mengel1 - plant physiologyplaint physiol. ('1967") 42, 6-14 ionic balance in different...

Plaint Physiol. ('1967") 42, 6-14

Ionic Balance in Different Tissues of the Tomato Plantin Relation to Nitrate, Urea, or Ammonium Nutrition

E. A. Kirkby and K. Mengel1Department of Agricultural Chemistry, the University, Leeds, 2, England

Received June 27, 1966.

Sumnwarv. An investigation was carried ouit to study the cation-anion balance indifferent tissues of tomato plants suipplied with nitrate, urea, or ammonium niitrogenin water culture.

Irrespective of the form of nutrition, a very close balance was found in the tissuesinvestigated (leaves, petioles, stems, and roots) between total cations (Ca, Mg, K andNa), and total anions (N03-, H2P04-, SO4--, Cl-) total non-volatile organic acids,oxalate, and uronic acids. In comparison with the tissues of the nitrate fed plants,the corresponding ammonium tissues contained lower concentrations of inorganiccations, and organic acids and a correspondingly higher proportion of inorganic anions.Tissues from the urea plants were intermediate between the other 2 treatments.These results were independent of concentration or dilution effects, caused bygrowth. In all tissuies approximately equivalent amounts of diffusible cations (Ca",\Ig+, K' and Nat), and diffusible anions (No3-, SO4--, H,PO4-, Cl-) and non-volatileorganic acids were found. An almost 1:1 ratio occurred between the levels of boundcalcium and magnesium, and oxalate and uronic acids. This points to the fact thatin the tomato plant the indiffusible anions are mainly oxalate and pectate. Approxi-mately equivalent values were found for the alkalinity of the ash, and organic anions(total organic acids including oxalate, and uronic acids).

The influence of nitrate, urea, and ammonitum nitrogen nutrition on the cation-anionbalance and the organic acid content of the plant has been considered and the effectsof these different nitrogen forms on both the pH of the plant and the nutrient mediumand its conseqtuences discussed.

Ionic equilibrium in plant tissues is maintainedby diffusible and indiffusible ions, in which bothorganic and inorganic cations and( anions play apart. It is known that this equilibrium is verymuch dependent on the form of nitrogen nutritionto the plant (6, 9, 14). Previous studies have cen-tered mainly on the comparison of nitrate andammonium sources on the diffusible ionic balancein leaves, (6, 9,14) or oIn the influence of thesenitrogen forms on the uptake of other ions (19).These experiments have shown that in comparisonwith ammonium fed plants, nitrate plants containhigher concentrations of cations and organic acidanions, while the content of inorganic anions islower. Such observations have been explained bythe axiomatic necessity for all plant tissues tomaintain ionic equilibrium.

In order to investigate how different plant tis-sues maintain this equilibrium and the contributionsof variouis components responsible for electroneu-

K. Mengel. Institut fuir Pflanzeilerinahrung der JustusLiebig IUniversitdit, Giessen, Germanv.

trality, both diffusible and indiffusible iOlns were

determined in the leaves, petioles, stems, and rootsof tomato plants supplied with nitrate, urea, orammonium nitrogen. The inclusioni of urea as acomparative treatment also made it possible tosubstitute one main ionic component of the nutrientsolution by an undissociated molecule. It was thuspossible to compare systems in which the nutrientbeing taken tip by the plant in greatest quantities,nitrogen, was in the form of an anioin, molecule orcation.

It is well known that the form of nitrogennutrition influences the pH of the nutrieint solution(20). As this pH shift should be related to nitro-gen metabolism, the present paper also considersthe relationships between electroneutrality of thetissue, pH shifts in the medium andc aspects ofnitrogein assimilation and metabolism.

Materials and Methods

Tomato plants (\var Ailsa Craig) N-ere groxwnfrom seed in the glasshouise in a soil comllpost mix-

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KIRKBY AND MENGEL-IONIC BALANCE IN RELATION TO NITROGEN NUTRITION

ture. At the 4 leaf stage of growth the plantswere removed from the soil, and by thoroughlyrinsing in running tap water followed by distilledwater all adhering soil particles were removedfrom the roots. The plants were then transferredto aerated nutrient solutions held in 7 liter blackpolythene containers, each container supporting 2plants held by rubber foam in a 2 cm thick poly-styrene sheet. The 3 treatments, nitrate, urea, andammonium were replicated 6 times.

