PLANT PHYSIOLOGY physiology volume27 july, 1952 number3 the uptake of phosphorus by bean plants with par-ticular referenceto theeffects of iron 0. ... results and discussion

PLANT PHYSIOLOGYVOLUME 27 JULY, 1952 NUMBER 3

THE UPTAKE OF PHOSPHORUS BY BEAN PLANTS WITH PAR-TICULAR REFERENCE TO THE EFFECTS OF IRON

0. BIDDULPH AND C. G. WOODBRIDGE

(WITH FOUR FIGURES)

Received September 21, 1951

This paper presents some of the findings of a study of the uptake ofphosphorus by bean plants as it is influenced by iron. As a result of thiswork a number of points have been brought out which have not been clearlystated previously. In order to complete the study successfully, the effectsof pH and the simultaneous effects of various phosphorus and iron supplieswere considered. The results show: (a) Absorbed iron ties up a portion ofthe " seed phosphorus " in an unusable condition. (b) A ferric phosphateprecipitate, in or on the roots, retards the flow of phosphorus to the activelygrowing leaves. (c) As the phosphorus content of the nutrient medium isincreased, roots, stems, and cordate leaves continue to build up in phos-phorus content even after trifoliate leaves are being adequately supplied.The excess phosphorus may be responsible for immobilizing iron and otherions.

The effect of the hydrogen ion concentration on the absorption of phos-phorus by higher plants was studied by ARNON et al. (2) as part of a morecomprehensive investigation of the absorption of inorganic nutrients. Theyshowed that the amount of phosphate absorbed varied both with the plantand also with the pH of the nutrient solution. With tomato, maximum ab-sorption occurred at pH 7, decreasing toward pH 3 and pH 9. With Bermudagrass there was no marked variation in the phosphorus absorption betweenpH 4 and pH 7, although a tendency toward a minimum appeared at pH 6.At a pH of 3 and pH 9, which are extremes under which plants barely per-sist, the phosphorus absorption was much lower. Because of this variationbetween plants, a study was undertaken of the absorption of phosphorus bybean plants grown in water cultures maintained at various pH values.

The effect of iron on the absorption of phosphorus has not been studieddirectly. However, relationships between the two have been reported by anumber of workers. AIYAR (1), using solution cultures, found that increas-ing concentrations of phosphorus caused an increase in the phosphorus con-tent of the roots, but a decrease in the nitrogen and iron contents. THONIAS

431

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PLANT PHYSIOLOGY

et al. (14) showed that phosphatic fertilizer reduced the absorption of ironby Calluna vulgaris. SIDEMtIS et al. (11) reported that in Ananas cornosusan increased supply of phosphorus increased slightly the amount of ironprecipitated on the plant's roots. These papers, while dealing more or lessindirectly with the effect of phosphorus on the absorption of iron, do notpresent a thorough elucidation of the phenomenon.

The relationship of phosphorus to chlorosis has been the subject of con-siderable study and a few papers have contained information of direct bear-ing on it. OLSEN (8, 9) demonstrated that certain plants (Lemna poly-thiza, Zea mays, and Xanthium spinosurn), when grown in water culturesolutions (Knop's solution at pH 7.0) having a high phosphate content butnormal iron content, became chlorotic even though the iron content of thechlorotic leaves compared favorably with that present in green leaves.WADLEIGH et al. (15) grew corn plants in solutions high in phosphate andfound that these plants were also severely chlorotic. FRANCO and LOOMIS(6) compared the absorption of phosphorus and iron from various nutrientsolutions and found that true iron-deficiency chlorosis could be prevented ifphosphorus was withheld for two to four days from the nutrient solution atthe start of an experiment.

