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MANOHARBHAI PATEL INSTITUTE OF

ENGINEERING AND TECHNOLOGY

GONDIA

F O R W A R D I N G L E T T E R

Forwarded herewith to the Rashtrasant Tukdoji Maharaj Univesity, Nagpur, and the

dissertation

TREATMENT OF DOMASTIC WASTE WATER BY

PYTOREMEDATION (LEMNA PLANT)

Submitted by- Sandeep P. Ajmire , in partial fulfillment of the award of the

degree of Master of Technology in Environmental Engineering.

Prof. A. L. Nashine Prof. Dr.S.S. Rathor

Head of department Principal

Dept. of Civil Engg. MIET Gondia

MIET , Gondia


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MANOHARBHAI PATEL INSTITUTE OF

ENGINEERING AND TECHNOLOGY

GONDIA

C E R T I F I C A T E

This is to certify that dissertation entitled

TREATMENT OF DOMASTIC WASTE WATER BY

PHYTOREMEDATION (LEMNA)

Submitted by Sandip P.Ajmire , in practical fulfillment of the requirement for the

award of Degree of Master of Technology in Environmental Engineering to The

Rashtrasant Tukdoji Maharaj University, Nagpur , is bonafide research work carried

out under my supervision and guidance. The work embodied in this dissertation has

not submitted previously for the award of any degree or diploma.

Prof . A.M. Deshpande Prof.AL Nashine

Supervisor Head Of Department

Dept. Of Civil Engineering Dept. Of Civil Engineering

MIET GONDIA MIET GONDIA


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A C K N O W L E D G E M E N T

I express my profound gratitude towards Prof. A.M. Deshpande ,Lecturer ,

Department of Civil Engineering. MIET Gondia, for this able guidance.

I am extremely Grateful to Hon President Mr. Choudhari &

CEO A.V. Dhoke, Municipal Council Gadchiroli . Mr .M.G. Nisal , Lab Asst .

Environmental Engineering Lab MITE ,Gondia , without whose help the project

might have been completed. Mr. S.P. Waghmare, Executive Engineer Jeewan

Pradhikarn Gadchiroli & his technical and non technical staff , without whose help

the project might have been completed.

I express heartfelt thankful to Prof. Dr. S.S. Rathod , Principal & Prof

.A.L.Nashine , H.O.D., Civil Engineering & Prof. P.E.Mishra Coordinate, PG

Deptt. Of Environmentel Engg.,MIET, Gondia, for providing necessary facilities in

the completion of this work and for his constant encouragement.

Sandeep P. Ajmire


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C O N T E N T SINTRODUCTION

1.1. General

1.2 . Polution Problem

1.3 . Standards of Disposal

1.4. Treatment methodology

1.5. Objective and scope of study

LITERATURE REVIEW

2.1. General

2.2. Characteristics of domestic waste water

2.3. Tretment Processes

2.4. Process selection criteria for treatment of various domestic waste water

2.5. Application of Phytoremedation to domestic waste water

PHYTOREMEDATION

3.1. History & back round

3.2. Definition & types of Phytoremedation

3.3.Introduction of Phytoremedation by Lemna

3.3.1

3.3.2

3.3.3

3.3.4

3.3.5

3.3.6

3.3.7

3.3.8

3.3.9

3.4.

3.5


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PLANTS AND METHOD

4.1

4.2

4.3

4.5

4.6

4.7

4.8

4.9

4.9.1

4.9.2

4.9.3

4.9.4

4.9.5

4.9.6

4.9.7

4.10

OBSERVATIONS ,RESULTS,AND DISCUSSION

5.1

5.2

5.3

5.4

5.5

5.6

5.7

Reference:

PHOTOGRAPHS


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I N T R O D U C T I O N

1.1. General

The population of glob is increasing, the problem of municipal &

industrial waste tedious day by day. The legacy of rapid urbanization ,

industrialization ,fertilizer & pesticide use has resulted in major pollution

problems in both terrestrial and aquatic environments. In developing

countries is major problem to treat the polluted water from above sources.

Chemical & mechanical menace are used for this purpose is expensive. In

response , conventital, remediation systems based on high physical andchemical engineering approaches have been developed and applied to avert

or restore polluted sites. Much as these conventional remediation systems

are efficient, they are sparsely adopted because of some economical and

technical limitations. Generally, the cost of establishment and running deter

their use and meeting the demand particularly in countries with week

economy. Logical this high cost technology can neither be applied justifiably

where

1. The discharge is abruptly high for short time but the entire

average load is relatively small.

