soil varied from

0.106 to 0.525.

y = 3123.x-0.06

R² = 0.509

1750

2000

2250

2500

2750

3000

0 250 500 750 1000

Ro

ot

Fe c

on

cen

trat

ion

(µ

g g-1

)

Root Cu concentration (µg g-1)

(a)

y = -35.2ln(x) + 329.4R² = 0.298

100

150

200

250

300

350

400

0 30 60 90 120

Stra

w F

e c

on

cen

trat

ion

(µ

g g-1

)

Straw Cu concentration (µg g-1)

(b)

93

Table 4.40: Effect of phosphorus and copper application on Cu: Fe concentration

ratios in grain

Copper

levels

(mg kg-1

soil)

Grain Cu: Fe ratio

Phosphorus levels (mg kg-1

soil)

0 25 50 100 200 400 Mean

Loamy sand

0 0.162 0.147 0.122 0.106 0.116 0.124 0.129

5 0.222 0.228 0.241 0.326 0.317 0.229 0.260

10 0.336 0.326 0.329 0.367 0.292 0.390 0.340

20 0.384 0.382 0.396 0.525 0.350 0.417 0.409

Mean 0.276 0.271 0.272 0.331 0.269 0.290

CD (5%) P levels NS Cu levels 0.025 P x Cu 0.061

Sandy loam

0 0.124 0.141 0.128 0.109 0.112 0.094 0.118

5 0.124 0.150 0.151 0.152 0.149 0.145 0.145

10 0.119 0.135 0.160 0.160 0.159 0.168 0.150

20 0.166 0.176 0.168 0.199 0.179 0.214 0.184

Mean 0.133 0.150 0.151 0.155 0.150 0.155

CD (5%) P levels 0.013 Cu levels 0.011 P x Cu 0.026

4.5.3 Straw Cu: Fe ratio

In loamy sand soil, straw Cu: Fe ratio decreased significantly from 0.208 in control to

a maximum of 0.100 and in sandy loam soil it decreased from 0.039 in control to 0.012 when

400 mg P kg-1

soil was applied (Table 4.41). A comprehensive plot (n=48) of Fe and Cu

concentration in straw (Fig. 4.17b) for both soils taken together also showed that as the Cu

accumulation in straw increased the concentration of Fe decreased. The concentration of Fe in

straw decreased steeply up to a level of 30 µg Cu g-1

in root dry matter and thereafter the rate

of decrease was slowed down. Application of Cu significantly increased the Cu: Fe ratio in

straw in both the soils. It increased from 0.026 in control to 0.061, 0.137 and 0.384 in loamy

sand calcareous soil and from 0.018 in control to 0.022, 0.022 and 0.036 in sandy loam with

application of 5, 10 and 20 mg Cu kg-1

soil, respectively. The straw Cu: Fe ratio increased by

94

about 134, 426 and 1376 per cent over control in loamy sand and by about 22, 22 and 100 per

cent over control in sandy loam with graded levels of applied Cu, respectively. Again the

magnitude of increase in straw Cu: Fe ratio was more pronounced in loamy sand calcareous

soil as compared to sandy loam. A significant interaction of P and Cu levels revealed that

straw Cu: Fe ratio increased significantly over control when 20 mg Cu kg-1

soil was combined

with any level of applied P in both the soils.

The straw Cu: Fe ratio in calcareous soil varied from 0.022 to 0.029 when no Cu was

applied. However, when 20 mg Cu kg-1

soil was combined with graded levels of P (P0Cu20,

P25Cu20, P50Cu20, P100Cu20, P200Cu20 and P400Cu20) the straw Cu: Fe ratio increased and varied

from 0.231 to 0.557. In sandy loam soil, in which no Fe chlorosis was observed with any

combination of P and Cu the straw Cu: Fe ratio with the above mentioned combinations of P

and Cu varied from 0.016 to 0.064. Overall, in sandy loam soil the straw Cu: Fe ratio varied

from 0.022 to 0.557 and in calcareous soil varied from 0.010 to 0.064.

Table 4.41: Effect of phosphorus and copper application on straw Cu: Fe ratios of wheat

Copper

levels

(mg kg-1

soil)

Straw Cu: Fe ratio

Phosphorus levels (mg kg-1

soil)

0 25 50 100 200 400 Mean

Loamy sand

0 0.029 0.028 0.024 0.025 0.027 0.022 0.026

5 0.062 0.063 0.071 0.068 0.049 0.055 0.061

10 0.183 0.143 0.156 0.142 0.107 0.092 0.137

20 0.557 0.492 0.415 0.286 0.323 0.231 0.384

Mean 0.208 0.181 0.166 0.130 0.127 0.100

CD (5%) P levels 0.018 Cu levels 0.014 P x Cu 0.036

Sandy loam

0 0.026 0.028 0.020 0.015 0.010 0.016 0.018

5 0.037 0.029 0.025 0.020 0.013 0.010 0.022

10 0.028 0.032 0.024 0.022 0.016 0.013 0.022

20 0.064 0.039 0.040 0.034 0.024 0.016 0.036

Mean 0.039 0.032 0.027 0.023 0.016 0.012

CD (5%) P levels 0.004 Cu levels 0.003 P x Cu 0.009

95

4.6 Upper critical levels of Cu in soil, root, grain and straw

Upper critical levels of Cu associated with 50 per cent reduction from maximum

yield, which are generally considered to be toxic for plants were determined by using the data

of all the 24 treatment combinations for both the soils taken together (n=48). The response of

relative dry matter yield of root (Fig. 4.18a), grain (Fig. 4.18b) and straw (Fig. 4.18c) to

DTPA extractable Cu in soil at harvest was observed to be quadratic with significant values of

coefficients of determination. A content of 6.56, 5.39 and 5.37 mg DTPA-extractable Cu kg-1

soil produced 50 per cent reduction from the maximum yield of root, grain and straw,

respectively which may be considered as the upper critical values for wheat (Table 4.42). In a

Typic Ustipsament, Chhibba et al (1994) reported that a content of 7.9 mg DTPA-Cu kg-1

soil

reduced the grain and dry matter yield of wheat by 50 per cent from the maximum yield. They

considered it to be the upper critical level of Cu in soil for wheat growth. The response of

relative dry matter yield of root (Fig. 4.18d), grain (Fig.4.18e) and straw (Fig. 4.18f) to Cu

concentration in root, grain and straw, respectively was also quadratic with significant

coefficients of determination. A concentration of >18 µg Cu g-1

in wheat tissue has been

reported to be toxic (Davis and Beckett 1978). In the present study, a Cu concentration of

436, 11.04 and 19.33µg g-1

reduced the yield of root, grain and straw, respectively by 50 per

cent from the maximum yield which may be considered as the upper critical levels of Cu in

root, grain and straw of wheat. Chhibba et al (1994) also observed that a concentration of 25.5

and 25.7µg Cu g-1

in flag leaves of wheat reduced the dry matter and grain yield by 50 per

cent, respectively indicating that Cu at these levels may prove toxic for wheat.

Table 4.42: Upper critical levels of Cu in soil and plant parts

Plant

part

Quadratic model R2 Critical level for 50% reduction

of maximum yield

DTPA-Cu (mg kg-1

soil)

Root Y= -1.617X2 + 9.466X + 57.58 0.159

* 6.56

Grain Y= -1.758X2 + 7.115X + 62.77 0.350

** 5.39

Straw Y= -1.704X2 + 7.562X + 58.56 0.307

** 5.37

Plant part (µg g-1

)

Root Y= 3E-05X2 – 0.071X + 75.27 0.398

** 436.0

Grain Y= -0.445X2 + 3.956X + 60.60 0.350

** 11.04

Straw Y= 0.008X2 -1.603X + 78.00 0.744

** 19.33

* Significant at 5%

** Significant at 1%

96

Fig.4.18: Relationship of relative root, grain and straw yield with DTPA-Cu in soil at

harvest (a, b, c) and Cu concentration in their respective plants parts (d, e, f)

in soil at harvest irrespective of the soil (n=48)

y = -1.617x2 + 9.466x + 57.58R² = 0.159

20

40

60

80

100

0 1 2 3 4 5 6 7 8

Re

lati

ve r

oo

t d

ry m

atte

r yi

eld

(%

)

DTPA- Cu (mg kg-1 soil)

(a)

y = -1.758x2 + 7.115x + 62.77R² = 0.350

0

20

40

60

80

100

0 1 2 3 4 5 6 7 8

Re

lati

ve g

rain

yie

ld (

%)

DTPA-Cu (mg kg-1 soil)