Methods

Red kidney bean seeds (Phaseolus vulgaris) were dusted with Semesan(active fungicidal ingredient, hydroxymercurichlorophenol) and placed be-tween dampened paper towelling in a shallow glass dish where they re-mained for two days. The radicles from viable seeds were then evident, andthe sprouted seeds were transferred to glass beads partially immersed in tapwater. The seedlings remained on the glass beads for five to six days.During this time, the hypocotyls grew sufficiently so that the young beanplants could be transferred to half-strength nutrient solution contained inenamelled pans of six liters capacity. The plants were supported by wrap-ping non-absorbent cotton about the stems and inserting them into slots cutin Lucite covers. Covers were painted with black asphaltum varnish onthe lower surface and with white paint on the upper surface. Each pan con-tained six plants. The day of transfer to the pans was always designatedas zero day, and periods in solution and age of plants were calculated fromthis base. The solutions were changed to full-strength on the fourth dayand changed again on the eighth day. Harvests were made on the twelfthday unless stated otherwise.

The nutrient solutions were varied with respect to their phosphorus con-tent, iron content, and pH. The variations are given with each experiment.The pH of each solution was adjusted twice daily (early morning and lateafternoon) by the addition of potassium hydroxide solution or sulphuricacid solution. The basic nutrient medium had the following composition:

Ca(NO3)2-0.0025 M H3BO3-0.5 p.p.m. BKNO3 -0.0025 M CUSO4-0.02 p.p.m. Cu

432



BIDD'ULPH AND WOODBRIDGE: 'UPTAKE OF PHOSPHORUS BY BEAN 433

MgS04 -0.001 M MnCI2-0.5 p.p.m. MnKH2P04 -0.00025 M (except as varied) MoO3 -0.01 p.p.m. MoFe(N03)3 -1.0 p.p.m. Fe (except as varied) ZnSO4-0.05 p.p.m. Zn

The solutions were continuously aerated by means of carbon pipe aeratorsand comiipressed air at three pounds pressure. Sunlight was supplementedduring the winter months with daylight fluorescent lamps, but the intensitywi-as still relatively low as compared to sunlight.

Leaves were harvested according to position upon the steini. The petioleswere considered as part of the stem. Tissues were dried at 80° C to con-stant weight, ground, and the aliquots weighed for analysis.

An aliquot of five mg. of dried tissues was used for analysis for totalph1osphorus (P31). This amount was transferred to a 10-ml. Pyrex volli-metric flask and was digested in one ml. of 5 N sulphuric acid solution andone drop of 60% perchloric acid solution. A sand bath was used, and aboutthree lhours were required for the digestion. After the solution had cleared,the flask was cooled, and 7 ml. of water were added followed by the reagentsfor development of the well-known phosphomolybdate blue (12). The in-tensity of the color was measured in a photelometer using a red (610-P)filter and was compared with standards prepared at the same time.

When radiophosphorus was to be determined, an aliquot (20 to 100 mg.)of dried material was weighed and transferred to a 30-ml. porcelain cruci-ble. The sample was ashed directly in a muffle furnace at dull red heat andthen cooled. The ash was dissolved in dilute hydrochloric acid to insureuniform spreading in the crucible. The solution was evaporated to drynessand counted. The geometry factor was constant for all determinations.Errors due to self-absorption, coincidence phenomena, and counter efficiencyw-ere low in comparison to the statistical errors of counting and were ig-nored. The quantity of radiophosphorus used in each experiment was cal-culated so that the net number of counts in the weakest sample wouldexceed twice the background. Statistical examination of the counting pro-cedure (7) indicates that the probable error of the results is not greaterthan 10% in cases of low activity and in the order of 5%v for higher activity.

Results and discussionTHE EFFECT OF PH UPON THE ABSORPTION OF PHOSPHORUS

It is known that the hydrogen ion concentration of a nutrient solutionexerts an effect both upon the nutrient solution and upon the roots of plantsgrowing in it. Within the nutrient solution the form of the phosphate ionis determined bv the pH, with H2P04- predominating below approximately6.7 and HP04: above that value (fig. 1). The hydrogen ion concentrationalso exerts an effect upon the permeability of the membranes of the absorb-ing cells. This is a comiplex phenomenon, and its discussion will not be at-teil)pted here. As it has been shown by ARNON et al. (2) that pH influencesphosphorus absorption differently in different plants, it was necessary to de-termiaine its effect on phosphorus absorption by bean plants.