The nutrient solutions are based on those usedby Chouteau (6). The nitrate solution was madeup as follows: KH9PO, (2 meq/l), MgSO4 (1.5meq/l), Ca(NO3 )2 (5 meq/l). In both the ammo-nium and urea solutions, Ca ( NO3) 2 was replacedby an equivalent of CaSO4 (5 meq/l), and thenitrogen level was kept constant by the addition ofthe equivalents of (NH4)2SO4 or CO(NH2)2.Thus sulfate was the only variable. The micro-nutrients were supplied in all the 3 treatments as:Fe-EDTA (2.80 jug Fe/ml), MnCl24H,O (0.550JUg Mn/ml), H3B,03 (0.330 Jag B/ml), ZnCl, (0.065jug Zn/ml), CuCl22H2O (0.064 ,ugC,u/ml), Na9MoO4(0.046 ug Mo/ml).

At the beginning of the experiment the pH ofall the solutions was increased to 5.5 with diluteCa(OH)2 solution. During the growth period thepH in all treatments was adjusted back to 5.5 every2 or 3 days with Ca (O1H) 2 solution or 0.02 NH2SO4. After 14 days the nutrient solutions werecompletely renewed. During growth occasionalqualitative checks were also made on the nitratecontent in the solutions of the ammonium and ureaseries, and for the ammonium nitrogen content inthe urea treatment. As these proved to be negativeit is perhaps legitimate to assume that nitrogen wasabsorbed in the forms supplied.

After 20 days growth in the nutrient media theplants were harvested and divided into leaves,petioles, stems, and roots and each replicate weighed.Similar tissues from each treatment were bulkedtogether. Weighed samples, obtained as soon aspossible after harvesting were stored in polythenebags at -15°. The remaining weighed fresh ma-terial was dried at 850 to constant weight andground to a fine powder using a micro hammermill.

Chemical estimations were made on both thefresh and dried plant material. The dried materialwas used for the estimation of total nitrogen,nitrate, inorganic sulfur, inorganic phosphorus,chloride, calcium, magnesium, potassium, sodium,oxalate, alkalinity of ash, and uronic acids. Totalnitrogen, nitrate, inorganic sulfur, inorganic phos-phorus and chloride were determined as describedin another publication (15). Calcium, magnesium,potassium, and sodium were determined on 0.1 Nhydrochloric acid extracts of the ash. Potassiumand sodiuim were estimated by flame photometry,calcium as oxalate and magnesium by atomic ab-sorption spectrometry. The same methods of esti-

mation were also used for calcium and magnesiumin boiling water extracts. Total oxalate was de-termined according to Baker (3). Free oxalate,oxalate soluble in boiling water, was estimated butfound only in trace amounts. Uronic acids alsowere determined using the manometric method ofTracey (21) in which CO2 evolved on decarboxvla-tion by 12 % (W/W) HCI is measured in a VanSlyke apparatus. The results were then calculatedin meq per 100 g dry plant material by dividing theweight of 0O2 evolved by 44 assuming all the CO2was derived from pectic substances.

The alkalinity of the ash was determined byashing 200 mg of dry plant material at 4800 for 24hours. By means of a back titration using 0.1 NHCI followed by 0.1 N KOH, excess alkalinitycould be determined. On the assumption that theinorganic anions other than nitrate are not affectedby ashing, this valtue should be equivalent to thetotal organic anions in the dry plant material.

Organic acids were determined on the freshplant material. Using samples of 5 g of tissue,the organic acids were extracted by repeated treat-ments with boiling water after maceration of theplant tissue. The acids were estimated by partitionchromatography using a silica gel column, aftertheir isolation, as described in detail by DeKockand Morrison (10).

The results of the analyses are expressed interms of meq per 100 g dry weight for comparativepurposes. Phosphorus is considered monovalentand sulfur divalent following the work of Dijk-shoorn (12).

Results

pH Changes in the Nutrient Soluitions DuringGrowth. The well-known tendency for an increasein pH in the external medium with nitrate nutritionand a corresponding decrease with ammoniumnutrition was found (fig 1). Urea had an inter-

7.0Sol ut ionschanged

pH

40

0 5 10 15 20GROWTH PERIOD (days)

FIG. 1. pH Changes in the nutrient media duringgrowth (pH adjusted back to 5.5 after every determina-tion). (A) NO3, (-) urea, ( 0 ) NH4.

7

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I'LANT PHYSIOLOGY

mediate effect, the nlutrieint me(liutm becomingaci(lic duiring the growth perio(l.