(b)

y = -1.704x2 + 7.562x + 58.56R² = 0.307

0

20

40

60

80

100

0 1 2 3 4 5 6 7 8

Re

lati

ve s

traw

yie

ld (

%)

DTPA -Cu (mg kg-1 soil)

(c)y = 3E-05x2 - 0.071x + 75.27

R² = 0.398

20

40

60

80

100

0 250 500 750 1000

Re

lati

ve r

oo

t d

ry m

atte

r yi

eld

(%)

Root Cu concentration (µg g-1)

(d)

y = -0.445x2 + 3.956x + 60.60R² = 0.350

0

20

40

60

80

100

2 4 6 8 10 12 14 16

Re

lati

ve g

rain

yie

ld (

%)

Grain Cu Concentration (µg g-1)

(e)

y = 0.008x2 - 1.603x + 78.00R² = 0.744

0

20

40

60

80

100

0 10 20 30 40 50 60 70 80 90 100

Re

lati

ve s

traw

yie

ld (

%)

Straw Cu concentration (µg g-1)

(f)

97

4.7 Effect of phosphorus and copper application on P, Cu and Fe uptake by root, grain

and straw

4.7.1 Phosphorus uptake by root

The data on P uptake by root as influenced by graded levels of applied P and Cu is

presented in Table 4.43. In loamy sand soil, mean P uptake by root increased by 100, 286,

434, 588 and 527 per cent over control with application of 25, 50, 100, 200 and 400 mg P kg-1

soil, respectively. It increased significantly from 0.215 mg pot-1

in control to a maximum of

1.48 mg pot-1

when P was applied @ 200 mg kg-1

soil and it decreased significantly to 1.35

mg pot-1

when P level was increased to 400 mg kg-1

soil. In medium textured soil, P uptake

by root increased significantly by 30.1, 29.2, 49.05, 92.4 and 113.2 per cent over control with

application of 25, 50, 100, 200 and 400 mg P kg-1

soil, respectively. It increased from 1.06 mg

pot-1

in control to a maximum of 2.26 mg pot-1

when P was applied @ 400 mg P kg-1

soil. A

significant positive coefficient of correlation of P uptake by root with root dry matter yield in

loamy sand (r=0.897**) and sandy loam (r=0.908**) was observed (Table 4.49). The

quadratic response of root P uptake to Olsen P indicated that in loamy sand (Fig. 4.19a,

R2=0.939) and sandy loam (Fig. 4.19b, R

2=0.971) soil maximum root P uptake was observed

up to a level of about 90 and 90 mg P kg-1

soil, respectively and thereafter it declined.

Table 4.43: Effect of phosphorus and copper application on phosphorus uptake by root

Copper

levels

(mg kg-1

soil)

Root P uptake (mg pot-1

)

Phosphorus levels (mg kg-1

soil)

0 25 50 100 200 400 Mean

Loamy sand

0 0.240 0.404 0.713 1.270 1.780 1.650 1.011

5 0.238 0.492 1.130 1.230 1.580 1.880 1.090

10 0.204 0.481 0.935 1.470 1.910 1.450 1.070

20 0.180 0.342 0.552 0.660 0.660 0.440 0.470

Mean 0.215 0.430 0.830 1.150 1.480 1.350

CD (5%) P levels 0.120 Cu levels 0.098 P x Cu 0.240

Sandy loam

0 1.04 1.46 1.45 1.35 2.38 2.22 1.65

5 1.19 1.45 1.42 1.48 2.08 2.33 1.66

10 1.10 1.35 1.32 1.69 14.99 2.22 1.61

20 0.91 1.24 1.29 1.80 1.72 2.28 1.54

Mean 1.06 1.38 1.37 1.58 2.04 2.26

CD (5%) P levels 0.146 Cu levels NS P x Cu NS

98

Copper application @ 5, 10 and 20 mg kg-1

soil significantly decreased P uptake by

root in loamy sand soil but its effect was not significant in sandy loam soil. In loamy sand soil

P uptake by root decreased significantly from 1.01 mg pot-1

in control to 0.47 mg pot-1

thereby resulting in a decrease of 53.5 per cent over control when 20 mg Cu kg-1

soil was

applied. The quadratic relationship of DTPA-Cu with root P uptake indicated that P uptake by

root in loamy sand (Fig. 4.20a, R2=0.996) as well as sandy loam (Fig. 4.20b, R

2=0.977)

decreased sharply when soil had >2 mg DTPA-Cu kg-1

soil.

A significant interaction effect of P and Cu levels on P uptake by root was observed

in loamy sand soil. At each level of applied P, it decreased significantly over control with

application of 20 mg Cu kg-1

soil. In sandy loam soil, P uptake by root decreased significantly

by 6.66 per cent over control with the highest level of applied Cu.

4.7.2 Phosphorus uptake by grain

The data on P uptake by grain is presented in Table 4.44. With the increasing levels

of applied P, its uptake by grain increased significantly over control in both the soils. In

loamy sand soil P uptake by grain increased significantly from 4.14 mg pot-1

in control to

9.53, 12.80, 17.72, 21.20 and 21.67 mg pot-1

resulting in an increase of 130, 209, 328, 412

and 423 per cent over control with P application @ 25, 50, 100, 200 and 400 mg kg-1

soil,

respectively.

Table 4.44: Effect of phosphorus and copper application on phosphorus uptake by grain

Copper

levels

(mg kg-1

soil)

Grain P uptake (mg pot-1

)

Phosphorus levels (mg kg-1

soil)

0 25 50 100 200 400 Mean

Loamy sand

0 6.96 11.58 17.29 27.01 30.43 37.64 21.82

5 5.58 13.76 15.83 21.98 28.63 25.14 18.49

10 3.86 11.64 16.72 19.99 24.08 22.03 16.38

20 0.16 1.15 1.37 1.91 1.66 1.88 1.35

Mean 4.14 9.53 12.80 17.72 21.20 21.67

CD (5%) P levels 1.93 Cu levels 1.58 P x Cu 3.87

Sandy loam

0 21.41 26.49 27.59 30.31 36.50 45.02 31.22

5 20.32 33.55 30.41 29.57 37.32 57.88 34.84

10 20.69 25.20 25.43 27.25 32.11 45.48 29.36

20 19.20 24.78 26.39 31.49 32.53 48.91 30.55

Mean 20.40 27.50 27.46 29.65 34.61 49.32

CD (5%) P levels 2.51 Cu levels 2.05 P x Cu 5.03

99

However, the differences in P uptake by grain observed with application of 200 and

400 mg P kg-1

soil were not significant. Similarly, in sandy loam soil P uptake by grain

increased significantly from 20.40 mg pot-1

in control to 27.50, 27.46, 29.65, 34.61 and 49.32

mg pot-1

which produced an increase of 34.8, 34.6, 45.3, 69.6 and 141.7 per cent over control

when P was applied @ 25, 50, 100, 200 and 400 mg kg-1

soil, respectively. The differences in

P uptake by grain with application of 25 and 50 mg P kg-1

soil were not significant. A

significant positive coefficient of correlation of P uptake by grain with grain yield in loamy

sand (r=0.918**) and sandy loam (r=0.933**) was observed (Table 4.49). The quadratic

response of grain P uptake to Olsen P indicated that in loamy sand (Fig. 4.19c, R2=0.907) P

uptake by grain reached a maximum level at 90 mg P kg-1

soil and then declined. In sandy

loam (Fig. 4.19d, R2=0.965) grain P uptake increased with increasing Olsen P.

Application of 5, 10 and 20 mg Cu kg-1

soil significantly decreased the mean P uptake

by grain from 21.82 mg pot-1

in control to 18.49, 16.38 and 1.35 mg pot-1

resulting in a

decrease of 15.2, 24.9 and 93.8 per cent over control, respectively. Whereas in sandy loam

soil mean P uptake by grain increased significantly by 11.5 per cent over control with

application of 5 mg Cu kg-1

soil and thereafter it decreased by 5.95 and 2.14 per cent over

control with application of10 and 20 mg Cu kg-1

soil, respectively. A significant reduction in

P uptake by wheat grain with application of 10 mg Cu kg-1

soil was also observed by Kumar

et al (2012). A linear negative relationship of P uptake by grain with DTPA-Cu in loamy sand

(Fig. 4.20c, R2=0.928) was observed. In sandy loam also grain P uptake decreased with

increasing Olsen P (Fig. 4.20d, R2= 0.204). A significant interaction effect of P and Cu levels

on grain uptake in loamy sand soil revealed that when each level of applied P was combined

with 20 mg Cu kg-1

soil, the grain P uptake was drastically reduced as compared to the control

which may be due to decrease in yield at this level (Table 4.21). Similarly, in sandy loam soil

grain P uptake significantly increased over control when 5 mg Cu kg-1

soil was combined

with either 25 or 400 mg P kg-1

soil.

