Plant Physiol. (1985) 79, 143-1480032-0889/85/79/0143/06/$0 1.00/0

Importance of Environmental pH during Root Development onPhosphate Absorption1

Received for publication March 11, 1985 and in revised form May 15, 1985

MICHAEL J. WEBB AND JACK F. LONERAGAN*School ofEnvironmental and Life Sciences, Murdoch University, Perth, Western Australia 6150

ABSTRACT

Wheat seedlings (Triticum aestivum L. cv Gamenya) were grown for4 days in culture solutions of differing pH prior to studying their subse-quent short-term absorption of 32Pi from solutions of the same or differentpH.

Increasing pH of the absorption solution from 5.5 to 7.0 or 8.0depressed 32Pi absorption from 1 and 10 micromolar Pi but had littleeffect at 100 and 1000 micromolar Pi. Increasing the pH of the culturesolution from 4.5 to 6.5 doubled or trebled subsequent 32Pi absorptionfrom nearly all absorption solutions over a wide range of Pi concentra-tions, pH, and nutrient compositions.When seedlings were transferred between culture pH treatments 4.5

and 6.5, their capacity for "Pi absorption remained unchanged for atleast 5 hours and adjusted by 60 to 80% after 24 hours and completelyafter 48 hours. This suggests that the root's capacity to absorb Piresponds to pH through slow structural changes in its mechanism of Piabsorption. P content and concentration of wheat seedlings reflected theresponse of 32Pi absorption to culture pH.

It is suggested that absorption pH affects an activity component of theprocess for Pi absorption and culture pH affects a capacity component.Failure to recognize the capacity component of the pH response explainswhy previously published results for short-term 32Pi absorption conflictwith those for long-term P accumulation in plants.

Most studies of the relationship of Pi absorption to pH havebeen done on plants previously grown under identical conditionsand then subjected to short periods of exposure to radioactivelylabeled orthophosphate (32Pi) under various conditions of Piconcentration and pH.Using this approach, Hagen and Hopkins (9) observed that

excised barley roots absorbed Pi at maximal rates from absorp-tion solutions with pH values between 4 and 5. Increasing thepH to 6.0 had relatively little effect on Pi absorption at higher Piconcentrations (10-100 ,M) but depressed it by as much as 60%at lower concentrations (1-5 AM). Further increasing the pH to7.0 and 7.7 progressively depressed Pi absorption at all Pi con-centrations. Similar results have been reported for rice roots (18)and bean (11) and white clover plants (6). These effects of Piconcentration and pH on Pi absorption by plant roots have beenattributed to changes in the concentrations ofmono- and divalentPi ions and to differences in their absorption by roots (2, 6-9,11, 18).By contrast with the effect of increasing pH in depressing 32Pi

absorption, plants grown for several weeks in culture solutionwith low and constant levels of Pi accumulated appreciably more

'Supported by the Australian Wheat Industry Research Council.

P at high than at low pH (1, 3). Clearly an anomaly existsbetween the results of short-term Pi absorption and long-term Paccumulation (3). This paper reports the results of experimentswhich resolve this anomaly; they show that, in addition to anyeffect it may have on the activity ofthe Pi absorption mechanismthrough changing ionic species of Pi, pH ofthe root environmentalso modifies the absorption of Pi by wheat plants through slowlyinduced changes in the capacity of the root mechanism for Piabsorption. Unless these changes are taken into account, short-term 32Pi absorption studies give a false representation of therelationship of pH to long-term Pi absorption.

MATERIALS AND METHODSGeneral Procedure. Except where otherwise stated in individ-

ual experiments, the following procedures were used for plantgrowth, isotope absorption, and plant analysis.

Plant Growth. Following 2 d (D22) imbibition in water, wheatseeds (Triticum aestivum L. cv Gamenya) were planted on plasticmesh in plastic cups (10 seeds/20 x 25-mm diameter cup). Threeto 12 cups, each representing a single experimental sample, wereplaced in black plastic lids over 4.8 L of aerated culture solutionof the following FN composition: KNO3, 5 mM; Ca(NO3)2, 2.5mM; MgSO4, 1 mM; KH2PO4, 1 mM; Fe EDTA, 100l m; H3B03,15 ,M; NaCl, 10 Am; ZnSO4, 2.5 ,M; CuSO4, 0.5 gm; MnSO4,0.5 AM; CoSO4, 0.2 /M; Na2MoO4, 0.1 M; pH maintained at thespecified value ± 0.5 with KOH. On D3, the pots were transferredto a growth room at 25°C with a 14-h photoperiod (1000-w high-intensity metal halide lamp; 450 AE/m2_s at the surface of thepots).Absorption Procedure. On D6, cups of plants were transferred

to aerated FN equilibration solutions of the same pH as theirsubsequent absorption pH treatment in a constant temperatureroom (20°C) with low light (8 ME/mi2s). After 1 h, cups weretransferred to 1 L of FN absorption solution containing 32Pi (orother isotope; experiment 4) with a specific activity of 185 MBq/mol P. Following absorption for 4 h, plant roots were rinsed for1 min in aerated CaB, blotted dry, and separated from shoots;roots and shoots were dried for at least 18 h at 70°C beforeanalysis.