Symitptomits at Harvest and Dry Hlaltter Yields.On harvesting the plants, marked(ldifferences be-tweeni the 3 treatments were observed. In com-parison with the nitrate plants, the ammonitim fedplants were small with very (lark green leaves and]a very poorly developed sttibby root system. Incontrast the nitrate plants were very well (levelope(Ian(l had an extremely large root sy-stem. The tipperparts of the urea plants were intermediate in sizealthouigh the growth was rather spinidly, the leavesshowing necrosis at the tips and a pale green color-verging on the appearance of nitrogen deficiency.Suirprisingly however, the roots were very well(levelope(l.

The lower yields obtained in all tisstues from theammoniulm plants in comparison with the corre-sponding tisstues of nitrate fe(d plants show out veryclearly (table I). This has been observed l-, other

Table I. Influence in the Form of Nitrogen Nutritionon the Dry Matter Yields of Leaves, Petioles, Stemiis

and Roots of Tomtato (g per 10 Plants)

ILeaves Petioles Stems RootsN Source

NO.,

NH.1

30.215.39.6

9.9 12.63.4 6.52.8 53

10.29.74.8

N-orkers uising tomato (8, 9). Urea is intermedliateini its effect except for the root weights which werealmost equial to those of the nitrate treatment. Asthe root system of the uirea plants developed undera low pH regime in the nuttrieilt mediuim, it istherefore very unlikely that the low pH is solelyresponsible for the poor root development in theammoniutm treatment.

Table II. Influienec of tli Form of Nitrogen Nutritionon thle Nitrogen Content of thle Leaz.es, Petioles,

Stems and Rooivisf TI Wato) (meq per 100 g drv wet)(A mllOnlovalcnt)

N Souirce

NO,UreaNH4

Leaves Petioles Stems Roots

280157313

13479

153

15273

192

294171340

Nitrogent Contentts. Muich lower contenits ofnitrogen occtirred in the tissuies of the turea plantsthan in tissues from corresponding tissues fromeither of the other 2 treatments (table II). Thesomewhat higher concentrations of nitrogen in theammonia plants as compared with the nitrate plantsmay be caused to some extent by concentrationleffects because of the lower yields in this treatment.

Cation Com1zposition. The cation contents invariouts tissues of the tomato plant in relationi tothe form of nitrogein nuitrition are shown in tableIII. The highest concentration of individualcations, other than soditum, occturred in the tissuesof nitrate plants and was lowest in the ammoniutmfed plants. The same is also true for the totalcation concentration. The well-known increase ofpotassiuim in stem and petiole tissues, and the re-duction in calcium concentration from the leaf tothe root was also apparent in all 3 treatments.The resutlts are generally in agreement with thefindings of a ntumber of the other writers, fromstuidies in nitrate and ammonium nultrition (1, 6,13, 14, 19, 22). As dry matter yields were alsolowest in the ammonitim plants the lower contentsof cations in this treatment can not have resulte(dfrom concentratioin or diluition effects and muist,therefore, be dependent oIn differences in ion uiptake.

Potassium and sodium occuir in plant tissuiesalmost totally in ionic form. \Vith calcitum anid(magnesiuim this is not the case. In order to obtain

Table III. Influence OJ tlh F )rm of N'itroqen Nutrition on Ihe Ino(ryanic Ca/ion Content inStems, (ind Roits of T(cmato( ( meq ter 100 g dry zet)

the Lcazc-es, Pe tiolet. .

C A T I 0 N SCa Mg K

(me(q per 100 g dry -\t)

16113462

12611061

868250

444038

3027

25

383017

353118

402711

583429

17616290

16212754

9310543

Tisstue

Leaves

Petioles

Stellm.

N Soutrce

NO.,UreaNH

N\O.,'UreaNH1

NO.,

Urea

NH14N O.,

Urea

NH4

Root

Na

191515

183026

182618

161513

Total

268210131

358332194

301266140

193187105

IR



KIRKBY AND MENGEL-IONIC BALANCE IN RELATION TO NITROGEN NUTRITION9

a measuire of diffuisible cations, so that the difftu-sible cation-anion balance couild be considered,boiling water extracts were made on the dry plantmaterial for calcium and magnesium estimation.The resuilts are shown in table IV. Althoutgh about50 % of the total calcium was present in ionic formin the leaves, only about 10 % was water soltiblein the petioles and 15 % in the stems and roots.A similar pattern is also true for magnesium exceptthat a higher percentage than calcitum was foundin ionic form in all tissues.