4.7.3 Phosphorus uptake by straw

In loamy sand soil, P uptake by straw increased significantly from 3.49 mg pot-1

in

control to 6.91, 10.2, 15.13, 19.95 and 21.89 mg pot-1

whereas in sandy loam soil it

increased significantly from 13.81 mg pot-1

in control to 17.78, 21.18, 22.77, 29.94 and 47.46

mg pot-1

when P was applied @ 25, 50, 100, 200 and 400 mg kg-1

soil, respectively (Table

4.45). The corresponding values of per cent increase in straw P uptake over control with these

levels of applied P were 98,192,333,471 and 527 for loamy sand and 28.7, 53.3, 64.8, 116.7

and 243.6 for sandy loam soil, respectively. A significant positive coefficient of correlation of

P uptake by straw with straw yield in loamy sand (r=0.855**) and sandy loam (r=0.849**)

was observed (Table 4.49). The quadratic response of straw P uptake to Olsen P indicated that

in loamy sand (Fig. 4.19e, R2=0.965) P uptake by straw increased to the maximum up to a

100

level of 90 mg P kg-1

soil and thereafter it declined. In sandy loam (Fig. 4.19f, R2=0.992)

straw P uptake increased with increasing Olsen P. Phosphorus uptake by straw decreased

steeply beyond a level of 2.0 and 3.4 mg DTPA-Cu kg-1

soil in loamy sand (Fig. 4.20e,

R2=0.986) and sandy loam (Fig. 4.20f, R

2= 0.873), respectively.

Table 4.45: Effect of phosphorus and copper application on phosphorus uptake by straw

Copper

levels

(mg kg-1

soil)

Straw P uptake (mg pot-1

)

Phosphorus levels (mg kg-1

soil)

0 25 50 100 200 400 Mean

Loamy sand

0 5.13 6.91 10.84 16.11 30.03 36.50 17.59

5 3.85 10.05 15.27 19.68 20.10 27.50 16.08

10 4.07 8.67 13.07 19.96 25.65 20.97 15.40

20 0.92 2.01 1.64 4.78 4.04 2.59 2.66

Mean 3.49 6.91 10.20 15.13 19.95 21.89

CD (5%) P levels 2.80 Cu levels 2.28 P x Cu 5.60

Sandy loam

0 14.11 17.40 20.50 21.67 29.30 42.18 24.19

5 12.96 22.24 19.52 20.90 34.51 56.39 27.75

10 16.66 16.49 24.59 23.26 29.02 47.15 26.19

20 11.51 14.97 20.12 25.24 26.94 44.13 23.82

Mean 13.81 17.78 21.18 22.77 29.94 47.46

CD (5%) P levels 2.85 Cu levels 2.32 P x Cu 5.70

Application of 20 mg Cu kg-1

soil significantly decreased the mean P uptake by straw

to the tune of 84.6 per cent over control in loamy sand soil. It increased significantly by 14.7

per cent over control (24.19 mg pot-1

) when 5 mg Cu kg-1

soil was applied in sandy loam soil.

A significant reduction in P uptake by wheat straw with application of 10 mg Cu kg-1

soil was

also observed by Kumar et al (2012). The results of this study indicated an adverse affect of

Cu on P utilization by wheat.

A significant interaction effect of P and copper levels in loamy sand soil revealed that

P uptake of straw decreased significantly over control when 20 mg Cu kg-1

soil was combined

with each level of P. In sandy loam soil, maximum P uptake of 56.39 mg pot-1

was observed

when 5 mg Cu kg-1

soil was combined with 400 mg P kg-1

soil.

101

Fig. 4.19: Relationship of Olsen P in soil at harvest with P uptake by root (a, b), grain

(c, d) and straw (e, f)

y = -0.000x2 + 0.035x + 0.096R² = 0.939

0.00

0.20

0.40

0.60

0.80

1.00

1.20

1.40

1.60

1.80

0 25 50 75 100 125 150

Ro

ot

P u

pta

ke (

mg

po

t-1)

Olsen P (mg kg-1 soil)

y = -0.000x2 + 0.040x + 0.633R² = 0.971

0.80

1.00

1.20

1.40

1.60

1.80

2.00

2.20

2.40

0 25 50 75 100

Ro

ot

P u

pta

ke (

mg

po

t-1)

Olsen P (mg kg-1 soil)

y = -0.002x2 + 0.437x + 4.04R² = 0.907

0

5

10

15

20

25

30

0 25 50 75 100 125 150

Gra

in P

up

take

(m

g p

ot-1

)

Olsen P (mg kg-1 soil)

y = 0.318x + 19.49R² = 0.965

10

20

30

40

50

0 25 50 75 100

Gra

in P

up

take

(m

g p

ot-1

)

Olsen P (mg kg-1 soil)

y = -0.002x2 + 0.421x + 2.209R² = 0.965

0

5

10

15

20

25

0 25 50 75 100 125 150

Stra

w P

up

take

(m

g p

ot-1

)

Olsen P (mg kg-1 soil)

y = 0.393x + 10.62R² = 0.992

10

20

30

40

50

0 25 50 75 100

Stra

w P

up

take

(m

g p

ot-1

)

Olsen P (mg kg-1 soil)

(a) (b)

(c) (d)

102

Fig. 4.20: Relationship of DTPA-Cu in soil at harvest with P uptake by root (a, b), grain (c, d) and

straw (e, f)

y = -0.030x2 + 0.129x + 0.977R² = 0.996

0.40

0.60

0.80

1.00

1.20

0 1 2 3 4 5 6 7

Ro

ot

P u

pta

ke (

mg

po

t-1)

DTPA-Cu (mg kg-1 soil)

y = -0.005x2 + 0.017x + 1.642R² = 0.977

1.52

1.54

1.56

1.58

1.60

1.62

1.64

1.66

1.68

0 1 2 3 4 5 6 7

Ro

ot

P u

pta

ke (

mg

po

t-1)

DTPA-Cu (mg kg-1 soil)

y = -3.164x + 24.30R² = 0.928

0

5

10

15

20

25

0 1 2 3 4 5 6 7

Gra

in P

up

take

(m

g p

ot-1

)

DTPA-Cu (mg kg-1 soil)

y = -0.146x2 + 0.665x + 31.54R² = 0.204

29

30

31

32

33

34

35

36

0 1 2 3 4 5 6 7

Gra

in P

up

take

(m

g p

ot-1

)

DTPA-Cu (mg kg-1 soil)

y = -0.454x2 + 0.908x + 17.06R² = 0.986

0

2

4

6

8

10

12

14

16

18

20

0 1 2 3 4 5 6 7

Sraw

P u

pta

ke (

mg

po

t-1)

DTPA-Cu (mg kg-1 soil)

y = -0.432x2 + 2.951x + 22.16R² = 0.873

23

24

25

26

27

28

0 1 2 3 4 5 6 7

Stra

w P

up

take

(m

g p

ot-1

)

DTPA-Cu (mg kg-1 soil)

(b) (a)

(c) (d)

(e) (f)

103

4.7.4 Copper uptake by root

The data on Cu uptake by root is presented in Table 4.46. Phosphorus application

significantly increased the mean Cu uptake by root in loamy sand soil. It increased significantly

from 0.19 mg pot-1 in control to 0.324, 0.354, 0.575 and 0.545 mg pot

-1 which represented an

increase of 34, 70, 86, 200 and 184 per cent over control with application of 50, 100, 200 and 400

mg P kg-1 soil, respectively. The effect of P on Cu uptake by root in sandy loam soil was not

significant. A quadratic relationship of Olsen P with root Cu uptake revealed that the maximum

uptake of Cu by root in loamy sand soil was observed when the soil had 90 mg P kg-1 soil (Fig.

4.21a, R2=0.960) and afterwards it exhibited a downward trend. However in sandy loam, Cu

uptake by root decreased with increasing Olsen P (Fig. 4.21b, R2= 0.233).