2. The discharge is very low but long term (entire load is medium).

3. The discharge is continuously decreasing over a long duration.

Thus conventional remediation approaches are best for circumstances

of high pollutants discharge like in industrial mining and domestic waste

water. Recently , it is evident that durability restoration and long term

contamination control in conventional remediation is questionable because


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in the long run the pollution problem is only is suspended or transferring

from one site to another.

The efficiency of duckweed (Lemna gibba L.) as an alternative cost

effective natural biological tool in wastewater treatment in general and

eliminating concentrations of both nutrients and soluble salts was examined

in an outdoor aquatic systems. Duckweed plants were inoculated into

primary treated sewage water systems (from the collector tank) for aquatic

treatment over eight days retention time period under local outdoor natural

conditions. Samples were taken below duckweed cover after every two days

to assess the plants efficiency in purifying sewage water from different

pollutants and to examine its effect on both phytoplankton and total and

fecal coliform bacteria.

The Lemnaceae family consists of four genera (Lemna, Spirodela,

Wolffia & Wolffiella) and 37 species have been identified so far. Compared

to most other plants, duckweed has low fiber content (about 5%), since it

does not require structural tissue to support leaves and stems. Of these,

applications ofLemna gibba L (duckweed) in wastewater treatment was

found to be very effective in the removal of nutrients, soluble salts, organic

matter, heavy metals and in eliminating suspended solids, algal abundance

and total and fecal coliform densities. Duckweed is a floating aquatic

macrophyte belonging to the botanical family Lemnaceae, which can be

found world-wide on the surface of nutrient rich fresh and brackish waters.

Outdoor experiments to evaluate the performance of the duckweed as a

purifier of domestic wastewater in shallow mini-ponds (20 & 30 cm deep)

showed that quality of resultant secondary effluents met irrigation reuse

criteria. Wastewater ammonia was converted into a protein rich biomass,

which could be used for animal feed or as soil fertilizer. The economic


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benefit of the biomass by-product reduced wastewater expenditures to

approx. US$ 0.05 per treated m3 of wastewater, which was in the range of

conventional treatment in oxidation ponds.

The present study was concerned with decreasing pollution of

municipal waste waster up to degree Standards of Disposal as per

National pollution control board.

1.2 . Pollution Problem

The quality of municipal wastewater of stagnant/ slow velocity may

create problem of high epidemics of malaria & other water born

diseases. The efficiency was tested by measuring some of

physicochemical characteristics of the control and plant treatments

after each seven days. The experiment lasted for a month, and recorded

the rates of reduction. The highest rates of reduction were for heavy

metals, accounting 99.8%, 99.6%, 98.7% and 72% for Copper,

Cadmium, Lead and Zinc, respectively, followed by Turbidity and

Nitrate recorded 64% and 57.1% respectively. The percentage

reduction of BOD5 was 49.6 %, while for COD 32.7%, either the rest of

the physicochemical characteristics as follows: 48.9% for Soluble Solids

and 43% of Oils and Grease, 41% of Total Alkalinity, 40% for Phenols,

39.1% for Sulfide, 38% of Suspended Solids and 30% of Phosphate.

Lower rates of reduction were recorded of each Temperature (17.2%),

pH and Sulfate (13.4%). The results showed that this aquatic plant canbe successfully used for wastewater pollutants removal


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Refinery wastewater is producing in a huge quantity in most the cities

of the country that contain a diverse range of pollutants including

Heavy Metals ,Oil and Grease ,Phenols, Sulphide, Sulphate ,Nitrate

,Phospate, Dissolved Solids, Suspended Solids, COD, BOD, which Its

disposal and treatment has become a challenge for the municipalities;

Many of the municipalities in growing cities neither have proper

disposal system nor have any treatment facility due to higher cost and in

such a situation refinery wastewater is discharge in to aquatic bodies

like river, ponds

and lakes, where it is posing a serious threat to the water quality and

become a big environmental problem throughout the development of

the petrochemical industry [1] [2].Physical, chemical, and biological technologies have been developed

to treat oily waste water and restore environmental quality; However

their costs are high and most of them are difficult to use under fieldconditions, hence in such a condition there is an urgent need to study

natural, simple, and cost-effective techniques for control pollution from

industrial effluents and treating such wastewater, such as

phytoremedation [1] [3].Viewing this fact Phytoremediation was assumed to be very useful, as

it is an innovative, eco-friendly and efficient technology in which naturalproperties of plant is used in engineered system to remediate hazardous

wastes through physical, chemical, and biological processes from

wastewater and sewage [4] [2].Phytoremedation is the utilization of plants accumulation capabilities

to remove contamination from water, soil and air, the capacity ofaquatic plants to remove pollutants from water is well documented [5].