Preparation ofPlant Materialfor Analysis. For P, Ca, Zn, orK (or their radioactive labels), the plant material was digested inHNO3 and HC104 (16). For [86Rb]K, 2 ml HCI were added tothe digestate to solubilize the K salts. For 35S, plant material wasdigested in 1 ml HNO3 at 120°C for 30 min followed by additionof 1 ml HNO3/HC104 (1/1, v/v) and digested at 220°C for 60min in long test tubes (220 x 22 mm diameter) to facilitaterefluxing of acids. For 36C1, 5 ml 5% (w/v) Na2CO3 in 10% (v/v) ethanol were added to plant samples in a 50-ml Pyrex beaker,

2 Abbreviations: DX, X days after placing seeds in water; FN, fullnutrient; CaB, 500 Mm CaSO4 plus 9 gM H3BO3; CaBP, CaB plus 1000tM H3PO4 adjusted to pH 5.5 with saturated Ca(OH)2; DM, dry matter.

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WEBB AND LONERAGAN

dried at 70°C, and ashed at 500C for 24 h (adapted from 12).The ash was neutralized with, and suspended in 5 ml 1 N HNO3.

AnalyticalProcedure. 32p, 35S, 36CI, 45Ca, and 'Rb were assayedin a Packard 3255 liquid scintillation spectrometer using 'Insta-gel' (Packard Corp.) as scintillant. 65Zn was assayed using a Searle1197 scintillation y-counter. Appropriate standards (made fromthe same diluted stock isotopes which were added to the absorp-tion solution) were assayed with the samples. Corrections forquenching were made when necessary by use of the externalstandard channels ratio technique (15). Using this approach fordigestion and analysis, isotope recoveries were greater than 95%.P was determined by a modification of the molybdovanado-phosphate method (4).

Details of Experiments. Experiment 1. The effect of culturesolution pH and absorption solution pH on 32Pi absorption wasstudied in a 3 x 3 factorial combination of the following treat-ments imposed in a randomized block design of three replicates:(a) culture solution: pH 4.5, 5.5, or 6.5; (b) absorption solution:pH 4.5, 5.5, or 6.5.Experiment 2. The effect of culture pH on 32Pi absorption

from absorption solutions of different Pi concentrations and pHwas studied in a 3 x 12 factorial combination of the followingtreatments imposed in a randomized block design of three rep-licates: (a) culture solution: pH 4.5, 6.5, or 7.5; (b) absorptionsolution: a factorial combination of 1, 10, 100, or 1000 Mm Pi atpH 5.5 or 7.0; or 1, 10, 100, or 300 Mm Pi at pH 8.0. Theabsorption period was 1 h in all treatments and 4.7 L of absorp-tion solution were used for 1 and 10 Mm Pi treatments only. Theabsorption solutions were labeled with 32P at a specific activityof 1480 MBq/mol P for 1 and 10 Mm Pi solutions and 740 MBq/mol P for 100, 300, and 1000 Mm Pi solutions.Experiment 3. The effect of culture solution pH, culture solu-

tion composition, and absorption solution composition on 32Piabsorption was studied in a 4 x 2 factorial combination of thefollowing treatments imposed in a randomized block design offour replicates: (a) culture solution, pH 4.5 or 6.5 in a factorialcombination with FN or CaB solutions; (b) absorption solutionat pH 5.5, FN or CaBP. Following germination, plants weretransferred to 1-L culture solution treatments. The pH of theCaB culture solutions was maintained at either 4.5 or 6.5 withH2SO4 or saturated Ca(OH)2.Experiment 4. The effect of culture solution pH on the sub-