434 PLANT PHYSIOLOGY

The problem of phosphorus absorption at different pH values was in-vesti, ated by measuring the uptake and the resultant distribution of thiselem At between the various plant organs. The pH values employed were4, 5, and 7, each under conditions of no iron supplementation and undercond ns wherein the solutions contained 1 p.p.m. Fe (0.000018 M). The

.9 me. OF H2P10 CHEMICALLY COMBINED WITH Fe

_ IONIC fORM OF P (CALC.)H2P°4\

_.0002 3

-.0001

HPO4,'

_10en-, _̂ ROOTS4 . ~~~~~~~~~~~~~~~~~~~~~~~~~~~~.-.....

Z _8X0.z .at C. LEAVES'-6

o T. LEAVESE 4

e. ..-- - STEMS

EE

4 5 6 7 8pH OF NUTRIENT MEDIUM

FIG. 1. Above: The amount of H2PO4- which is chemically combined with iron atv-arious pH levels accordifig to Swenson, Cole, and Sieling (reproduced by permission ofthe copyright holders) and the icnic form of phosphorus (calculated) expressed in molarconcentrations at various pH levels. Below: The concentration of phosphorus in variousparts of four groups of bean plants, each group grown in a similar nutrient solution(containing iron) but maintained at a different pH level. Results for each pH level arefrom two separate pans of six plants each. The curves represent the average value ateach pH. C. Leaves, cordate or unifoliate leaves. T. Leaves, trifoliate leaves.

results are expressed graphically in figure 1. Several forms of iron havebeen employed with little difference in the amount and distribution of ironin the aerial parts of the plant. Ferric nitrate was used in this study.

The results indicate that there exists a characteristic uptake of phos-phorus for each pH level of the nutrient medium. Total uptake is highest



BIDDULPH AND WOODBRIDGE: UPTAKE OF PHIOSPHORUS BY BEAN 435

froml a mediumii of pH 4.0 with a minimum occurring at or near pH 5.0. Asecond maxinmmui occurs at or near pH 6.0 with a general decrease at l-igherpH. The behavior of individual parts is interesting, but it should be,- earlystated that the root concentrations include both absorbed and a rbedphosphorus since we have not found a successful mieans of distin .t ihingbetween them. Our root values are, therefore, without great signif Lnce asto absorbed phosphorus. Stem concentrations decrease with decreasingacidity to pH 6.0, but they show an increase between pH 6.0 and 7.0. Leafconcentrations decrease markedly at pH 6.0 to 7.0. This indicates thatmovement of phosphorus from stems and petioles to leaf blades is impairedat pH 7.0. The resultant accumulation of phosphorus in stems and petiolesat pH 7.0 constitutes a medium rich in phosphorus through which other ionsbeing transported to the leaf blades must pass. Under conditions of higlpH (7.0) and with accumulation of phosphorus, some difficulty in the suc-cessful passage of iron ions to the blades has been found (10).

As a result of this and other studies, a pH of 6.0 has been adopted as asuitable pH at which to grow this variety of bean while attempting to studythe effect of iron upon the uptake of phosphorus. At this acidity and withthe nutrient solutions employed, very little drift in pH occurs and there areno unusual effects of pH upon the internal distribution of phosphorus be-tween plant parts.

PHOSPHORUS UPTAKE AS A FUNCTION OF NUTRIENT PHOSPHORUS

CONCENTRATION

We have found it possible to grow beans at a wide range of phosphoruslevels. The lower and upper limits may be defined respectively as that con-centration at which phosphorus deficiency occurs, and as that concentrationat which chlorosis develops. In general, phosphorus deficiency symptomsare slow to develop because of the extreme mobility of phosphorus withinthe plant (3) and it is possible for metabolically active tissues to continuetheir growtlh at the expense of phosphorus in other parts of the plant. Thephosphorus requirement of beans is not high. Levels as low as 0.00005 -Msupplied at the rate of one liter per plant per four days (less than 0.4 mg.P,1plant/day) have proved sufficient for plants having several trifoliateleaves. At the upper extreme, chlorosis becomes a problem in bean plantsgrowing in a solution in which the phosphorus concentration is 0.005 A.Our experiments on phosphorus uptake were designed to: (a) demonstratethe tolerable levels of phosphorus both in the presence and absence of nutri-ent iron; (b) furnish information on the nature of the relationship betweennutrient phosphorus and tissue phosphorus; and (c) furnish a basis for theselection of proper phosphorus levels to be employed in further experimentsdealing with uptake and metabolism of phosphorus.