The recent application of phytoremediation technology by duckweed inwastewater treatment and management is quite interesting andrevealing. Phytoremediation systems by duckweed are one of the

options that have been widely applied for combined handling of

wastewater with the nutrients used for poultry and aqua-culturalprojects [6] [7].


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Lemna minor L. known as common duckweed is a small, freefloating aquatic plant fast growing, adapt easily to various aquatic

conditions and play an important role in extraction and accumulation of

pollutants from waters [8]. In particular, species of Lemna are reportedto accumulate toxic metals and therefore are being used as experimental

model systems to investigate heavy metal induced responses,

Bioavaibility and bioaccumulation of various heavy metals in aquaticand wetland ecosystems is gaining tremendous significance globally [9].

Aquatic macrophytes take up metals from the water producing an

internal concentration several fold greater than their surroundings.Many of the aquatic macrophytes found to be the potential scavengers

of heavy metals from aquatic environment and are being used inwastewater renovation systems [10] [11]. Aquatic plants have shown

their efficiency in absorbing nutrients from various sources of polluted

water, [12] [13].This study aimed to assessing the efficiency of duckweed (Lemna minor)

in phytoremediate the pollutants of Basrah oil refinery wastewater.

Pour dterminer la tolrance et la capacit de phytoaccumulation ducuivre (Cu) et du nickel (Ni) par

une espce de lentilles deau, Lemna gibba L., les plantes sont exposes

diffrentes concentrationsde Cu et Ni (0,1 2,0 mg/L) dans une solution de Coc et Lesaint dilu

1/4. Le pH est maintenu

constant 6,0 ( 0,1) et le flux de lumire est de 12 h/jour. Le cuivre etle nickel sont tolrs par L.

gibba des concentrations 0,3 mg/L et 0,5 mg/L, respectivement.

Cependant, la croissance desplantes diminue de 50% (I50) quand le milieu de culture contient 0,45

mg/L de Cu ou 0,75 mg/L de Ni.La plus faible concentration causant une inhibition complte (LCI) est

de 0,5 et 1,0 mg/L

respectivement en prsence de Cu et Ni. Les rsultats de lanalyse dumtal dans les tissus des

plantes rvlent une grande accumulation de Cu et une faibleaccumulation de Ni dans les tissusvgtaux (pour la concentration ne causant aucune inhibition dans la

croissance). Une diminution de

la concentration de mtal dans leau est galement observe. On peutconclure que L. gibba peut tre

un bon candidat pour lpuration des eaux contamines par le cuivre.


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ABSTRACTTo assess the tolerance and phytoaccumulation ability of the duckweed

Lemna gibba L. to copper

(Cu) and nickel (Ni), the aquatic plants were exposed to differentconcentrations of Cu and Ni (0.1

2.0 mg/L) in quarter Coc and Lesaint solution at pH = 6.0 ( 0.1) and

under a daily regime of 12 hlight. Copper and nickel were tolerated by L. gibba at concentrations

0.3 mg/L and 0.5 mg/L,

respectively. However, plant growth decreased by 50% (I50) when themedium contained 0.45 mg/L of

Cu or 0.75 mg/L of Ni. The observed LCI (lowest concentration causingcomplete inhibition) were 0.5

and 1.0 mg/L respectively in the presence of Cu and Ni. Results from

metal analysis in plant tissuesrevealed a high accumulation of copper and a low accumulation of

nickel within the plant (for

concentrations causing no growth inhibition) and a correspondingdecrease of metals in the water.

The duckweed L. gibba L. could be a good candidate for the removal of

low concentrations of copperfrom polluted water.

Certain plants are able to accumulate in their tissues several metals

without showing any signs oftoxicity. This natural accumulation is related with the resistance which

represents response of plants

to metal stress conditions. According to Papazoglou et al. (2005), metalresistance can be based on

either avoidance or tolerance mechanisms. Avoidance reflects the cellprotection against the metal

whereas tolerance is the cell capability to protect themselves against

injury by the metal (Sabreen &sugiyama, 2008). Kanoun-Boul et al. (2008) reported in the tolerance of

duckweed to copper that therelease of organic anions might be involved in the protection of plantsby chelating the metal ions in

the rhizosphere to form non toxic complexes. However, according to

Sabreen and Sugiyama (2008),tolerant populations can be characterized by a lower metal

accumulation than the sensitive one.