sequent absorption of radioactively labeled Pi, K+, S042-, Ca2',Cl-, and Zn2+ was studied in three separate experiments each ofwhich was a 2 x 6 factorial combination of: (a) culture solution,pH 4.5 or 6.5; (b) one of the following sets of six absorptionsolution treatments at pH 5.5 imposed in a randomized blockdesign of three replicates: (a) FN solution labeled with 32p, 86Rb(181 MBq/mol K), 45Ca (185 MBq/mol Ca), 35S (185 MBq/molS), 36C1(3700 MBq/mol Cl), or 65Zn (74,000 MBq/mol Zn). (b)9 AM H3BO3 and 1,000 Mm H3PO4, 500 Mm H2SO4, or 1,000 AMHCI titrated to pH 5.5 with saturated Ca(OH)2, labeled witheither 45Ca or 32p, 35S or 36CI, and designated Ca,P; Ca,S; orCa,CI, respectively. (c) 9 AM H3BO3, 100 MM Ca(OH)2, and 1,Mm H3PO4, 500 AM H2SO4, or 1,000 AM HCI titrated to pH 5.5with KOH, labeled with either 'Rb or 32p, 35S or -6Cl, anddesignated K,P; K,S; or K,C1, respectively. The specific activitiesof the solutions (b) and (c) were: 4'Ca, 370 MBq/mol Ca; 'Rb,206 MBq/mol K; 35S, 370 MBq/mol S; 3Cl, 185 MBq/mol Cl.Immediately following the usual 1-min rinse, roots were desorbedfor a further 19 min in a solution of the same composition asthe equilibration solution.Experiment 5. The effect of altering the pH of the culture

solution during plant growth on the subsequent 32Pi absorptionwas studied in two separate experiments.Experiment Sa. A 2 x 4 factorial combination of the following

culture solution treatments was imposed on wheat seedlings in a

randomized block design of three replicates: (a) culture solutionfor 4 d: pH initially 4.5 then transferred to 6.5 or initially 6.5then transferred to 4.5. (b) days at final pH: 1, 2, 3, or 4 d. Thetreatments were imposed by placing plants over two culturesolutions on D2, one at pH 4.5, and one at 6.5 in each replicate.On D3, D4, and D5, one cup ofplants from the pH 4.5 treatmentwas interchanged with one cup from the pH 6.5 treatment ofthesame replicate. This resulted in plants having from 0 to 3 d atpH 6.5 followed by 4 to 1 d at pH 4.5 and vice versa. In alltreatments, absorption was measured in FN solution at pH 5.5.Experiment Sb. A 3 x 2 factorial combination of the following

culture solution treatments was imposed on wheat plants previ-ously grown at pH 5.5 in a glasshouse for 19 d in a randomizedblock design of three replicates: (a) culture solution: pH 4.5, 5.5,or 6.5; (b) time ofpH treatment: 1 or 2 d.

RESULTS AND DISCUSSIONEffects of pH on Growth, P Content, and P Concentration of

Wheat Seedlings. Increasing the pH of the culture solution for 4d tended to increase the dry weight of roots but not of shoots in6-d-old wheat seedlings. Differences in root weight were smalland, in individual experiments, often not significant (e.g. exper-iment 3, Table III). However, when the data from all experimentswere pooled, root weight increased by 18% (P < 0.001) as thepH ofthe culture solution increased from 4.5 to 6.5. Roots grownat pH 6.5 were generally longer and appeared to have a greatermass of root hairs than those grown at pH 4.5. Culture pH didnot affect the appearance of the shoots and absorption solutionpH had no effect on roots or shoots.Although increasing the pH of the culture solution from 4.5

to 6.5 had a relatively small effect on root dry matter, it increasedthe P content and P concentration of wheat seedlings substan-tially as, for example, in experiment 1 where it increased Pcontent by 90% and P concentration by 76% (Table I). However,while increasing P content of seedlings, increasing culture pHhad little or no effect on the distribution ofP between roots andshoots (e.g. Table I). Other workers (1, 3) have also reportedincreased P content and concentration in plants grown at in-creased pH. Thus, at some stage during the growth of theseedlings, those grown at high pH must have had a greater rateof Pi absorption than those grown at low pH. This is clearlyinconsistent with the reported depression of short-term absorp-tion of 32Pi by increasing pH above 6 (3).