Anion Composition. The inorganic anion con-tents in table V show that in the leaves and petiolesthe nitrate plants have the lowest inorganic anionconcentration, and urea the highest. It would beexpected that the inorganic content of nitrate fedplants shoutld be low as nitrate the major anionabsorbed by the plant is converted largely to organicform. This is especially the case in the upperplant parts at the significant site of nitrate re-

Table IV. Influence of tile Form of Nitrogen Nutritionon the Content of Water Soluble Ca and M1g in the

Leaves, Petioles, Stems, and Roots of TomtatoResults expressed as a percentage of total Ca and

total Mg.

Tissue N Source % Ca % Mg

Leaves NO. 50 85

Petioles

Stems

Roots

UreaNH4N03UreaNH4NO3UreaNH4

NO3UreaN,H

4836

1199

141716

171521

6553

373935

272327

443689

duiction in tomato (5). When ammonitum ions aretaken up higher amounts of inorganic anions arealso necessary to maintain ionic balance. As thediffusible cation concentration in the tissues of theammonium plants is rather low, however, the some-what lower concentrations of inorganic anions, thanin the urea plants and the corresponding stems androot tisstues of the nitrate fed plants, may be ex-plained. This effect can be seen from the restultsof the inorganic anions expressed as a percentageof the diffusible cations where the urea and nitratetissues have a lower percentage of their diffulsiblecations bound with organic anions than corre-sponding ammonium tissues.

The individual inorganic anions in the planttissues investigated follow the same pattern in all3 treatments, phosphate being highest in the roots,chloride in the stems and petioles and sulfate inthe leaves.

In most plant tissuies the prevailing pH is sutchthat organic acids are present as the salts of in-organic cations. It is only in a few rather atypicalplants that free acids are present to any markedextent. The non-volatile organic acids shown intable VI may thus be considered to contribute tothe ionic equilibrium largely as organic anions.The influence of the form of nitrogen nutrition isshown very clearly, nitrate tissues having very muchhigher concentrations of organic acids than corre-sponding ammonitum tissues. This confirms thefindings of Clark (8) and Coic (9) for tomatoleaves and other workers using a nuimber of plantspecies (6,13, 14). From the literatuire there ap-pears to be no earlier work on the effect of uireanitrogen on the concentration or composition of theorganic acid fraction in the plant. Our resuiltsshow the intermediate position of turea betweennitrate and ammonium nutrition in this respect.There is also a marked effect on the concentrationof the acids between tissuies of the same plant, theorganic acids falling in concentration from the leafto the root.

Table V. Infliuence of tile Form of Nitrogen XN'utrition on Content of Inorga,nic Ani ns in the Leaves, Petioles,Stems and Roots of T( mato (meq per 100 g dry zi't)

INORGANIC ANIONSSO4-- H2PO4- C1-

(meqI per 100 g dry wt)N03 Total % of

Duf fusible cations

223135

122029

131515

152120

122314

308949

8 13 2612 20 59

14 15 27

103515

27

3521

133217

Tissv:e N Source

Lea, es

Petioles

Stems

Roots

NO3UreaNH4NO..

NH4NO3UreaNH4

NO.,U-eaNH4

4..

16

..

20

35..

..

516964

7313098

679156

858453

285381

336177

335266

636272

9



PLANT PHYSIOLOGY

Diffusible Cation-Anion Balantce. Figure 2illustrates graphically the relationship between thediffusible cations and diffusible anions. The re-sults show a very highly significant relationship(r = 0.992, b = 0.916+0.056, P <0.001), whichinfers that the most important diffusible cationsand anions have been taken into account. Varia-tions from the ideal 1:1 ratio may be accounted forby the cumulative errors of chemical analysis oromissions from the balance suich as proteins andorganic bases.

Non-Diffusible Cations and Anions. Table VIIshows the comparison of the totals of bound calciumand magnesium against total oxalate and uronicacids. There is a remarkably good agreement be-tween the cation and anion totals (r = 0.944,b = 1.119±0.124, P<0.001). It must be consid-ered, however, that althouigh oxalate was found tobe present almost totally in salt form, this may not

be the case for the uironic aci(ds. Boiliig wateralso is not ideal for the extraction of diffuisiblecalcium and magnesium. Moreover, some non-dif-fusible magnesium is present in chlorophyll, althoughthis is only a very small fraction of the totalmagnesium concentration (16). The close agree-ment between the results in table VII infers, how-ever, that non-diffusible calciuim and magnesiuimions are largely associated with pectate and oxalate.