Experimental plants were grown at four levels of phosphorus, 0.00000,0.00005, 0.0005, and 0.005 molar. At each of these phosphorus levels, twoiron levels, 0.00000 and 0.000018 molar (1 p.p.m.), were used. The plants



PLANT PHYSIOLOGY

received half-strength nutrient solution on the first day, and on the fourthand eighth days, the solutions were replaced with fresh full-strength nu-trient. The pH of all nutrient solutions was maintained at or near 6.0.The plants were harvested when the first indications of chlorosis wereclearly visible in those plants receiving 0.005 M phosphorus with no supple-mentary iron (10 days). At the time of harvest, the leaves of the plantsgrown with no supplementary phosphorus were a dark bluish-green whichis typical of phosphorus deficiency in this species. The results are showngraphically in figure 2.

50 so

I PPM. Fe R NO Fe SUPPLEMENTATION

,/'

z 1.0 _- 1.0_,

_ ~ PHSHOU COTNOFNTINTMDU

TL

innurin souin cotiigdfeetaonso hshrsad1ppm eAB

0 .00005 .0005 .005 0 .00005 .0005 .005 M

PHOSPHORUS CONTENT OF NUTRIENT MEDIUM

FIG. 2. A. The concentration of phosphorus in various parts of bean plants grownin nutrient solutions containing different amounts of phosphorus and 1 p.p.m. Fe.Results are from two separate pans of six plants each. The iron concentration (1 p.p.m.)would be equimolar with phosphorus at the position of the arrow on the abscissa.B. The concentration of phosphorus in various parts of bean plants grown in nutrientsolutions containing different amounts of phosphorus, but with no iron supplementation.Results are from two separate pans of six plants each. R, roots; S, stems; CL, cordateleaves; TL, trifoliate leaves.

PLANTS GROWN WITH 1 P.P.M. IRON.-A ranking of plant parts with re-gard to their phosphorus concentration showed roots to be highest, with tri-foliate and cordate leaves somewhat lower, and stems lowest of all. Theconcentration of phosphorus in the root was particularly high at the highlevels of nutrient phosphorus, but the total amount included absorbed aswell as adsorbed phosphorus, with the latter apparently contributing anappreciable amount. This is the characteristic distribution of phosphorus

436



BIDDULPH AND 'WOODBRIDGE: UPTAKE OF PHOSPHORUS BY BEAN 437

between the various parts of the bean plant. Other elements, of course,assume their own characteristic distribution. No two are strictly alike.

As the concentration of phosphorus in the nutrient solution was in-creased from the lowest level upwards, all plant parts showed increasedconcentrations of phosphorus but the relation was not linear. Initially, therate of increase was relatively low, particularly for root concentrations andto a lesser extent for stem and cordate leaf concentrations. Rapid gainsN-ere made only after the phosphorus concentration of the nutrient mediumexceeded the iron concentration. The point of equivalence (equimolar con-centrations) between iron and phosphorus in the nutrient medium is indi-cated by an arrow on the abscissa of figure 2 A.

The trifoliate leaves showed an initial rapid rise with the first increasein nutrient phosphorus, but the rate of rise was not as rapid as it was withplants receiving no supplementary iron. The accumulating power of theseactively metabolizing leaves is very striking. The young trifoliate leaves,together with the meristematic tissues of the stem tip, were able to acquireand maintain higher concentrations of phosphorus from nutrient solutionsof low phosphorus content (0.00005 M) than tissues of the root, stem, orcordate leaves. With further increases in nutrient phosphorus, tissue con-centrations began to level off and lesser gains were made at the higher nu-trient ranges.