Copper uptake by root increased significantly over control with increasing levels of

applied Cu in both the soils. It increased from 0.042 mg pot-1

in control to 0.351, 0.674 and

0.433 mg pot-1

in loamy sand soil and from 0.049 mg pot-1

in control to 0.058, 0.091 and

0.139 mg pot-1

in sandy loam soil with Cu application @ 5, 10 and 20 mg kg-1

soil. This

increase in Cu uptake by root over control was of the order of 735, 1504 and 930 per cent in

loamy sand and 18, 85 and 183 per cent in sandy loam with application of 5, 10 and 20 mg Cu

kg-1

soil, respectively. The relationship of DTPA-Cu with root Cu uptake indicated that in

loamy sand (Fig. 4.22a, R2=0.948) Cu uptake by root increased to the maximum up to a level

of 4.6 mg DTPA-Cu kg-1

soil and then it decreased sharply whereas in sandy loam Cu uptake

by root increased with increasing DTPA- Cu in soil (Fig. 4.22b, R2=0.937).

Table 4.46 Effect of phosphorus and copper application on root Cu uptake of wheat

Copper

levels

(mg kg-1

soil)

Root Cu uptake (mg pot-1

)

Phosphorus levels (mg kg-1

soil)

0 25 50 100 200 400 Mean

Loamy sand

0 0.030 0.037 0.039 0.049 0.050 0.046 0.042

5 0.190 0.283 0.385 0.384 0.410 0.440 0.351

10 0.180 0.306 0.462 0.564 1.340 1.170 0.674

20 0.370 0.400 0.411 0.421 0.480 0.510 0.433

Mean 0.190 0.256 0.324 0.354 0.570 0.540

CD (5%) P levels 0.069 Cu levels 0.056 P x Cu 1.39

Sandy loam

0 0.049 0.059 0.057 0.043 0.044 0.044 0.049

5 0.053 0.073 0.058 0.059 0.061 0.046 0.058

10 0.087 0.099 0.083 0.089 0.094 0.092 0.091

20 0.142 0.138 0.132 0.148 0.136 0.139 0.139

Mean 0.083 0.092 0.082 0.085 0.084 0.081

CD (5%) P levels NS Cu levels 0.006 P x Cu NS

104

4.7.5 Copper uptake by grain

In sandy loam soil, Cu uptake by grain increased significantly from 0.035 mg pot-1

in

control to 0.076, 0.082, 0.110, 0.133 and 0.142 mg pot-1

with application of 25, 50, 100, 200

and 400 mg P kg-1

soil which represented an increase of 117, 134, 214, 280 and 305 per cent

over control, respectively (Table 4.47). Similarly, in sandy loam soil Cu uptake by grain

increased significantly from 0.122 mg pot-1

in control to 0.150, 0.137, 0.144, 0.145 and 0.170

mg pot-1

with application of 25, 50, 100, 200 and 400 mg P kg-1

soil thereby resulting in an

increase of 22.9, 12.2, 18.0, 18.8 and 39.3 per cent over control, respectively. A significant

positive coefficient of correlation of Cu uptake by grain with grain yield in loamy sand

(r=0.763**) and sandy loam (r=0.441**) was observed (Table 4.49). The quadratic response

of grain Cu uptake to Olsen P indicated that in loamy sand (Fig. 4.21c, R2=0.891) Cu uptake

by grain reached a maximum level at 90 mg P kg-1

soil and then declined. In sandy loam (Fig.

4.21d, R2=0.704) grain Cu uptake increased with increasing Olsen P.

Table 4.47: Effect of phosphorus and copper application on copper uptake by grain

Copper

levels

(mg kg-1

soil)

Grain Cu uptake (mg pot-1

)

Phosphorus levels (mg kg-1

soil)

0 25 50 100 200 400 Mean

Loamy sand

0 0.045 0.067 0.066 0.095 0.105 0.115 0.082

5 0.047 0.108 0.115 0.146 0.186 0.187 0.131

10 0.048 0.116 0.135 0.173 0.223 0.248 0.157

20 0.026 0.014 0.013 0.026 0.020 0.019 0.016

Mean 0.035 0.076 0.082 0.110 0.133 0.142

CD (5%) P levels 0.011 Cu levels 0.009 P x Cu 0.022

Sandy loam

0 0.114 0.136 0.127 0.103 0.101 0.099 0.113

5 0.113 0.161 0.135 0.127 0.154 0.185 0.146

10 0.114 0.134 0.136 0.145 0.156 0.186 0.145

20 0.146 0.171 0.151 0.201 0.170 0.210 0.175

Mean 0.122 0.150 0.137 0.144 0.145 0.170

CD (5%) P levels 0.012 Cu levels 0.010 P x Cu 0.024

Grain Cu uptake due to Cu application in loamy sand soil increased significantly by

59.7 and 91.4 per cent over control (0.082 mg pot-1

) with application of 5 and 10 mg Cu kg-1

soil, respectively but it decreased significantly to 80.4 per cent over control when Cu was

applied @ 20 mg kg-1

soil. However, in sandy loam soil grain Cu uptake increased

105

significantly over control with increasing levels of applied Cu. It increased by 29.2, 28.3 and

54.8 per cent over control (0.113 mg pot-1

) with application of 5, 10 and 20 mg Cu kg-1

soil,

respectively. The quadratic response of grain Cu uptake to DTPA-Cu indicated that in loamy

sand (Fig. 4.22c, R2=0.982) Cu uptake by grain reached a maximum level at 3 mg DTPA-Cu

kg-1

soil and then declined sharply. In sandy loam (Fig. 4.22d, R2=0.910) grain Cu uptake

increased with increase in DTPA-Cu.

A significant interaction effect of P and Cu levels on grain Cu uptake was observed in

both the soils. It was observed that in light textured loamy sand soil, grain Cu uptake

decreased significantly over control at each level of applied P when Cu was applied @ 20 mg

kg-1

soil. But in sandy loam soil, the grain Cu uptake increased significantly over control

when 20 mg Cu kg-1

soil was combined with any level of applied P.

4.7.6 Copper uptake by straw

The data on Cu uptake by straw is presented in Table 4.48. Copper uptake by straw

increased significantly over control with increasing levels of applied P in loamy sand soil but

in sandy loam soil a decreasing trend was observed. In loamy sand soil it increased

significantly from 0.147 mg pot-1

in control to 0.262, 0.279, 0.316, 0.325 and 0.266 mg pot-1

with P application @ 25, 50, 100, 200 and 400 mg kg-1

soil which represented an increase of

78.2, 89.7, 115, 121, and 81 per cent over control, respectively. But in sandy loam soil, Cu

uptake by straw decreased significantly from 0.198 mg pot-1

in control to 0.158, 0.149 and

0.145 mg pot-1

with application of 100, 200 and 400 mg P kg-1

soil resulting in a decrease of

20.2, 24.7 and 26.7 per cent over control, respectively. The differences in Cu uptake observed

with application of 100, 200 and 400 mg P kg-1

soil were not significant. The quadratic

response of straw Cu uptake to Olsen P indicated that in loamy sand (Fig. 4.21e, R2=0.607)

Cu uptake by straw increased to the maximum up to a level of 75 mg P kg-1

soil and thereafter

it declined. In sandy loam (Fig. 4.21f, R2=0.944) straw Cu uptake decreased up to a level of

70 mg P kg-1

soil and then followed an upward trend. Cu uptake by straw increased up to a

level of 3.6 mg DTPA-Cu kg-1

soil in loamy sand (Fig. 4.22e, R2=0.854) and thereafter it

declined. In sandy loam (Fig. 4.22f, R2= 0.813) Cu uptake by straw increased with increase in

DTPA-Cu.

Copper application significantly increased Cu uptake by straw over control in both

the soils. In loamy sand soil it increased from 0.127 mg pot-1

in control to 0.265, 0.465 and

0.214 mg pot-1

and in sandy loam soil it increased from 0.115 mg pot-1

in control to 0.168,

0.151 and 0.252 mg pot-1

with application of 5, 10 and 20 mg Cu kg-1

soil, respectively. The

corresponding values of per cent increase in straw Cu uptake over control with these levels of

applied Cu were 101, 266 and 68.5 for loamy sand and 46.1, 31.3 and 119 for sandy loam

soil, respectively.