The form of nitrogen nutrition shows exactlythe same effect with the oxalate content as withthe diffusible non-volatile organic acids discuissedpreviously, highest concentrations being fouin(d inthe tissues of nitrate fed plants. In agreementwith other workers rather high levels of oxalatewere fouind (8, 9). It is of interest to note thatthe tironic acid content was not so greatly influ-enced by the form of nitrogen nutritioni as thenon-volatile organic acids. This is, as would be

Table VI. Influence of t11 Form of Nitrogen Nutrition on the Content of Non-Volatile Organic Acid Anions in theLcazcs, Pctiolcs, Stems, and Roots of Tomato (mneq per kg fJ ret)

Tissue N source

Leaxves NO3UreaNH4

Petioles NO3UreaNH4

Stems

Roots

NO3UreaNH4

NO3UreaNH4

Fumaric Succinic(meq per kg fr wt)

1.20.40.5

1.00.3o.5

1.8030.9

0.90.21.5

1.20.908

0.30.30.8

0.60.80.6

2.1160.8

Table VII. Influence of tlhc Formn of Nitrogen Nutrition on the Contents of Bound Calciwnsl and Ilagncsium, andOxalate and Uronic Acids in the Leaves, Petioles, Stems, and Roots of Tomtato (meq per 100 g dry ret)

Tissuie

Leaves

N Source

NO3UreaNH4

Petioles NO3UreaNH4

Stems

Roots

NO3UreaNH4

NO3

UreaNH4

Cations TotalCa Mg

(meq per 100 g dry w-t)816940

11210156

6842

373430

51012

241911

252313

22171

8679

13612067

1009155

595131

.AiioilsOxalate Uronic Acids

4125

8

614916

584318

2216

1

444046

696863

54

50

_5

42

Malonic

1.80.81.1

0.4

0.60.42.4

3.06.3

Malic

130.225.55.9

95.666 514.7

70.143.67.4

13.79.11.2

Citric

47.455.610.6

11.54.81.1

2.52.71.1

9.33.00.3

Total

181.883.218.9

108.471.917.$

75.647.812.9

29.020.23.8

Total

85rfi;

13011,79

112Q3,3~13~,s

10



KIRKBY AND 'MENGEL-IONIC BALANCE IN RELATION TO NITROGEN NUTRITION1

100 200DIFFUSIBLE ANIONS(meq/100g)

FIG. 2. The relationship between the diffusible ca-tions (Ca++, Mg++, K+, Na+) and diffusible anions(NO3-, S045 , H2P04-, Cl- and non-volatile organicacid n-iions) in meq per 100 dry wt. (A NO3, (N) urea,(0) NH4. y = 23.45 + 0.916 x.

expected, that the formation of primarv structuresand cell walls of the plant are less dependent ofthe form of nitrogen nutrition.

Total Organic Anions. The results discussedearlier indicate the very close relationship betweenthe total cations and anions in all plant tissues. Inorder to confirm that all the anions and cationscontributing to the balance had been taken intoaccount, the total amounts of organic anions asobtained by the alkalinity of the ash method weregraphed against the sum of the organic anions

Table VIII. Inzfluence of the

obtained by the other chemical estimations describedearlier. The results are shown in figure 3. Thevery high significant relationship (r= 0.975, b =

0.997±+ 0.071, P<0.001) between these 2 independentestimations of organic anions gives a very goodindication that no anions or cations have beenomitted from the balance in the results discussedpreviously. The results also demonstrate the use-fulness of the alkalinity of the ash method in esti-mating organically bound anions.

Total Cation-Anion Balance. The contributionof the individual fractions to the total cation-anionbalance in the various plant tissues in relation tothe form of nitrogen are shown in table VIII. Theoverall effect shows out very clearly in all tissues.A very close balance occurs in all treatments.Nitrate tisstues are highest in cations and anions,ammonium tissues lowest and those of urea areintermediate.

Discussion

The experimental data give very good stupportto the concept of a cation-anion balance in differentplant tissues, maintained by the diffusible and in-difftusible organic and inorganic, cations and anionswhich were determined. In each tisstue, independentof the form of nitrogen nutrition, the total cationsare fairly well balanced by total anions. There isalso a remarkably good agreement between thebound calcium and magnesium, and oxalic anduronic acids. It thus seems probable that these 2organic constituents are mainly balanced by calciumand magnesium ions.