The principal difference between the plants grown with and without ironin the nutrient medium, was in the phosphorus concentration of the respec-tive parts of the plants grown at the zero level of nutrient phosphorus.Those plants which made their growth in the presence of an ample supplyof iron had a distinctly higher phosphorus concentration than those receiv-ing no supplementary iron. Of course, all growth in this case (zero P in fig.2 A and B) was made by utilization of phosphorus which was present in theseed as no other source was available. The time allotted for growth (12days) was sufficient to insure the utmost dilution of phosphorus which waspossible under the conditions of growth. The failure of the plants suppliedwith iron to affect as great a dilution of seed phosphorus indicates a lessefficient or thorough utilization of that phosphorus. The most logical inter-pretation of this observation is that at least a portion of the iron taken intothe plant, or held on the root surfaces, was precipitated with a portion ofthe seed phosphorus rendering it unavailable for metabolic use. As a result,the tissues were relatively rich in total phosphorus at the time that growthceased from a lack of metabolically usable phosphorus.

At the higher levels of nutrient phosphorus, the plants grown with andwithout supplementary iron were essentially similar in the phosphorus con-centration of their tissues. Evidently, the amount of phosphorus held byiron is quite insignificant in comparison to the total phosphorus presentwlhen an ample supply is furnished to the roots.

PLANTS GROWN WITHOUT IRON SUPPLEMENTATION.-In those experimentswhere iron was intentionally omitted from the nutrient medium, and with



PLANT PHYSIOLOGY

particular reference to the plants grown at the zero level of nutrient phos-phorus, the concentration of phosphorus in the tissues of all plant parts isconsiderably lower than is the case where iron was present in the nutrientmedium. Furthermore, the initial gain in phosphorus concentration wasmore rapid for all plant parts as the phosphorus content of the nutrientmedium was increased. The initial flattened part of the curves as seen infigure 2 A is absent. The phosphorus concentration of the trifoliate leavesbuilt up particularly rapidly indicating a completely unobstructed move-ment of phosphorus to these rapidly metabolizing parts. With further in-creases in nutrient phosphorus beyond 0.00005 M, only small gains weremade. Beyond this point, the metabolic use of phosphorus by the trifoliateleaves, kept the incoming supply diluted to a value of approximately 6 mg.P/gm. of dry matter. Apparently this concentration within leaf tissue wasadequate for normal metabolic functions.

The phosphorus content of roots, cordate leaves, and stems increasedrather linearly with increases in nutrient phosphorus. At a nutrient valueof 0.0005 M, a nutrient concentration 10 times as high as was necessary formaintenance of adequate trifoliate leaf concentrations, the phosphorus con-centration of cordate leaves tended to level off. Stems and roots were stillslowly increasing in phosphorus content and continued to do so even up toa nutrient concentraton of 0.005 M phosphorus. Thus it is evident that cor-date leaves, stems, and roots progressively accumulate available phosphorusin excess of the values characteristic of these organs when the rapidly grow-ing leaves of the upper stem are adequately supplied. It is this additionalaccumulation of phosphorus by roots, stems and cordate leaves, which ap-pears in excess of the minimum requirements for the growth of the plant asa whole, that is responsible for disturbances in the metabolic use of certainother ions as these must enter and pass through tissues so rich in phos-phorus as to cause precipitation reactions and immobilization (5). Iron isso frequently immobilized in this manner that we have adopted the term,phosphorus-induced-chlorosis, to designate this type of disturbance.

EXPERIMENTS WITH TRACER PHOSPHORUS.-In the above experiments thetotal uptake of phosphorus which had occurred during the experimental pe-riod was measured. In the experiment to be reported, the absorption andtranslocation of an aliquot of marked phosphorus was measured after a pre-treatment period during which the different groups of plants were grown atphosphorus and iron levels indicated in figure 3. Plants were given half-strength solution at pH 6.0 during the first four days then full-strengthsolution for four days. This treatment was followed by a second four-dayperiod in full-strength solution in which the added phosphorus containedp32 at a known P32/P31 ratio. The plants were then analyzed for totalphosphorus and marked phosphorus, the marked phosphorus being thatphosphorus absorbed during the last four day period. One of the values ofthe tracer method lies in the fact that this fractioin can be quantitativelymeasured even in the presence of the fraction which was absorbed previ-

438



BIDDULPH AND WOODBRIDGE: UPTAKE OF PHOSPHORUS BY BEAN 439

50 ~~~~~R4~~~~~~~~~~~~~-0. .--0-_,_ . T.L.