106

Table 4.48 Effect of phosphorus and copper application copper uptake by straw

Copper

levels

(mg kg-1

soil)

Straw Cu uptake (mg pot-1

)

Phosphorus levels (mg kg-1

soil)

0 25 50 100 200 400 Mean

Loamy sand

0 0.078 0.112 0.093 0.129 0.191 0.160 0.127

5 0.115 0.246 0.269 0.301 0.294 0.311 0.256

10 0.269 0.424 0.539 0.539 0.579 0.440 0.465

20 0.125 0.265 0.215 0.295 0.235 0.151 0.214

Mean 0.147 0.262 0.279 0.316 0.325 0.266

CD (5%) P levels 0.034 Cu levels 0.027 P x Cu 0.068

Sandy loam

0 0.139 0.146 0.120 0.095 0.100 0.092 0.115

5 0.168 0.231 0.206 0.116 0.139 0.146 0.168

10 0.159 0.167 0.160 0.131 0.137 0.152 0.151

20 0.326 0.247 0.238 0.291 0.221 0.189 0.252

Mean 0.198 0.198 0.181 0.158 0.149 0.145

CD (5%) P levels 0.026 Cu levels 0.021 P x Cu 0.052

Table 4.49: Linear coefficients of correlation of P, Cu and Fe uptake by root, grain

and straw with crop yield

Plant parameter Loamy sand Sandy loam

Root dry matter yield

P uptake 0.897**

0.908**

Cu uptake 0.153 -0.114

Fe uptake 0.991**

0.915**

Grain yield

P uptake 0.918**

0.933**

Cu uptake 0.763**

0.441

Fe uptake 0.936**

0.765**

Straw yield

P uptake 0.855**

0.849**

Cu uptake 0.265 -0.401

Fe uptake 0.940**

0.826**

*Significant at 5%

**Significant at 1%

107

Fig. 4.21: Relationship of Olsen P in soil at harvest with Cu uptake by root (a, b),

grain (c, d) and straw (e, f)

y = -5E-05x2 + 0.009x + 0.142R² = 0.960

0.1

0.2

0.3

0.4

0.5

0.6

0.7

0 25 50 75 100 125 150

Ro

ot

Cu

up

take

(m

g p

ot-1

)

Olsen P (mg kg-1 soil)

y = -6E-05x + 0.086R² = 0.233

0.080

0.082

0.084

0.086

0.088

0.090

0.092

0.094

0 25 50 75 100

Ro

ot

Cu

up

take

(m

g p

ot-1

)

Olen P (mg kg soil-1)

y = -1E-05x2 + 0.002x + 0.039R² = 0.891

0.00

0.02

0.04

0.06

0.08

0.10

0.12

0.14

0.16

0 25 50 75 100 125 150

Gra

in C

u u

pta

ke (

mg

po

t-1)

Olsen P (mg kg-1 soil)

y = 0.000x + 0.128R² = 0.704

0.10

0.12

0.14

0.16

0.18

0.20

0 25 50 75 100

Gra

in C

u u

pta

ke (

mg

po

t-1)

Olsen P (mg kg-1 soil)

y = -3E-05x2 + 0.004x + 0.180R² = 0.607

0.10

0.15

0.20

0.25

0.30

0.35

0.40

0 25 50 75 100 125 150

Staw

Cu

up

take

(m

g p

ot-1

)

Olsen P (mg kg-1 soil)

y = 2E-05x2 - 0.002x + 0.233R² = 0.944

0.12

0.14

0.16

0.18

0.20

0.22

0 25 50 75 100

Stra

w C

u u

pta

ke (

mg

po

t-1)

Olsen P (mg kg-1 soil)

(a)

(b)

(c) (d)

(e) (f)

108

Fig. 4.22: Relationship of DTPA-Cu in soil at harvest with Cu uptake by root (a, b),

grain (c, d) and straw (e, f)

y = -0.035x2 + 0.313x - 0.046R² = 0.948

0

0.1

0.2

0.3

0.4

0.5

0.6

0.7

0.8

0 1 2 3 4 5 6 7

Ro

ot

Cu

Up

take

(mg

po

t-1)

DTPA-Cu (mg kg-1 soil)

y = 0.017x + 0.024R² = 0.937

0.00

0.02

0.04

0.06

0.08

0.10

0.12

0.14

0.16

0 1 2 3 4 5 6 7

Ro

ot

Cu

Up

take

(m

g p

ot-1

)

DTPA-Cu (mg kg-1 soil)

y = -0.009x2 + 0.057x + 0.067R² = 0.982

0.00

0.04

0.08

0.12

0.16

0.20

0 1 2 3 4 5 6 7

Gra

in C

u u

pta

ke (

mg

po

t-1)

DTPA-Cu (mg kg-1 soil)

y = 0.010x + 0.107R² = 0.910

0.10

0.12

0.14

0.16

0.18

0 1 2 3 4 5 6 7

Gra

in C

u u

pta

ake

(m

g p

ot-1

)

DTPA-Cu (mg kg-1 soil)

y = -0.023x2 + 0.178x + 0.067R² = 0.854

0

0.1

0.2

0.3

0.4

0.5

0 1 2 3 4 5 6 7

Stra

w C

u u

pta

ke (

mg

po

t-1)

DTPA-Cu (mg kg-1 soil)

y = 0.022x + 0.091R² = 0.813

0.10

0.15

0.20

0.25

0.30

0 1 2 3 4 5 6 7

Stra

w C

u u

pta

ke (

mg

po

t-1)

DTPA-Cu (mg kg-1 soil)

(a) (b)

(c) (d)

(e) (f)

109

4.7.7 Iron uptake by root

In loamy sand soil, mean root Fe uptake increased significantly from 1.56 mg pot-1

in

control to 2.03, 2.34, 2.58, 2.80 and 2.65 mg pot-1

which represented an increase of 30.1, 50.0,

65.3, 79.4 and 69.8 per cent over control with an application of 25, 50, 100, 200 and 400 mg

P kg-1

soil, respectively (Table 4.50). In sandy loam soil, it increased significantly from 3.39

mg pot-1

in control to 4.29, 4.20, 4.74, 5.12 and 4.99 mg pot-1

resulting in an increase of 26.5,

23.8, 39.8, 51.0 and 47.1 per cent over control with the same levels of applied P, respectively.

The decrease in mean root uptake with the highest level of applied P in both the soils was not

significant over the root uptake recorded with the application of 200 mg P kg-1

soil. A

significant positive coefficient of correlation of Fe uptake by root with root yield in loamy

sand (r=0.991**) and sandy loam (r=0.915**) was observed (Table 4.49). A quadratic

relationship of Olsen P with root Fe uptake revealed that uptake of Fe by root in loamy sand

(Fig. 4.23a, R2=0.827) increased to the maximum up to a level of 90 mg P kg

-1 soil whereas

the corresponding value for sandy loam was about 70 mg P kg-1

soil (Fig. 4.23b, R2=0.887)

and afterwards the uptake of Fe by root decreased in both soils.

Table 4.50: Effect of phosphorus and copper application on iron uptake by root

Copper

levels

(mg kg-1

soil)

Root Fe uptake (mg pot-1

)

Phosphorus levels (mg kg-1

soil)

0 25 50 100 200 400 Mean

Loamy sand

0 1.91 2.61 2.74 3.25 3.55 3.20 2.88

5 1.63 2.37 2.92 2.98 3.18 3.14 2.70

10 1.50 1.91 2.45 2.78 3.26 3.07 2.49

20 1.21 1.24 1.25 1.31 1.22 1.18 1.23

Mean 1.56 2.03 2.34 2.58 2.80 2.65

CD (5%) P levels 0.257 Cu levels 0.210 P x Cu 0.514

Sandy loam

0 3.88 4.55 4.36 4.61 5.04 5.06 4.58

5 3.69 4.97 4.17 5.18 5.46 5.08 4.76

10 3.27 3.99 4.37 4.62 5.34 4.81 4.40

20 2.73 3.65 3.93 4.54 4.65 5.00 4.08

Mean 3.39 4.29 4.20 4.74 5.12 4.99

CD (5%) P levels 0.349 Cu levels 0.285 P x Cu NS

In loamy sand soil, mean root Fe uptake decreased significantly from 2.88 mg pot-1

in control to 2.49 and 1.23 mg pot-1

thereby resulting in a decrease of 13.5 and 57.2 per cent

over control with application of 10 and 20 mg Cu kg-1

soil, respectively. A significant

110

decrease of 10.9 per cent in root Fe uptake in sandy loam soil was observed only when Cu

was applied @ 20 mg kg-1

soil. A significant interaction effect of P and Cu levels on root Fe

uptake in loamy sand soil revealed that Fe uptake by root decreased significantly over control

when 20 mg Cu kg-1

soil was applied at each level of P. The quadratic relationship of DTPA-

Cu with root Fe uptake indicated that Fe uptake by root in loamy sand (Fig. 4.24a, R2=0.996)

as well as sandy loam (Fig. 4.24b, R2=0.900) decreased sharply when soil had >2 mg DTPA-

Cu kg-1

soil.