Effect of pH on Short-Term 32Pi Absorption from 1000 gM Pi.Increasing the pH of the culture solution not only increased therate of Pi absorption by wheat seedlings while they were in thatsolution, but also enhanced the rate of 32Pi absorption in asubsequent short-term absorption period. This enhancement wasindependent of the conditions imposed during the absorptionperiod. Thus, increasing the pH of the culture solution from 4.5to 5.5 and 6.5 enhanced subsequent 32Pi absorption by roots ofwheat seedlings by nearly 50 and 150%, respectively. By contrast,increasing the pH of the absorption solution from 4.5 to 6.5 hadno effect on the 32Pi absorption of any culture pH pretreatment(Fig. 1).The effects of culture solution pH on the 32p content of shoots

were similar to its effects on 32Pi absorption and on the contentand concentration of total P in seedlings after 4 d (Table I). Itseffects on 32Pi absorption therefore appear to reflect true absorp-tion rather than simply exchange with unlabeled Pi in the roots.This conclusion is supported by the results ofan experiment withseedlings grown in the absence of any Pi; such plants grown atpH 6.5 had identical P status to those grown at pH 4.5 (TableIII), yet responded to culture pH in their 32Pi absorption andtransport in the same general way as did those grown in thepresence of Pi (Fig. 2).The failure of 32Pi absorption to respond to increasing pH of

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ROOT ENVIRONMENTAL pH AND PHOSPHATE ABSORPTION

Table I. Effect ofCulture Solution pH on the P Content and Concentration in Wheat Seedlings, theProportion of Total P in the Shoots, and the Proportion and Amount of32P Transported to the Shoots in 6-d-

old Wheat SeedlingsExperiment 1. Germinated wheat seedlings were grown for 4 d in culture solutions containing all nutrients

including 1000 Mm Pi. Culture solution pH treatments were imposed in triplicate. From each replicate pot ofeach culture solution pH treatment, wheat seedlings were transferred to each ofthree absorption pH treatments.As absorption pH treatments had no effect (P > 0.05) on any parameter in the table, the data for absorptionpH treatments were pooled to give means + SE of nine values for each culture solution pH treatment.

p 32PCulture Content Proportion TransportSolution Concentration in to

Total Shoot shoot shoot

pH % DM Mg/plant % oftotal % oftotal gmol/gDMroot 4h4.5 1.01 ± 0.02 174 ±4 61.4 ± 0.6 22.1 ± 1.0 9.4 ± 0.55.5 1.49 ± 0.01 275 ± 8 58.2 ± 0.3 21.5 ± 0.6 13.6 ± 0.46.5 1.78 ± 0.04 332 ± 5 58.4 ± 1.2 21.7 ± 1.4 22.0 ± 2.1

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ABSORPTION SOLUTION pHFIG. 1. Effect of culture solution pH (4.5 [0]; 5.5 [0]; 6.5 [x]) for 4

d on the subsequent 32Pi absorption from full nutrient solution with 1000Mm Pi at pH 4.5, 5.5, or 6.5. Values are means of three replicates: wherethey exceed the size of the symbols, the SE are shown as vertical bars.Experiment 1.

the absorption solution at 1000 Mm Pi in the present experiment,is in agreement with an earlier experiment with excised barleyroots, which at the highest concentration used (100 ,uM), showedlittle response to changing the absorption solution pH from 4.0to 6.0 (9). It is also consistent with the observation that Piabsorption by wheat seedlings is not responsive to changingconcentrations of monovalent Pi ions from 50 to 1000 Mm (8).That absorption pH had no effect on 32Pi absorption of any

culture pH treatment means that the effect of culture pH treat-ment persisted for 5 h after changing the pH environment of theroots. This suggests that culture pH affected some structuralcomponent ofthe absorption mechanism. In its ability to inducea lasting change in 32Pi absorption, the effect ofpH ofthe culturesolution on wheat roots resembles that of Pi supply in barleyroots. In barley roots, Clarkson et al. (5) have shown that anearly period of P stress, induced by low P supply, enhancedsubsequent 32Pi absorption and transport to the shoot. P stresswas not a factor in the response of wheat roots to culture pH inthe present experiment since all plants had high levels of P and,moreover, the effects ofculture pH on P status and on subsequent32Pi absorption proceeded in the same rather than oppositedirections.

Effect of pH on Short-Term 32Pi Absorption from 1 to 1000MM Pi. At each concentration from 1 to 1000 Mm Pi and eachpH of the absorption solution, increasing from 4.5 to 6.5 the pH

of the culture solution in which plants were grown for 4 dsubstantially increased subsequent 32Pi absorption generally dou-bling or trebling it (Table II). Increasing the culture pH to 7.5had more variable effects, sometimes increasing.subsequent 32Piabsorption further and sometimes having no further effect. How-ever, in all cases, with the exception ofabsorption from a solutionofpH 8.0 at 1 uM Pi, increasing pH of the culture solution from4.5 to 7.5 at least doubled or trebled subsequent 32Pi absorption.The results thus establish that the pH of the root environmentduring growth of wheat seedlings is an important factor in theirsubsequent capacity to absorb 32Pi over a wide range of Pi andhydrogen ion concentrations in the absorption solution.The results also establish that in their 32Pi absorption, the

response of wheat roots to an earlier culture pH treatment isindependent of and additional to their response to the pH andPi concentration of the absorption solution (culture pH byabsorption pH interaction P> 0.05). The present results showingno effect of increasing absorption solution pH above 5.5 on 32Piabsorption from 1000 and 100 Mm Pi (P > 0.05), but markedeffects on absorption from 10 and 1 Mm Pi (P < 0.05) areconsistent with earlier observations in wheat seedlings where 32Piabsorption responded weakly to increasing concentrations ofmonovalent Pi ions from 50 to 1000 Mm (8) but strongly to