z 10_JU. -,-..Roots .T.L.z

z ' \o \~z0

z

0

CL)

Ul)0

a. T.Leoves

0.1.2 .2 .2 1 1 20 20 20 PO 1 2 0 1 2 0 1 2 fe

PHOSPHORUS AND IRON CONCENTRATION OF NUTRIENT MEDIUM. MOLAR X 10-5

FIG. 3. The percentage increase in the phosphorus concentration of parts of beanplants as indicated by the accumulation of radioactive tracer phosphorus during a four-dav interval prior to harvest. The N-arious groups of plants wvere grown throughout (12days) in nutrient solutions containing the phosphorus and iron concentrations indicatedon the base lines. Resuilts are from two separate pans of six plants each. R, roots;TL, trifoliate leaves.

ously. A measurement of the marked phosphorus present in each organ, asw-ell as the total phosphorus, allowed the calculation of the percentage in-crease in phosphorus during the experimental period. The increase w-ascalculated as followvs:

incrase n P marked P x 10070ncraseln =total P -marked P'

The results are shown graphically in figure 3. The percentage increase inphosphorus reflects the amount of flow of currently acquired phosphorusinto a given tissue. A high value in a selected tissue results if there hasbeen an unimpaired flow of phosphorus to that particular area. A low valueresults when absorption is limited by the amount of phosphorus present, orby an impairment in the flow to the tissue.

By comparing plants grown at three different phosphorus levels, it isevident that the percentage gain in phosphorus is dependent first of all uponthe amount of phosphorus present in the nutrient medium. Those plantsgrowing at the highest nutrient levels acquire the most phosphorus. For



PLANT PHYSIOLOGY

plants growing at the same phosphorus levels the determining factor in theacquisition of phosphorus is the amount of iron present. The roots gainphosphorus (adsorbed plus absorbed) in direct relation to the iron concen-tration of the nutrient medium, while the trifoliate leaves gain phosphorusin inverse relation to the iron concentration of the nutrient medium. Thiseffect is greatest at the low nutrient phosphorus levels and becomes almostindiscernible at the high phosphorus levels. Iron, therefore, exerts a rela-tively greater influence in solutions of low phosphorus content. The effectis to cause a precipitation of phosphorus upon the roots which then ob-structs the flow of phosphorus to the aerial parts. It will be seen that wherethere occurred ten or more atoms of phosphorus to each one of iron, theeffect of iron upon the uptake of phosphorus into the trifoliate leaves wasrelatively insignificant. The explanation perhaps lies in the fact that thepreponderance of phosphorus over iron is responsible for a rapid precipita-tion of the iron and a subsequent suppression of its occurrence in a solubleform in the nutrient medium. Plants are then growing in an essentiallyiron-free solution wherein ample phosphorus is present.

When iron is present in superabundance over phosphorus, the occurrenceof ionic phosphorus in solution is suppressed by the formation of a ferric

20%/ 0-lo

|8 c

4

z~~~~~~~~~~~

0. p~~~~~~~~~6l%oxl ok z

o~~~~~~~~~~~

0gQ%. 4~~~c W4

w >_

z

IRON CONG. OF T. LEAVES

pI I~~~!xI05M.Fe O 1 2

P/Fe IN NUTRIENT MEDIA

FIG. 4. The percentage increase in phosphorus, as indicated by the accumulationof radioactive tracer phosphorus during the four-day interval prior to harvest, and thecorresponding iron concentrations at the time of harvest of parts of bean plants grownfor 12 days in nutrient solutions containing various ratios of phosphorus and iron.Results are those of the central curves in figure 3 together with iron values as deter-mined by John Rediske.

440



BID1)ULPH AND WOODBRIDGE: UPTAKE OF PHOSPHORUS BY BEAN 441

phosphate complex. The excess iron forms hydroxides which also contrib-ute toward its precipitation. Iron, then, influences the uptake of phos-phorus into the aerial parts of the plants first, by removing some phosphorusby precipitation, and second, by the formation of a precipitate upon theabsorbing surfaces which impairs the rapid entrance and passage of phos-phorus across the cortex of the root. Therefore, when precipitated phos-phorus and iron accumulates upon the roots, the passage of phosphorus tothe aerial parts is decreased.