4.7.8 Iron uptake by grain

The data on the effect of P and Cu levels on Fe uptake by grain is presented in Table

4.51. In loamy sand soil, Fe uptake by grain increased significantly from 0.161 mg pot-1

in

control to 0.332, 0.372, 0.467, 0.582 and 0.617 mg pot-1

thus resulting in an increase of 106,

131, 190, 261 and 283 per cent over control with application of P @ 25, 50, 100, 200 and 400

mg kg-1

soil, respectively. A significant positive coefficient of correlation of Fe uptake by

grain with grain yield in loamy sand (r=0.936**) and sandy loam (r=0.765**) was observed

(Table 4.49).The quadratic response of grain Fe uptake to Olsen P indicated that in loamy

sand (Fig. 4.23c, R2=0.895) Fe uptake by grain reached a maximum level at 90 mg P kg

-1 soil

and then declined. In sandy loam (Fig. 4.23d, R2=0.730) grain Fe uptake increased with

increase in Olsen P. In sandy loam soil, the effect of P and Cu levels, and their interaction on

Fe uptake was not significant.

Table 4.51: Effect of phosphorus and copper application on iron uptake by grain

Copper

levels

(mg kg-1

soil)

Grain Fe uptake (mg pot-1

)

Phosphorus levels (mg kg-1

soil)

0 25 50 100 200 400 Mean

Loamy sand

0 0.276 0.458 0.547 0.897 0.912 0.924 0.669

5 0.219 0.475 0.479 0.446 0.591 0.845 0.508

10 0.145 0.361 0.428 0.474 0.167 0.654 0.471

20 0.070 0.036 0.035 0.050 0.059 0.048 0.039

Mean 0.161 0.332 0.372 0.467 0.582 0.617

CD (5%) P levels 0.055 Cu levels 0.045 P x Cu 0.110

Sandy loam

0 0.924 0.962 0.992 0.941 0.900 1.070 0.965

5 0.911 1.070 0.904 0.844 10.400 1.270 1.009

10 0.965 0.998 0.853 0.930 0.987 1.100 0.972

20 0.869 0.973 0.902 1.010 0.959 0.990 0.951

Mean 0.917 1.002 0.913 0.932 0.973 1.110

CD (5%) P levels 0.081 Cu levels NS P x Cu NS

111

Grain Fe uptake decreased significantly by 24.1, 29.5 and 94.1 per cent over control

(0.669 mg pot-1

) in loamy sand soil with application of 5, 10 and 20 mg Cu kg-1

soil,

respectively. But a significant interaction effect of P and Cu levels on grain Fe uptake in

loamy sand soil was observed and it decreased significantly over control when 20 mg Cu kg-1

soil was combined with each level of applied P. A linear negative relationship of Fe uptake by

grain with DTPA-Cu in loamy sand (Fig. 4.24c, R2=0.947) was observed. In sandy loam,

grain Fe uptake increased up to a level of 3 mg DTPA-Cu kg-1

soil and then followed a

decreasing trend (Fig. 4.24d, R2= 0.688).

4.7.9 Iron uptake by straw

The data on Fe uptake by straw is presented in Table 4.52. Iron uptake by straw

increased significantly from 1.56 mg pot-1

in control to 2.88, 2.92, 3.71, 4.81 and 4.61 mg

pot-1

in loamy sand soil and from 5.15 mg pot-1

in control to 6.44, 6.63, 6.72, 9.50 and

11.95 mg pot-1

in sandy loam soil when P was applied @ 25, 50, 100, 200 and 400 mg kg-

1 soil, respectively. The corresponding values of per cent increase in straw Fe uptake over

control with these levels of applied P were 84.6, 87.1, 137.8, 208 and 195 for loamy sand

and 25, 28.7, 30.4, 84.4 and 132 for sandy loam soil, respectively. The differences in

straw Fe uptake observed with 25 and 50 mg P kg-1

soil were not significant in both the

soils.

Table 4.52: Effect of phosphorus and copper application on iron uptake by straw

Copper

levels

(mg kg-1

soil)

Straw Fe uptake (mg pot-1)

Phosphorus levels (mg kg-1

soil)

0 25 50 100 200 400 Mean

Loamy sand

0 2.71 3.98 3.91 5.31 6.98 7.39 5.05

5 1.84 4.02 3.82 4.69 5.95 5.61 4.32

10 1.45 2.98 3.46 3.79 5.57 4.77 3.67

20 0.22 0.54 0.52 1.04 0.75 0.66 0.62

Mean 1.56 2.88 2.92 3.71 4.81 4.61

CD (5%) P levels 0.55 Cu levels 0.44 P x Cu 1.10

Sandy loam

0 5.39 5.25 5.85 6.40 9.47 10.67 7.17

5 4.53 8.56 8.14 5.89 10.96 14.47 8.77

10 5.70 5.23 6.61 5.98 8.41 11.08 7.17

20 4.98 6.72 5.92 8.61 9.17 11.50 7.81

Mean 5.15 6.44 6.63 6.72 9.50 11.95

CD (5%) P levels 1.24 Cu levels 1.01 P x Cu NS

112

Fig. 4.23: Relationship of Olsen P in soil at harvest with Fe uptake by root (a, b), grain

(c, d) and straw (e, f)

y = -0.000x2 + 0.032x + 1.617R² = 0.827

1.00

1.50

2.00

2.50

3.00

0 25 50 75 100 125 150

Ro

ot

Fe u

pta

ke (

mg

po

t-1)

Olsen P (mg kg-1 soil)

y = -0.000x2 + 0.081x + 2.727R² = 0.887

3.00

3.50

4.00

4.50

5.00

5.50

0 25 50 75 100

Ro

ot

Fe u

pta

ke (

mg

po

t-1)

Olsen P (mg kg-1 soil)

y = -5E-05x2 + 0.009x + 0.178R² = 0.895

0

0.1

0.2

0.3

0.4

0.5

0.6

0.7

0 25 50 75 100 125 150

Gra

in F

e u

pta

ke (

mg

po

t-1)

Olsen P (mg kg-1 soil)

y = 0.002x + 0.895R² = 0.730

0.85

0.95

1.05

1.15

0 25 50 75 100

Gra

in F

e u

pta

ke (

mg

po

t-1)

Olsen P (mg kg-1 soil)

y = -0.000x2 + 0.077x + 1.573R² = 0.903

1

2

3

4

5

6

0 25 50 75 100 125 150

Stra

w F

e u

pta

ke (

mg

po

t-1)

Olsen P (mg kg-1 soil)

y = -0.000x2 + 0.143x + 3.591R² = 0.969

4

6

8

10

12

0 25 50 75 100

Stra

w F

e u

pta

ke (

mg

po

t-1)

Olsen P (mg kg-1 soil)

(a) (b)

(c) (d)

(e) (f)

113

Fig. 4.24: Relationship of DTPA-Cu in soil at harvest with Fe uptake by root (a, b),

grain (c, d) and Straw (e, f)

y = -0.04x2 + 0.025x + 2.857R² = 0.996

1.00

1.50

2.00

2.50

3.00

0 1 2 3 4 5 6 7

Ro

ot

Fe u

pta

ke (

mg

po

t-1)

DTPA-Cu (mg kg-1 soil)

y = -0.033x2 + 0.139x + 4.518R² = 0.900

4

4.1

4.2

4.3

4.4

4.5

4.6

4.7

4.8

0 1 2 3 4 5 6 7

Ro

ot

Fe u

pta

ke (

mg

po

t-1)

DTPA-Cu (mg kg-1 soil

y = -0.095x + 0.715R² = 0.947

-0.10

0.00

0.10

0.20

0.30

0.40

0.50

0.60

0.70

0 1 2 3 4 5 6 7Gra

in F

e u

pta

ke (

mg

po

t-1)

DTPA-Cu (mg kg-1 soil)

y = -0.004x2 + 0.028x + 0.948R² = 0.688

0.94

0.96

0.98

1.00

1.02

0 1 2 3 4 5 6 7

Gra

in F

e u

pta

ke (

mg

po

t-1)

DTPA-Cu (mg kg-1 soil)

y = -0.076x2 - 0.145x + 5.033R² = 0.997

0

1

2

3

4

5

6

0 1 2 3 4 5 6 7

Stra

w F

e u

pta

ke (

mg

po

t-1)

DTPA-Cu (mg kg-1 soil)

y = -0.063x2 + 0.485x + 7.061R² = 0.112

7.00

7.50

8.00

8.50

9.00

0 1 2 3 4 5 6 7

Stra

wFe

up

take

(m

g p

ot-1

)

DTPA-Cu (mg kg-1 soil)

(a) (b)

(c) (d)

(e) (f)

114

Straw Fe uptake decreased significantly from 5.05 mg pot-1

in control to 4.32, 3.67

and 0.62 mg pot-1

when Cu was applied @ 5, 10 and 20 mg kg-1

soil in loamy sand soil

resulting in a decrease of 14.4, 27.3 and 87.7 per cent over control, respectively. In sandy

loam soil, Fe uptake by straw increased significantly by 22.3 per cent over control (7.17 mg

pot-1

) with application of 5 mg Cu kg-1

soil. A significant positive coefficient of correlation

of Fe uptake by straw with straw yield in loamy sand (r=0.940**) and sandy loam

(r=0.826**) was observed (Table 4.49). The quadratic response of straw Fe uptake to Olsen

P indicated that in loamy sand (Fig. 4.23e, R2=0.903) Fe uptake by straw increased to the

maximum up to a level of 90 mg P kg-1

soil and thereafter it declined. In sandy loam (Fig.