Table II. Effect ofCulture Solution pH on the Subsequent 32PiAbsorption by 6-d-old Wheat Seedlingsfrom Absorption Solutions of

Different pH and Pi ConcentrationValues are means ± SE of three replicates. Experiment 2.

Absorption Solution Culture Solution pH

concePration pH 4.5 6.5 7.5

MM Mmol 32Pi absorbed/gDM root- h1 5.5 7±3 21 ±9 16±3

7.0 5±2 14±8 12±78.0 4±2 12+5 6 1

10 5.5 25±6 53±4 94±227.0 9+2 51 17 49 148.0 9±3 32+8 33 13

100 5.5 28±4 49±6 97±257.0 27± 1 65±3 118 168.0 31 ±6 91 ± 10 118±24

300 8.0 27±2 73± 10 104±31000 5.5 40±3 82± 14 149+31

7.0 34±2 94±22 175±21

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WEBB AND LONERAGAN

decreasing concentrations below 50 uM. In other species as wellas wheat, 32Pi absorption has been observed to decrease rapidlywith decreasing concentrations of monovalent Pi ion concentra-tions below 50 Mm brought about by either depressing Pi concen-tration at pH 5.6 (8) or by increasing pH at fixed Pi concentration(6, 9).The fact that 32Pi absorption responds to culture solution pH

in addition to absorption solution pH suggests that pH affectstwo distinct processes. The process affected by the culture solu-tion pH involves slow adjustment of the absorption mechanismand may result from structural changes affecting the root's ca-pacity to absorb Pi, such as, for example, by influencing thenumber of absorption sites. The process affected by the absorp-tion solution pH involves immediate adjustment of the activityofthe Pi absorption mechanism possibly through conformationalchanges in the absorption sites, or through concentrations ofsubstrate(s) or inhibitor(s) of Pi absorption. Clearly the resultantrate of Pi absorption depends on the effects of pH on both thecapacity and activity components of the Pi absorption process.

Characteristics of the Culture Solution pH Effect on Subse-quent Pi Absorption. Effect ofNutrient Composition on the 32PiAbsorption Response to pH. Changing the composition of eitherthe culture solution or the absorption solution from a completeFN solution to a simple CaB or CaBP solution generally de-pressed but did not change the nature of the 32Pi absorptionresponse to culture solution pH (Fig. 2). These results suggestthat H+ ions influence the capacity component of the Pi absorp-tion process either directly or through an interaction with Ca, S,or B.Changing the culture solution from FN to CaB also depressed

plant growth (Table III) suggesting that CaB grown plants weredeficient in one or more nutrients. This may account for theirlower subsequent 32Pi absorption. The depression of 32Pi absorp-tion observed when the absorption solution composition waschanged from FN to CaBP, can probably be attributed to differ-ences in the ionic strength of the absorption solutions since ionicstrength is known to affect rates of absorption of ions includingPi (17, 19).The proportion of 32p in the shoots responded weakly to

culture solution pH and absorption solution nutrient composi-tion but strongly to culture solution nutrient composition (TableIII). The plants grown in CaB culture solution transported only7 to 12% of the total 32Pi absorbed to their shoots, whereas thosegrown in FN culture solution transported 20 to 39%. The reten-tion of P by tissues low in P is well known (13) and this may

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CULTURE SOLUTION pHFIG. 2. Effect of culture solution pH and composition on subsequent

32Pi absorption from solutions with 1000 Mm Pi and of different compo-sition. Values are means of four replicates: where they exceed the size ofthe symbols, the SE are shown as vertical bars. Experiment 3. (-), CultureFN, absorption FN. (0), Culture FN, absorption CaBP. (U), CultureCaB, absorption FN. (0), Culture CaB. absorption CaBP.

explain the depressed transport of P by seedlings grown in CaB.But while CaB culture treatment depressed P transport, bothabsorption and transport responded to culture pH in the sameway as they did in FN treatments.