The data from the 0.00001 2I phosphorus level of figure 3 are presentedin figure 4 together with the iron values for the roots and trifoliate leaves.This figure furnishes additional convincing proof that the interpretation ofresults on the basis of precipitation reactions is valid. The percentage gainin phosphorus by roots is greatest from solutions of high iron concentration.Likewise the amount of iron associated with the roots is also highest in so-lutions of high iron content. However, as the iron concentration and thepercentage gain in phosphorus by roots increase, the gain in phosphorus byleaves is correspondingly decreased.

The nature of the precipitate which was present on the root surfaceshas not been precisely determined, but the precipitate which formed withinthe nutrient solutions alone at pH values of 6 or lower consisted of approxi-mately stoichiometric proportions of iron and phosphorus. At pH valuesabove 6, some calcium was present in the precipitate and on a quantitativebasis calcium plus iron was equal to phosphorus.

SWANSON et al. (13) have shown that the following reaction takes placewhen ferric chloride is titrated with sodium hydroxide in the presence of anexcess of phosphate:

Fe (H20)++6 + 2 0H- + H2PO4- -> Fe (H20)3 (OH) 2H2P04 + 3 H20.The H2PO4- occupies only one of the coordination positions of the hexa-hydrated iron ions. It is assumed from the work of the above authors andfrom our own studies, that under the conditions of the experiments hereinreported, i.e., in nutrient solutions, the formation of the basic iron phos-phate is possible and may take place to the extent of full participation ofavailable phosphate ions up to the equivalence point. That phosphatewhich is present in superabundance over iron does not react and, except forsome slight adsorption, remains available to the plant.

On the basis of this and other investigations, we now expect to find thata number of nutritional disturbances (other than chlorosis) will be explain-able on the grounds of an immobilization of essential nutrients in precipi-tation and absorption reactions in the conductive tissues of the plant. It isknown that certain soils are constituted, in part, of elements whose defi-ciency symptoms are often manifest in the plants which grow in them. Theexplanation frequently given is that the elements in question are tied up inthe soil in a manner which renders them unavailable to the plants. Fromthe data presented herein, it is shown that a more detailed explanation in-



PLANT PHYSIOLOGY

volves precipitation reactions in and on the surface of the roots as well asin the conductive tissues, particularly near the distal ends, which renderssome elements unavailable for metabolic use. Those elements which forminsoluble precipitates with p4osphorus are to be particularly suspected asparticipating in such a mechanism.

Summary

Absorption and translocation of phosphorus by bean plants was unim-paired at pH 4.0. A minimum in phosphorus accumulation occurs at ornear pH 5.0 which reflects the slight solubility of basic ferric phosphate ator near this pH. It is conceivable that this minimum also reflects the mini-mal absorption encountered at the isoelectric points of certain cellular con-stituents active in phosphorus absorption. As the pH of the nutrient solu-tion was raised from 6.0 to 7.0, the concentration of phosphorus in the stemsand petioles increased, whereas the concentration in the leaf blades de-creased. This observation indicates a lodging of incoming phosphorus inthe stems and petioles at high pH values, creating tissues rich in phosphorusthrough which ions being transported to the leaf blade must pass.

As the phosphorus concentration of the nutrient medium is increasedfrom low to higher values, the tissue concentrations also increase, but atdifferent rates. The general effect is a rapid rise, followed by lesser in-creases until a leveling off occurs. The phosphorus concentrations of thenutrient solutions at which the different plant organs level off in phosphorusconcentrations are as follows: trifoliate leaves, 0.00005 M; cordate leaves,0.0005 M; stems, 0.005 M. Root concentrations begin leveling off at0.005 M.