4.23f, R2=0.969) straw Fe uptake increased with increase in Olsen P. Iron uptake by straw

decreased sharply beyond a level of 2.0 mg DTPA-Cu kg-1

soil in loamy sand (Fig. 4.24e,

R2=0.997). In sandy loam Fe uptake by straw was poorly related with DTPA-Cu (Fig. 4.24f,

R2= 0.112).

A significant interaction effect of P and Cu levels on straw Fe uptake revealed that at

each level of applied P, the Fe uptake by straw decreased over control when Cu was applied

@ 20 mg kg-1

soil. But in sandy loam soil this interaction effect was not significant.

The data on linear coefficients of correlation of chemical pools of Cu with Cu

uptake by root, grain and straw revealed that in loamy sand soil amorphous oxides Cu

(r=0.527**

) and residual Cu (r=0.408*) was significantly positively correlated with Cu

uptake by root. However, Cu uptake by grain was significantly negatively correlated with

almost all the chemical pools of Cu (Table 4.53). But in sandy loam soil, Cu uptake by

root, grain and straw was significantly positively correlated with various pools of Cu

studied. Iron uptake by root, grain, and straw in loamy sand soil was significantly

negatively correlated with all the pools of Cu. In sandy loam, Fe uptake by root was

significantly negatively correlated with amorphous oxides Cu (r=-0.420*) and Crystalline

oxides Cu (r=-0.453*). However, organically bound Cu was significantly positively

correlated with grain (r=0.456*) and straw Fe uptake (=0.629**). DTPA-Fe was also

significantly positively correlated with Fe uptake by root, grain and straw. Phosphorus

uptake by root, grain and straw was negatively correlated with all the pools of Cu in

loamy sand. However, organically bound Cu was significantly positively correlated with

P uptake by grain (r=0.586**

) and straw (r=0.589**

) in sandy loam soil. Phosphorus

uptake by root, grain and straw was significantly positively correlated with DTPA-Zn, Mn

and Fe in both soils (Table 4.53).

115

Table 4.53: Linear coefficients of correlation of P, Cu and Fe uptake by root, grain and straw with chemical pools of Cu and DTPA-

extractable micronutrient cations

Phosphorus uptake Copper uptake Iron uptake

Root Grain Straw Root Grain Straw Root Grain Straw

Loamy sand

EXCH-Cu -0.495* -0.746

** -0.637

** 0.134 -0.663

** -0.123 -0.820

** -0.782

** -0.814

**

SAD-Cu -0.451* -0.697

** -0.592

** 0.183 -0.601

** -0.103 -0.761

** -0.727

** -0.761

**

CARB-Cu -0.299 -0.645**

-0.502* 0.358 -0.446

* 0.101 -0.708

** -0.693

** -0.718

**

MnOX-Cu -0.419* -0.675

** -0.571

** 0.168 -0.561

** 0.012 -0.736

** -0.717

** -0.735

**

AMPOX-Cu -0.289 -0.636**

-0.487* 0.527

** -0.275 0.387 -0.686

** -0.696

** 0.721

**

CRYOX-Cu -0.445* -0.717

** -0.586

** 0.370 -0.441

* 0.243 -0.740

** -0.739

** -0.769

**

OM-Cu -0.432* -0.610

** -0.551

** 0.155 -0.468

* -0.084 -0.609

** -0.618

** -0.643

**

RES-Cu -0.363 -0.690**

-0.554**

0.408* -0.407

* 0.165 -0.760

** -0.761

** -0.772

**

DTPA-Cu -0.354 -0.682**

-0.539**

0.470* -0.365 0.212 -0.735

** -0.782

** -0.761

**

DTPA-Zn 0.573**

0.397* 0.529

** 0.425

* 0.402 0.152 0.269 0.356 0.339

DTPA-Mn 0.569**

0.389* 0.523

** 0.429

* 0.397 0.154 0.261 0348 0.330

DTPA-Fe 0.577**

0.403 0.535**

0.422* 0.405

* 0.150 0.275 0.363 0.346

Sandy loam

EXCH-Cu -0.096 -0.091 -0.078 0.926**

0.597**

0.816**

-0.417* -0.110 -0.003

SAD-Cu -0.110 -0.120 -0.097 0.951**

0.605**

0.815**

-0.408* -0.133 -0.005

MnOX-Cu 0.026 0.057 0.075 0.926**

0.656**

0.642**

-0.223 -0.099 0.112

AMPOX-Cu -0.162 -0.150 -0.114 0.929**

0.612**

0.791**

-0.420* -0.142 -0.050

CRYOX-Cu -0.200 -0.234 -0.180 0.907**

0.537**

0.791**

-00.453* -0.222 -0.141

OM-Cu 0.352 0.586**

0.589**

0.510**

0.763**

0.286 0.185 0.456* 0.629

**

RES-Cu -0.035 -0.035 0.002 0.950**

0.690**

0.720**

-0.220 -0.095 0.063

DTPA-Cu -0.159 -0.144 -0.095 0.936**

0.616**

0.749**

-0.372 -0.160 -0.047

DTPA-Zn 0.850**

0.916**

0.946**

0.072 0.480* -0.198 0.560

** 0.612

** 0.870

**

DTPA-Mn 0.846**

0.913**

0.944**

0.084 0.488* -0.187 0.555

** 0.609

** 0.868

**

DTPA-Fe 0.852**

0.918**

0.948**

0.061 0.473* -0.206 0.565

** 0.614

** 0.871

**

*Significant at 5%

**Significant at 1%

CHAPTER V

SUMMARY

As only one- third of the applied phosphorus (P) is taken up by the current crop, it

may lead to the buildup of P in plough layer of soils over a period of time. Phosphorus

accumulation in soils can cause various types of nutrient interactions in soils and plants.

Depending upon the nutrient supply, the interactions between nutrients can either induce

deficiencies and/or toxicities that can modify the growth response. In the present investigation

a pot experiment was conducted to study the effect of P and Cu application on transformation

and availability of Cu to wheat. The two soils used were i) calcareous loamy sand (ls) Typic

Ustipssament (pH 8.1, EC 0.37 dS m-1

, OC 0.15%, available P 8.5, CaCO3 0.13%, DTPA-Cu

0.18 and DTPA-Fe 3.2 mg kg-1

soil) and ii) non calcareous sandy loam (sl) Typic Haplustept

(pH 6.5, EC 0.295 dS m-1

, OC 0.38%, available P 12.65, DTPA-Cu 1.05 and DTPA-Fe 51.6

mg kg-1

soil). Six levels of P (0, 25, 50, 100, 200 and 400 mg P kg-1

soil) as monocalcium

phosphate monohydrate and four levels of Cu (0, 5, 10 and 20 mg Cu kg-1

soil) as Cu-EDTA

were applied in all possible combinations to eight kg of each soil per pot with three

replications. Wheat (cv HD 2967) was grown, soil, root, grain and straw samples were

collected at maturity. Soil samples were analysed for pH, available P, DTPA-Cu, DTPA-Fe

and chemical pools of Cu viz. exchangeable (EXCH), specifically adsorbed (SAD), carbonate

bound (CARB), manganese oxides bound (MnOX), amorphous oxides bound (AMPOX),

crystalline oxides bound (CRYOX), organically bound (OM) and residual (RES) and plant

samples were analysed for total P, Cu and Fe.

The quadratic relationship of levels of applied P with soil pH indicated that in ls soil

pH increased but in sl soil it decreased up to a point when soil had about160 mg P kg-1

soil and

thereafter the trend was reversed. Application of 20 mg Cu kg-1

soil decreased soil pH

significantly from 8.06 in control to 7.87 in ls soil and from 6.12 in control to 6.01 in sl soil.