Effect ofpH on the Absorption ofOther Ions. While increasingthe culture solution pH from 4.5 to 6.5 increased the subsequentabsorption of 32Pi in all absorption solutions, it affected thesubsequent absorption of other labeled ions in various ways(Figs. 3 and 4). It had no effect on [35S04]2- absorption, depressed[86Rb]K' absorption, and enhanced both [45Ca]2" and [65ZnJ2+absorption; it enhanced [36C1]- from 10 Mm Cl- in EN and from1000 gM Cl- in K,Cl but depressed it from 1000 AM Cl- in Ca,Cl.This result implies that any structural response induced by pHtreatment of the roots and affecting their capacity to absorb 32Pishows some degree of specificity to Pi absorption.

Critical Timefor the Induction Phase. Whatever the nature ofthe changes induced in the subsequent 32Pi absorption of wheatroots by culture pH, the present results show that they requirean induction period of between 1 and 2 d to respond fully.Changing the pH of the culture solution from 4.5 to 6.5 (andvice versa) for 2 d completely reversed the effect of the previouspH treatment on subsequent 32Pi absorption and transport toshoots by 6-d-old wheat seedlings (Fig. 5). Although this capacityresponse is slow compared with the activity response, it is quiterapid when compared with root growth.The concentration of 32Pi in roots (total absorption minus

transport to shoots in Fig. 5) responded most rapidly, beingcompletely reversed after only 1 d of reversing treatment pH.Absorption of 32Pi responded to altered pH more rapidly thandid transport, taking only 1 d of treatment to be 78% complete.The changing rates of 32Pi absorption induced by altered culturesolution pH were reflected, but more slowly, in the long-term Paccumulation (Fig. 5), establishing that the measured 32Pi ab-sorption rates are representative of net Pi absorption.The faster response of 32Pi absorption compared with transport

may infer two sites of action of pH. Alternatively and morelikely, the culture pH probably only affects the absorption of Piwith its subsequent effect on transport being a function of, andcontrolled by, the P accumulation in the roots, but which takeslonger to adjust than the absorption mechanism.The effects presented here are not restricted to young seedlings

because in 20-d-old wheat plants, as in seedlings, 32Pi absorptionand transport to shoots responded to changing culture pH (TableIV); after 1 d of transferring plants from pH 5.5 (median value)to pH 6.5 culture solution, the subsequent 32Pi absorption andtransport to shoots was greater than by those transferred to pH4.5. The same pattern was also evident 2 d after transfer. Theseobservations that the capacity of wheat plants to absorb Pi isslowly altered by changing the pH of the root environment andis of greater consequence than the effects of pH on the short-term absorption activity have important implications for thelong-term P nutrition of plants.

DISCUSSIONThe present results establish that the pH ofthe culture solution

induces a slow response in roots which modifies their capacityfor Pi absorption and that this is an important factor in subse-quent absorption of 32Pi and long-term P nutrition of wheatplants. Under the conditions of the present experiments, theeffect of culture pH on the capacity of 32Pi absorption was muchgreater than, and additional to the effect of the absorptionsolution pH on the activity of 32Pi absorption. Moreover, themarked response of plants to culture pH appears to be relevantto plants growing in a wide range of conditions since it wasobserved in seedlings grown at low and high nutrient supply, in20-d-old plants as well as seedlings, and in absorption solutionsof varying nutrient composition and over a wide range of pH

0

S U

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ROOT ENVIRONMENTAL pH AND PHOSPHATE ABSORPTION

Table III. Effect of Culture Solution pH and Nutrient Composition on Dry Matter and P Concentration of6-d-old Wheat Seedlings, and on the Proportion of32P Transported to the Shootfrom Absorption Solutions of

Different CompositionValues are means ± SE of four replicates. Experiment 3.

Culture Solution Absorption Dry Matter P Concentrationa 32pSolution Proportion

Composition pH Composition Root Shoot Root Shoot in Shoot

mg/plant % DM % oftotal

FN 4.0±0.1 8.3±0.2 20± 14.5 CaBP 3.9 ± 0.2 8.0 ± 0.4 30 ± 1

FN 4.3 ± 0.4 8.3 ± 0.4 1.36 ± 0.04 1.40 ± 0.02 27 ± 16.5 CaBP 4.2 ± 0.2 8.4 ± 0.5 39 ± 1

FN 3.3±0.2 5.3±0.4 7± 14-5 CaBP 3.5 ± 0.2 5.7 ± 0.1 0.34 ± 0.00 0.67 ± 0.01 12 ± 1CaB FN 3.3 ± 0.2 5.2 ± 0.0 8 ± 16.5 CaBP 3.6 ± 0.1 5.6 ± 0.2 0.35 ± 0.02 0.65 ± 0.02 12 ± 1

ap concentrations are those of seedlings prior to absorption period.