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Duckweed commonly refers to a group of floating, flowering plants ofthe family Lemnaceae. The

different species (Lemna, Spirodela, Wolffia and Wolfiella) are

worldwide distributed in freshwater andwetlands, ponds and some effluents are the most common sites to find

duckweed. The plants are fast

growing and adapt easily to various aquatic conditions. They are able togrow across a wide range of

pH, from pH 3.5 to10.5 but survive best between pH 4.5 to 8.3

(Environnement Canada, 1999;Cayuela et al., 2007). The plants are found in temperate climates and

serve as an important foodsource for various water birds and fish (Drost et al., 2007).

Some studies indicate that duckweed plants are sensitive to toxicity.

Other studies however, reportthat duckweed plants are tolerant to environmental toxicity (Wang,

1990). To assess the tolerance of

the species L. gibba to heavy metals, plants were exposed toconcentrations of copper and nickel

higher than those used in medium cultures. Toxic effect of pollutant on

duckweed is generallyevaluated by phytotoxicity tests based on growth inhibition (Geoffroy et

al., 2004). Copper and nickel

were chosen as the metals for this study for a number of reasons. Theirpresence above trace levels

in the environment is an indicator of industrial pollution. On the other

hand, they are essentialmicronutrients for plants; copper is a structural and catalytic

component of many proteins andenzymes involved in metabolic pathways (Teisseire & Vernet, 2000) and

nickel has an important role

in the urease and hydrogenase metabolism (Harish et al., 2008).However, when the concentration

reaches a threshold value, these essential metals become first inhibitoryand afterwards toxic. Copperis responsible for many alterations of the plant cell (respiration,

photosynthesis, pigment synthesis

and enzyme activity) (Teisseire & Vernet, 2000; Kanoun-Boul et al.,2009). Nickel inhibits


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germination, chlorophyll production and proteins (Zhou et al., 2009) inplants; several animal

experimental studies have shown an increased cancer incidence

associated with chronic exposure tonickel.

Each plant species has different resistance and tolerance levels to

different contaminants (Kamal etal., 2004). Therefore, several studies have been performed to elucidate

heavy metal toxicity to plants.

In earlier study, we demonstrated that the duckweed L. gibba L. nativeto the north-east of Algeria

tolerated Zn up to 18 mg/L and was effective in removing 60% of themetal pollutant from the nutrient

medium (Khellaf & Zerdaoui, 2009). The present study investigates

copper and nickel toxicity toduckweed to determine tolerance of this aquatic species to Cu and Ni.

The study also investigates the

potential accumulation of these two metals by the duckweed L. gibba L.The goal was to assess the

possibility to use L. gibba L. for phytoremediation of Cu and Ni

contamination in water.

1.3 . Standards of Disposal

1.4. Treatment methodology


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1.5. Objective and scope of study


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MATERIALS AND METHODS

Preparation steps. Primary treated sewage water were transferred to the

laboratory from the tertiary sewage water treatment plant after the

preliminary sieving step to get rid of large suspended solids. The transferred

water was immediately collected into two opaque tanks (as replicates) to

prevent light entering except at the top (Parr et al., 2002), each tank with

dimensions of 50 cm long, 35 cm wide and 25 cm deep and was filled with

25 L primary treated sewage water. Duckweed (Lemna gibba L.) plants ere

collected from Ganabiet-Tersa drain (Fig. 1). The stock were cleaned by

tap water then washed by distilled water inocula of Lemna plants were

transferred to the water systems for aquatic treatment. The experiment was

kept under outdoor local environmental conditions for eight days retention

time.

Water sampling. Subsurface (under duckweed mt) water samples for

physico-chemical, biological and bacteriological parameters were collected

in polyethylene bottles from all sides of each tank and then mixed. This

procedure carried out every 2 days. Samples volume taken every two days

for each of phytoplankton count and chlorophyll a determination was 100

mL.

Parameters measured. Physico-chemical analyses (Table

I) were carried out according to standard methods for e examination of water

and wastewater (APHA, 1992). Field parameters (pH, conductivity &

dissolved oxygen) were measured in situ using the multi-probe system

(model Hydralab-Surveyor) and rechecked in laboratory using bench-top

equipment to ensure data accuracy for biological parameters including total


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coliform count and fecal coliform. count, phytoplankton identification and

counting andchlorophyll a determination.

Determination of duckweed growth rate. This was determined for fresh

and dry weights. Samples of 20 cm2 area ofLemnaplants were harvested

periodically at the designated time periods (every 2 days) and filtered using

filter papers then fresh weights were determined. These samples were then

dried at 60oC for 48 h to a constant weight and then dry weights were

calculated.

Protien content. Duckweed organic nitrogen content was estimated at the

beginning of the experiment and after 8 days retention time by using Micro-

Kjeldahl method, then the obtained values were multiplied by 6.25 to obtain

protein content values.


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