Soil EC increased up to a point when it contained about 300 mg P kg-1

soil and thereafter it

exhibited a downward trend in both the soils. The EC increased significantly with Cu

application in both the soils. In ls soil, the availability of P increased from 8.32 mg kg-1

soil in

control to a maximum of 130.6 mg P kg-1

soil in ls and from 12.62 mg kg-1

soil in control to

94.62 mg kg-1

soil in sl soil with the highest level of P. More amount of applied P remained in

solution form in light textured ls soil as compared to sl soil.

DTPA-Cu decreased from 3.17 mg kg-1

soil in control to 2.84 mg kg-1

soil with an

application of 100 mg P kg-1

soil in ls but in sl soil it decreased significantly from 3.65 mg kg-

1 soil in control to 3.31 mg kg

-1 soil with application of 400 mg P kg

-1 soil. Application of Cu

significantly increased DTPA-Cu in both the soils. In ls soil, DTPA-Fe decreased

significantly from 3.25 mg kg-1

soil in control to 3.04 mg kg-1

soil with application of 200 mg

117

P kg-1

soil. However, in sl soil a significant decrease in DTPA-Fe was observed with

application of 100, 200 and 400 mg P kg-1

soil. A significant decrease in DTPA- Fe in

calcareous ls soil was observed when Cu was applied @ 20 mg kg-1

soil as compared to

application of 10 mg Cu kg-1

soil. In both the soils, DTPA-extractable Zn, Cu, Fe and Mn

decreased with increase in soil pH.

When Cu is applied to soil though external sources many chemical reactions like

adsorption, precipitation and complexation take place. The percent recovery of added Cu @

20 mg kg-1

soil at different levels of applied P in a particular chemical pool was calculated by

taking the difference of the content of that fraction in Cu treated pot and the corresponding no

Cu pots. In ls soil only 3.27 and 2.93 per cent and in sandy loam soil 2.81 and 1.44 per cent of

applied Cu @ 20 mg kg-1 soil entered in to exchangeable and specifically adsorbed fractions,

respectively which are considered to be the most plant available forms. A major portion (43-45%)

of the added Cu entered in to residual mineral fraction followed by amorphous oxides (32-34%)

and crystalline oxides (7.1-12.7%) in both the soils. It is due to the fact that Cu has a strong

tendency to associate with the crystalline structures of the minerals and organic ligands. About

6.08 per cent of added Cu also entered in to carbonate bound fraction in alkaline calcareous loamy

sand soil because metals can also be associated with soil carbonates. Only 1.29 and 2.47 per cent

of added Cu entered in to OM-Cu in ls and sl soils, respectively. Mean values of per cent recovery

of applied Cu @ 20 mg kg-1 soil in various chemical pools of Cu were used to study the

distribution and movement of Cu in to its various chemical pools in the presence and absence of

applied P. The data indicated that in ls soil, with P application plant available forms like EXCH-

Cu and SAD-Cu decreased but CARB-Cu increased and most of the applied Cu redistributed in to

the various chemical pools and a very little of it got in to RES-Cu.. In sl soil, applied P decreased

EXCH-Cu, AMPOX-Cu and CRYOX-Cu but increased the Cu content in OM-Cu and RES-Cu.

Mean root dry matter, grain and straw yield increased significantly over control with

increasing levels of applied P in both the soils. But the yield of each plant part studied,

decreased significantly over control with increasing levels of applied Cu in ls calcareous soil.

But in sl soil a maximum increase of 62.5, 74.3 and 63.7 per cent in root, grain and straw

yield was observed when 400 mg P was applied in combination with 5 mg Cu kg-1

soil,

respectively. A significant negative interaction of P and Cu levels on root dry matter yield

indicated that in ls soil higher levels of applied Cu may prove toxic for the growth of roots as

more amounts of applied Cu remained in available form as compared to medium textured soil

as indicated by the higher content of DTPA-Cu in ls soil.

In alkaline calcareous ls soil, severe Fe chlorosis of leaves was observed when

each level of applied P was combined with 20 mg Cu kg-1

soil and the growth of the

crop was drastically reduced as compared to control. But in the absence of applied Cu,

118

deficiency symptoms of Fe were not observed even with the highest level of applied

P. This was confirmed by a significant decrease in SPAD values of the leaves as an

indicator of chlorophyll content at 65 days after sowing. The specific activity of super

oxide dismutase increased significantly by 84 per cent over no Cu application when

20 mg Cu kg-1

soil was applied which may be considered as a circumstantial evidence

of enhanced production of reactive oxygen species as a defensive mechanism of

wheat plants against the toxic effects of Cu.

Path coefficient analysis of grain yield of wheat against various pools of Cu revealed

that in ls soil DTPA-Cu, CARB-Cu and AMPOX-Cu produced significant direct effects on

grain yield of wheat. About 21.9, 45.5 and 18.6 percent variation in grain yield was directly

controlled by DTPA-Cu, CARB-Cu and AMPOX-Cu, respectively. Exchangeable Cu and

SAD-Cu, the most plant available pools controlled a variation of 17.8 and 17.0 per cent in

grain yield indirectly through DTPA-Cu. An indirect variation of 39.8 and 13.3 per cent, in

grain yield was controlled by EXCH-Cu through CARB-Cu and AMPOX-Cu and 37.9 and

12.7 per cent by SAD-Cu through CARB-Cu and AMPOX-Cu, respectively. The results

indicated the importance of DTPA-Cu, EXCH-Cu, SAD-Cu and AMPOX-Cu effecting the

grain yield of wheat in ls calcareous soil but in sl acidic soil oxides and organic matter forms

of Cu played an important role either directly or indirectly in effecting the grain yield.

Phosphorus concentration in root, grain and straw increased significantly with

increasing levels of applied P in both the soils. Also Cu concentration in each plant part

increased with graded levels of applied Cu in both the soils but in sl soil Cu concentration in

root, grain and straw decreased significantly with increasing levels of applied P. In ls

calcareous soil CARB-Cu played a significant role in effecting the Cu concentration of wheat

grain. All the different pools of Cu studied together controlled an indirect variation of 44.4

per cent in grain Cu concentration through AMPOX-Cu. Like that on grain yield, the oxides

and residual Cu played an important role in controlling grain Cu concentration in sl soil.

Accumulation of Cu in roots decreased Fe absorption by roots. The Cu: Fe

concentration ratio in root, grain and straw at which severe Fe chlorosis of leaves was

observed varied from 0.304 to 0.429, 0.384 to 0.525 and 0.231 to 0.557, respectively.

Upper critical levels of Cu associated with 50 per cent reduction from maximum

yield, which are generally considered to be toxic for plants were determined by using the data

of all the 24 treatment combinations for both the soils taken together (n=48). The response of

relative dry matter yield of root grain and straw to DTPA extractable Cu in soil at harvest

was observed to be quadratic with significant values of coefficients of determination. A

content of 6.56, 5.39 and 5.37 mg DTPA-extractable Cu kg-1

soil produced 50 per cent

reduction from the maximum yield of root, grain and straw, respectively which may be

119

considered as the upper critical values for wheat. A Cu concentration of 436, 11.04 and

19.33µg g-1

reduced the yield of root, grain and straw, respectively by 50 per cent from the

maximum yield which may be considered as the upper critical levels of Cu in root, grain and

straw of wheat.

Mean P and Fe uptake by root, grain and straw increased with increasing levels of

applied P but decreased with Cu application in both the soils. However, Cu uptake by each

plant part increased significantly with graded levels of applied Cu. In ls soil P and Fe uptake

by root, grain and straw was significantly negatively correlated with most of the chemical

pools of Cu in soil.

120

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132

VITA

Name of the student : Harpreet Kaur

Father’s name : S. Jatinder Singh

Mother’s Name : Smt. Manjeet Kaur

Nationality : Indian

Date of Birth : 05 December, 1989

Permanent Address : Punia Colony, St.No.10, H.No.181, Sangrur-

148001

EDUCATIONAL QUALIFICATIONS

Bachelor’s degree : B.Sc. (Hons.) Agriculture

University and year of award : Punjab Agricultural University, Ludhiana,

(2012)

OCPA : 7.11/10.00

Master’s degree : M. Sc. (Soil Science)

University and year of Award : Punjab Agricultural University, Ludhiana,

(2014)

OCPA : 7.88/10.00

Title of Master’s Thesis : Transformation and availability of copper to

wheat (Triticum aestivum L) as influenced by

phosphorus fertilization