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Absorption Solution CompositionFIG. 3. Effect of culture solution pH on the subsequent absorption of

anions. Values are means + SE for three replicates of the absorption ofeach anion (gmol/g DM root.4 h) by wheat seedlings cultured at pH 4.5expressed as a percentage of its absorption by seedlings cultured at pH6.5. Ions measured were 32Pi, [35so42-, and ['Cl]-. Details ofabsorptionsolutions are given in "Materials and Methods." Experiment 4.

and Pi concentrations.Although the present experiments establish that culture pH

induced a large and general effect on P nutrition ofwheat plants,they say little about the nature of the phenomenon. They do,however, indicate that the changes induced by pH are specificfor Pi absorption and probably involve changes in the capacityof the Pi absorption mechanism rather than changes in overallabsorption parameters, such as, for example, surface area ofroots. As no appreciable changes occurred within 5 h, it is clearthat the culture pH effect did not act through immediate confor-mational alterations in the absorption sites or rapid metabolicchanges such as enzyme or absorption site activation. It is morelikely that culture pH induced some metabolic changes whichresulted in structural modifications that ultimately affected thecapacity of the Pi absorption mechanism. Such modificationsmight involve changes in the properties of the root cell mem-

o 1500~

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FIG.40 %3z Li..ZCLMAbsorption Solution CompositionFIG. 4. Effect of culture solution pH on the subsequent absorption of

cations. Values are means + SE for three replicates of the absorption ofeach cation (umol/g DM root-4 h) by wheat seedlings cultured at pH4.5 expressed as a percentage of its absorption by seedlings cultured atpH 6.5. Ions measured were [45Ca]f+, ['Rb]K+, and [65Zn]2+. Details ofabsorption solutions are given in "Materials and Methods." Experiment4.

branes which have been postulated to account for the change in32Pi absorption induced by P stress in barley seedlings (5).The separate responses to pH of the capacity and the activity

components of Pi absorption in wheat roots could be expectedto act simultaneously in plants grown for sustained periods inenvironments of constant pH values. In such plants, the largerresponse to environmental pH of the capacity component of thePi absorption mechanism of wheat would override the smallerresponse of the activity component. Recognition of this fact

147

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WEBB AND LONERAGAN

80

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FIG. 5. Effect of ct6.5 (0, ,A); pH 6.5(0, 0), 32P transportseedlings. Absorption Iare means of three repthe SE are shown as ve

Table IV. Effect of(2 d on the SubsequeiGrown for 19 d in FuBoth culture and al

are means ± SE of thr

Days ofTreatment

2

resolves the appareresults of long-termof pH on Pi absorpother plant specieswheat (1, 3) suggescomponent of Pi atto other species.

In addition to itsof Pi absorption to

tions for other aspects of P nutrition. For example, changes inPi absorption during or following treatments imposed duringgrowth may result more from effects of the treatments on rhi-zosphere pH than from any other direct effect on Pi absorptionitself.The present results may also be relevant to the complex

problems ofplants absorbing Pi from acid soils. Acidity is knownto depress Pi absorption by plants in some soils (10). It hasgenerally been attributed to an indirect effect ofpH in increasingthe concentration of aluminum ions in the soil solution whichdepresses absorption of Pi by plants (10). While increasing con-centrations ofaluminum do depress Pi absorption by plants (14),the present results show that a direct effect of acidity on rootstructure or metabolism may also be involved and may need tobe considered in any program for developing plant cultivarssuitable for acid soils.

LITERATURE CITD

1. ALAM SM 1981 Effects ofsolution pH on the growth and chemical composition__ _ __ _ __ ___I of rice plants. J Plant Nutr 4: 247-260

. . * .s 2. BIELESKI RL, IB FERGUSON 1983 Physiology and metabolism ofphosphate and4 3 2 1 0 its compounds. In A Lauchli, RL Bieleski, eds, Encyclopedia of Plant

DAYS AT INITIAL pH Physiology. Inorganic Plant Nutrition, Vol ISA. Springer-Verlag, Berlin, pp~~423-4 9,_____ ,_____ ,_____._____ .____ 3. BLAMEY FPC, DG EDWARDS, Cl ASHER, Moo-KEY KIM 1982 Response of

0 1 2 3 4 sunflower to low pH. Scaife, ed, Plant Nutrition 1982. Proceedings of

the 9th International Plant Nutrition Colloquium. Commonwealth Agricul-DAYS AT FINAL pH tural Bureaux. Slough, United Kingdom, pp 66-77