The different forms of nitrogen ntutrition re-sulted in very different yields and growth habits ofthe plants. It is clear that there is a very close

Formii of Nitrogen Nutrition on the Cation-anion Balance in the Leaves, Pctioles, Stemns,and Roots of Tomato (meq fcr 100 g drv wt'

CATIONS Non ANIONSvolatile

N Indiffus- Diffus- OrganicTissue Source ible ible Total acids Inorganiic Uronic Oxalate Total

(meq per 100 g dry wt)

Leaves NO3 86 182 268 117 51 44 41 253UreaNH4

Petioles NO3UreaNH4

Stems

Roots

NO3UreaNH4NO3UreaNH4

7952

13612067

999155

595131

13179

222212127

20217585

13413674

210131

358332194

301266140

193187105

4111

1479418

1145812

4530

6964

7313098

679156

858453

4046

696863

5450535565242

258

614916

58431822161

175129

350341195293242141

208182101

11



PLANT PHYSIOLOGY

° 200

0-

-E

z

<I:lOO1

0 100 200

ORGANIC ANIONS (meq /100g)

FIG. 3. The relationshlii) betsveen the alkalinity of theitrate free ash and the conitent of organic anions (total

organic acild aniions and uronic acids) in me( per 100drx-wt. (A) NO., (U) uirea, (* NHj. y17.17 + 0.997 x

initeraction between the uiptake of ionls by the plantanld drv matter yield. ariationis in vield maytherefore produce concentratioin or dilution effectsonl catioIn anid anlioIn constituienits. These effects,however, shouil(d be similar for both catioils aindanioins. As the restilts showed, this was not thecase, so that the influiencc of yield differeincescaninot accouint for constitueint changes in thecation-anion equilibrium brouight about by the 3forms of ilitrogen nutritioin.

\Vhein conisiderinig the questioin of cation-ainionbalance in relationi to different nitrogen soturces itmuist be kept in mind that in comparison with othernittrieints, the plant needs very high quaintities ofnitrogeni in or(ler to realize its inherent reqtiire-menits. Thus with nitrate nultritioin the plant hasto take tip high amouints of anions while withamiimoniutnm Inlutritioin high quLaintities of cationis mustbe absorbed. Already at this initial step with theuiptake of niitrate or ammoniulm ions into the cell,a halance has to occur otherwise the electropoten-tial drop between the cell and medium wouldl bedeleteriouis to the cell. The uiptake of nitrate has,therefore, to he accompainie(d by cations or by an

exchainge of anions, juist as the uptake of ammo-

niulmln iOlns muist be accompainied by anionls or the

exchange of other cations from the root. Fromthe resuilts showun in figuire 4 it is clear that theexchanige process mtist occuir. The excretion ofhydroxyl or bicarbonate accouints for a greateranioin thain cation tuptake with nitrate nuitritionwhile hydrogen ioIn exchange resuilts from a highercatioin absorption with uirea and amlmoniumtl1 nitrogen

supply. WN hether the process of the uiptake ofnitrate or ammoniuim ions is an active processmediated by a carrier or a passive process along anlelectro-chemical gradient is a secondary quiestionin this context. \WNhat is important is that electro-neutrality mtust be maintainied both in the plant andthe nuitrient medium.

Nitrate taken tip from the nuitrienit soltution doesnot remain in ionic form. It is reduiced and (luringthis reduiction the negative charge shifts from NO3-to OH- which increases the pH of the system.(N5+03)±811SH-+ +8e- N3+H3 + 2H10 + OH-This pH increase may lead to the accuimulation oforganic acid anions either by the dissociationi oforganic acids metabolically synthesized, as for ex-ample in the tricarboxylic acid cycle, or by theproduiction of HCO0:- from CO2. As w,as demon-strated l)y Bedri et al (4) and Chouteau (7), thepresence of bicarbonate in the nlultrient solutioinincreases the organic acid content of the plant. Itis feasible that a higher bicarbonate concentrationin the tissue enhances bicarbonate incorporationinto organic acids via CO,, fixation.

Not all bicarbonate produced dulring niitrate re-(Luction will remain in the tissuie, some will diffuiseouit of the root and increase the pH of the nuttrieintmedliuim. This is in accordance with ouir observa-tionls. The assimilation of nitrate therefore affectsthe cation-anion balance insofar as it provides acontinuiouis source of negative charges which maybe transferred to form organic aIniolns in the plantor retuirned to the nuttrienit mediuim as bicarbonateions. The assimilation of stulphate ioIs shouldI alsoproduice the same effect buit to a mulch less sig-nificant extent. \NVhether an organic anioin isformedl depenids oti whether or not the uiptake ofthe nitrate ion is accompaniied by the uiptake of a

100

e' 80 -NH4

CL 60a-

UREACLa

D

20

0N

crLULu 0c 20

C]<

40

FIG. 4. Differential remiioval of cationls anid aniionIsfromii the niutrienlt media ( tptake in meie by 10 plants).