A concentration of approximately 6 mg. of P/gm. of dry matter in tri-foliate leaves is attained from solutions at 0.00005 M P and this value issufficient for continued growth of leaves. The corresponding combined stemand petiole concentration is approximately 2 mg. P/gm. dry matter whenthe leaves are adequately supplied. The stems and petioles will, however,build up to 4 mg. P/gm. dry matter as more phosphorus is made available.Cordate leaves correspondingly rise from 3 to 7 mg. P/gm. dry mlatter. Itis this additional accumulation of phosphorus, beyond the concentrationwhich is adequate for leaf growth and stem extension, which causes dis-turbances in the metabolic use of other ions, particularly iron. The passageof such ions through tissues rich in phosphorus is interfered with and muchof the iron is precipitated along the conductive tissue. The principal effectof high phosphorus concentrations on the development of chlorosis is ex-plainable on this basis.

In a nutrient solution containing both phosphorus and iron in nutrientquantities, a precipitate will form which will reduce the amount of availableions of both elements. The precipitate of iron and phosphorus forms bothupon the container and upon the plant roots. The effect of the precipitationis twofold. First, it removes some of the materials from solution reducing

442



BIDDULPH AND WOODBRIDGE: UPTAKE OF PHOSPHORUS BY BEAN 443

the effective concentration. Second, the presence of the precipitate uponabsorbing surfaces furnishes a barrier to the rapid entrance of either speciesof ion, i.e., phosphate and iron. Under certain conditions, precipitation re-actions may also occur in the conductive tissues of the stemn and leaf and soconstitute a block in the transfer of certain ions froml- the conductive tissueto the leaf mesophyll and growing points.

The P32 herein employed was obtained fromn the Isotopes Division,Atomic Energy Commission, Oak Ridge, Tennessee.

DEPARTMENT OF BOTANYSTATE COLLECE OF WASHINGTON

PULLMAN, WASSHINGTON

AND

DIvISION- OF CHEMISTRYDEPARTMENT OF AGRICULTURE

SUMMERLAND, BRITISH COLUMBIA, CANADA

LITERATURE CITED1. AIYAR, S. P. Effects of phosphate deficiency on rice. Proc. Indian

Acad. Sci. 23B: 165-193. 1946. Chem. Abstracts 40: 7473. 1946.2. ARNONT D. I., FRATZKE, W. E., and JOHNSON, C. 'M. Hydrogen ion con-

centration in relation to absorption of inorganic nutrients by higherplants. Plant Physiol. 17: 515-524. 1942.

3. BIDDULPH, 0. Diurnal migration of injected radiophosphorus frombean leaves. Amer. Jour. Bot. 28: 348-352. 1941.

4. BIDDULPH, 0. Proceedings of the Auburn Conference on the Use ofRadioactive Isotopes in Agricultural Research. p. 90-102. Ala-bama Polytechnic Institute, Auburn. 1948.

5. BIDDULPH, 0. The translocation of minerals in plants. In: MineralNutrition of Plants, E. Truog, Ed. p. 261-275. University ofWisconsin Press, MIadison. 1951.

6. FRANCO, C. M. and LoomIIs, W. E. The absorption of phosphorus andiron from nutrient solutions. Plant Physiol. 22: 627-634. 1947.

7. KAMEN, M. D. Radioactive Tracers in Biology. Academic Press, NTewYork. 1947.

8. OLSEN, C. Iron absorption and chlorosis in green plants. Com-apt.Rend. Trav. Lab. Carlsburg Ser. Chim. 21: 15-52. 1935.

9. OLSEN, C. Experiments with different quantities of iron salts given tomaize in water culture. Compt. Rend. Trav. Lab. Carlsburg Ser.Chim. 21: 301-313. 1938.

10. REDISKE, J. H. The translocation of radio iron in the bean plant.Thesis, The State College of Washington. 1950.

11. SIDERIS, C. P., YOUNG, H. Y., and KRAUSS, B. H. Effects of iron on thegrowth and ash constituents of Ananas comnosus (L.) Merr. PlantPlhysiol. 18: 608-632. 1943.



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12. SNELL, F. D. and SNELL, C. J. Colorimetric Methods of Analysis. D.Van Nostrand Co., New York. 1936.

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PLANT PHYSIOLOGY physiology volume27 july, 1952 number3 the uptake of phosphorus by bean plants with par-ticular referenceto theeffects of iron 0. ... results and discussion