4. BOLTZ DF, CH LUECK 1958 Phosphorus. In DF Boltz, ed, Colorimetrichanging the pH of the culture solution (pH 4.5 to determination of non-metals. Interscience, New York, pp 29-46to 4.5 (0, 0, A) on the subsequent 32pi absorption 5. CLARKSON DT, J SANDERSON, CB SCATrERGOOD 1978 Influence ofphosphate-(U, 0), and P content (A, A) of 6-d-old wheat stress on phosphate absorption and translocation by various parts of thefromfullnutrient solution with 1000,um Pi. Values roots system ofHordeum vulgare L. (barley). Planta 139: 47-53

from full nutrient solution with 1000 iSM Values 6. DUNLOP J, DJF BOWLING 1978 Uptake of phosphate by white clover. II. The

Aicates: where they exceed the size of the symbols, effect of pH on the electrogenic phosphate pump. J Exp Bot 29: 1147-1153-rtical bars. Experiment Sa. 7. EDWARDS DG 1968 The mechanism of phosphate absorption by roots. In JW

Holmes, ed, Transcripts of the 9th International Congress on Soil Science,Adelaide, Vol 2. The International Society of Soil Science and Angus and

1hanging the pH ofthe Culture Solution for and Robertson Ltd., Sydney, pp 183-190nt 32Pi Absorption of Wheat Seedlings Previously 8. EDWARDS DG 1970 Phosphate absorption and long-distance transport in wheatll Nutrient Solution at a Median Value ofpH 5.5 seedlings. Aust J Biol Sci 23: 255-264

9. HAGEN CE, HT HOPKINS 1955 Ionic species in orthophosphate absorption bybsorption solutions contained 1000 AM Pi. Values barley roots. Plant Physiol 30: 193-199

ee replicates. Experiment Sb. 10. HA aNEs RJ 1982 Effects of liming on phosphate availability in acid soils. ATreatment Total32pi 32P Transport critical review. Plant Soil 68: 289-308

Treatment Total 32Pi 32P Transport 11. HENDRIX JE 1967 The effect of pH on the uptake and accumulation ofpH Absorption to Shoot phosphate and sulfate ions by bean plants. Am J Bot 54: 560-564

12. HORwITz Wed 1975 Official Methods ofAnalysis ofthe Association ofOfficialjimolig DM root- 4 h Analytical Chemists, Ed 12. Association of Official Analytical Chemists,

4.5 49 ± 1 9 ± 1 Washington, DC, p445.5 54 ± 3 12 ± 0 13. LONERAGAN JF 1978 The physiology of plant tolerance to low phosphorus

6.5 59 ± 1 14 ± O availability. Jung, ed, Crop Toleranceto Low Phosphorus Availability.

- American Society of Agronomy, Houston, pp 329-3434.5 45 ± 4 7 ± 0 14. MUNNS DN 1965 Soil acidity and growth ofa legume. II. Reactions aluminium

and phosphate in solution and effects of aluminium, phosphate, calcium,5.5 49 2 9 0 and pH on Medicago sativa L. and Trifolium subterraneum L. in solution

6.5 57 ± 2 13 ± I culture. Aust J Agric Res 16: 743-75515. Packard Instrument Co., Inc. 1977 Instruction manual. Model 3255 Tricarb

Liquid Scintillation Spectrometer System. Manual 2136. Downers Grove,Dnt anomaly that has existed between the ILand short-term experiments on the influence 16. REUTER DJ, AD RoBSON, JF LONERAGAN, DJ TRANTHIM-FRYER 1981 Copper)tion (3). That long-term P accumulation by nutrition of subterranean clover (Trifolium subterraneum L. cv. Seaton

responds to culturepHin a similarwayto Park). II. Effects ofcopper supply on distribution ofcopper and the diagnosisresponds to culture pH in a similar way to ofcopper deficiency by plant analysis. Aust J Agric Res 32: 267-282

sts that the response to pH of the capacity 17. ROBSON AD, DG EDWARDS, JF LONERAGAN 1970 Calcium stimulation of)sorption reported here for wheat may apply phosphate absorption by annual legumes. Aust J Agric Res 21: 601-612

18. TAN VAN HAl, H LAUDELOUT 1966 L'absorption des phosphates par les racinesde riz. Ann Physiol Veg 8: 13-24

intrinsic interest, the response of the process 19. VIris JR FG 1944 Calcium and other polyvalent cations as accelerators of ionculture solution pH has important implica- accumulation by excised barley roots. Plant Physiol 19: 466-480

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148 Plant Physiol. Vol. 79, 1985

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