A/

Al74.

.7s;J-~~

*/~~o

12



KIRKBY AND LMENGEL-IONIC BALANCE IN RELATION TO NITROGEN NUTRITION

cation. If no cation is taken ulp the net result isthe exchange of an equivalent amiount of nitratefrom the nutrient solution for bicarbonate from theplant.

The assimilation of ammonium ions leads to theproduction of hydrogen ions.

NH4+ -* NH3 + H+Thus with ammonium nutrition the pH of the

tissue should be lowered. This assumption is inaccordance with Mulder's observations (17) oflower pH in root and leaf tissues of peas suppliedwith ammonium rather than with nitrate nitrogen.Our own results show the same effect as can beseen in table IX. Differences are particuilarly

Table IX. Influtencc of this Form of Nitrogen Nutritionon the pH of Maccrated Tissues of Leaves, Petioles,

Stems and Roots of Tomato

Leaves Petioles Stems Roots

NO3 5.50 5.45 5.45 5.60Urea 5.5o 5.30 5.30 5.00NH1 5 00 4.70 4.80 4.70

noticeable in the root tissues at the site of ammo-

nium assimilation. As discussed above this loweredpH should reduce the accumulation of organic acidanions. This concept explains why with ammoniumnutrition the content of organic anions is low andthat of inorganic anions high. \Vith nitrate suipplythe reverse is true. Hydrogen ions produced duringammonium assimilation will also diffuse out lower-ing the pH of the nutrient medium (fig 1) andalso reduce cation uptake by competition effects(2). As urea is taken up as a molecule there is no

need for it to be balanced by other ions. The totalcations and organic anions should be intermediatebetween nitrate and ammonium nutrition. This isexactly what the results show (see tables III, V,VI). The assimilation of urea is assumed to beinduced by urease (18) splitting urea into NH3and CO. This process and the further incorpora-tion of ammonia does not result in a change in pH.Thus the pH of the tissues of this treatment shouldlie between nitrate and ammonium tissues (tableIX). In agreement with this result, the pH valueof the nutrient solution is intermediate between theother 2 treatments as is also the content of organicacids (table VI).

The urea plants had a lower total nitrogen con-

tent than the plants of the other 2 treatments. Itis probable that in this case reduced growth incomparison with the nitrate plants was caused byan insufficient supply of nitrogen. From theseresults it is impossible to say whether the limitingfactor was the uptake of urea or urease activity.In the case of ammonium nultrition the lowered pHof the tissue may have an important influence on

growth processes. The pH values of macerated

plant tissues give only a rough indication of thereal pH decrease which may occur at metabolicsites. It is possible for example that at low pH,photosynthetic CO2 fixation may be reduiced.

The effect of the different nitrogen forms onthe cation-anion balance is the same in leaves,petioles, stems and roots. Thus in addition to theresult of Coic et al (9) for leaves, it is shown thatthe form of nitrogen nutrition exerts an overalleffect on the plant. It is of interest that the upperparts of the plants contain higher concentrationsof organic acids than the roots. Calcium shows asomewhat similar distribution. The high calciumcontent of the leaves relates to the uipward trans-port of calcium ions in the transpiration stream.During this transport, calcium ions muist be accom-panied by equivalent amounts of anions. \Vithnitrate nutrition a rather high proportion of theseanions will be nitrate. At the site of nitrate re-duction which in the tomato occurs more especiallyin leaf tissue, organic anions may be accuimulatedto balance calcium ions on the metabolic removalof the accompanying nitrate ions.

The cation-anion ratios in all tissues were closeto unity, so it must be assumed that the main com-ponents responsible for this equilibriuim have beentaken into account. Other components in the plantwith positive or negative charges do occuir andinclude phosphorylated compouinds, proteins, aminoacids, hydrogen ions, and organic bases, but theircontribution to the whole balance is low. A balanicebetween cations and anions in the plant infers thatdifferent cation species compete for the bulk ofanions and vice versa. This is probably the reasonwhy the increase in suipply of 1 ion species in thenuitrient medium decreases the uptake of a similarlycharged ion species, where there is no specificcompetition for a carrier site.

Acknowledgments

The authors express their thanks to Professor T. XV.Walker, Department of Soil Science, Lincoln College,University of Canterbury. New Zealand, and to Dr. D.H. Jennings, Department of Botany, University of Leeds,England, for their interest in this +vork.

Literature Cited

1. ARNON, D. I. 1939. Effect of amiiioniiiui and ni-trate nitrogeni on the mineral and sap character-istics of barley. Soil Sci. 48: 295-307.

2. ARNON, D. I., W. E. FRATZKE, AND C. M. JOHN-SON. 1942. Hydrogen ion concentration in rela-tion to absorption of inorganic nutrients byIhiglherplants. Plant Physiol. 17: 515-24.

3. BAKER, C. J. L. 1952. The determinationi of oxa-lates in fresh plant material. Analyst 77: 340-44.

4. BEDRI, A. A., A. WALLACE, AND W. A. RHOADS.1960. Assimilation of bicarbonate bh roots ofdifferent plant species. Soil Sci. 89: 257-63.

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PLANT PHYSIOLOGY

5. BONNER, J. 1950. Plant Biochemistry. AcademicPress, New York, p 223.

6. CHOUTEAU, J. 1960. Balance acides-bases de lacomposition de plantes de tabac alimentees en azotenitrique on en azote ammoniiacal. Ann. Phiysiol.V'6getale 4: 237-47.

7. CHOUTEAU, J. 1963. Etude de la nutrition nitriqueet ammoniacale de la plante de tabac en presencedes doses croissantes de bicarbonate dans le milieunutritif. Ann. Inst. Exp. Tabac Bergerac 4:319-32.

8. CLARK, H. E. 1936. Effect of ammonium and ni-trate nitrogen on the composition of the tomatoplant. Plant Physiol. 11: 5-24.

9. Coic, Y., C. LESAINT, ET F. LE Roux. 1961. Com-parison de l'influence de la nutrition nitrique etammoniacale combinee on non avec une deficienceen acide phosphorique sur l'absorption et le me-tabolisme des anions-cations et plus particule&ere-ment des acides organiques le mais. Comparison(tL mais et de la tomate quant a l'effet de lanature de l'alimentation azot6e. Ann Phv siol.Vegetale 3: 141-63.

10. DEKOCK, P. C. AND R. I. MORRISON. 1958. Themetabolism of chlorotic leaves. 2. Organic Acids.Biochem. J. 70: 272-77.

11. DIJKSHOORN, W. 1958. Nitrate accumulation, ni-trogen balance and cation-anion ratio during re-growth of perennial ryegrass. Neth. J. Agr. Sci.6: 211-21.

12. DIJKSHOORNt, W. 1962. Metabolic regulation ofthe alkaline effect of nitrate utilization in plants.Nature 194: 165-67.

13. EHRENDORFER, K. 1964. Einfluss der Stickstof-form atif Mfineralstoffaufnahme und Substanzbil-

dung bei Spinat (Spinacca oleracca L) Bodenkul-tur 15: 1-13.

14. ERGLE, D. R. AND F. M. EATON. 1949. Organicacids of the cotton plant. Plant Physiol. 24: 373-88.

15. KIRKBY, E. A. AND P. C. DEKOCK. 1965. Theinfluence of age on the cation-anion balance inthe leaves of Brussels sprouts (Brassica oleraceavar Gemmifera) Z. Pflanzenernihr. Dung. Bo-denk. 111: 197-203.

16. MICHAEL, G. 1941. Uber die Aufnahme und Ver-teilung des Magnesiums und dessen Rolle in derh6heren grunen Pflanze. Bodenkunde u. Pflan-zenerniihr 25: 65-121.

17. MULDER, E. G. 1948. Investigationis Onl the niitro-gen nutrition of pea plants. Plant Soil 1: 179-212.

18. REINBOTHE, H. AND K. MOTHES. 1962. Urea,ureides, and guanidines in plants. Ann. Rev. PlantPhysiol. 13: 129-50.

19. SCHARRER, K. UND J. JUNG. 1955. Weitere Unter-suchungen fiber die Nahrstoffaufnahme und dasVerhaltnis von Kationen zu Anionen in der Pflanze.Z. Pflanzenernaihr Dung. Bodenk. 71: 97-113.

20. STREET, H. E. AND D. E. G. SHEAT. 1958. Theabsorption and availability of nitrate and ammonia.Encyclopaedia of P,lant Physiol. Vol. 8: 150-65.

21. TRACEY, M. V. 1948. A manometric method forthe estimation of milligram quantities of uronicacid. Biochem. J. 43: 185-89.

22. WELTE, E. UND W. WERNER. 1962. lonen aus-tancherversuche iiber die Beeinflussung der Kati-onenaufnahme der Pflanzen durch die Stickstoff-Form. Agrochimica 6: 337-48.

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