Bioavailability of soil inorganic P in the rhizosphere as affected by root-induced chemical changes: a review

In most soils, inorganic phosphorus occurs at fairly low concentrations in the soil solution whilst a large proportion of it is more or less strongly held by diverse soil minerals. Phosphate ions can indeed be adsorbed onto positively charged minerals such as Fe and Al oxides. Phosphate (P) ions can also form a range of minerals in combination with metals such as Ca, Fe and Al. These adsorption/desorption and precipitation/dissolution equilibria control the concentration of P in the soil solution and, thereby, both its chemical mobility and bioavailability. Apart from the concentration of P ions, the major factors that determine those equilibria as well as the speciation of soil P are (i) the pH, (ii) the concentrations of anions that compete with P ions for ligand exchange reactions and (iii) the concentrations of metals (Ca, Fe and Al) that can coprecipitate with P ions. The chemical conditions of the rhizosphere are known to considerably differ from those of the bulk soil, as a consequence of a range of processes that are induced either directly by the activity of plant roots or by the activity of rhizosphere microflora. The aim of this paper is to give an overview of those chemical processes that are directly induced by plant roots and which can affect the concentration of P in the soil solution and, ultimately, the bioavailability of soil inorganic P to plants. Amongst these, the uptake activity of plant roots should be taken into account in the first place. A second group of activities which is of major concern with respect to P bioavailability are those processes that can affect soil pH, such as proton/bicarbonate release (anion/cation balance) and gaseous (O₂/CO₂) exchanges. Thirdly, the release of root exudates such as organic ligands is another activity of the root that can alter the concentration of P in the soil solution. These various processes and their relative contributions to the changes in the bioavailability of soil inorganic P that can occur in the rhizosphere can considerably vary with (i) plant species, (ii) plant nutritional status and (iii) ambient soil conditions, as will be stressed in this paper. Their possible implications for the understanding and management of P nutrition of plants will be briefly addressed and discussed.     

氮素形态对专用小麦中后期根际土壤微生物和酶活性的影响

采用盆栽方法研究了酰胺态氮、铵态氮和硝态氮对强筋小麦(Triticum aestivum L.)"豫麦34"、中筋小麦"豫麦49"和弱筋小麦"豫麦50"生育中后期根际微生物和土壤酶活性的影响.结果表明,专用小麦根际真菌、细菌、放线菌数量和土壤脲酶、蛋白酶、硝酸还原酶活性以及根际pH值对氮素形态的反应不同."豫麦34"施用硝态氮,对根际土壤真菌、细菌(除成熟期外)和放线菌数量均具有明显的促进作用;"豫麦49"施用铵态氮,根际土壤细菌和放线菌数量最大,根际真菌数量在孕穗期和开花期以酰胺态氮处理最大,而成熟期以硝态氮处理最大;"豫麦50"施用硝态氮,对根际土壤真菌、细菌和放线菌数量均具有明显的促进作用.不同专用小麦品种均表现为在酰胺态氮处理下,根际土壤脲酶活性最高;在铵态氮处理下,根际土壤蛋白酶活性最高;在硝态氮处理下,根际土壤硝酸还原酶活性和pH值最高.     

氮肥运筹对免耕水稻根系生长、根际土壤特性及产量的影响

为探索氮肥运筹对免耕条件下水稻根系生长以及对根际土壤特性、产量的影响,以金优253为材料进行试验。结果表明：平衡施肥显著提高单株根系干重、根长、单株生物量、根半径、单株根表面积、根长密度及根系活力,实收单产高于重穗肥和重基肥处理,且与重基肥差异达95％的显著水平,主要是有效穗数、结实率的增加。平衡施肥显著提高0~10 cm土层的0~2 mm根际土壤有机质、碱解氮含量及脲酶、蔗糖酶活性。因此平衡施肥能明显促进免耕水稻根系生长和有效穗数的增加,对提高水稻产量具有促进作用。     

Non-destructive measurement of acid phosphatase activity in the rhizosphere using nitrocellulose membranes and image analysis

We developed a method using nitrocellulose membranes and image analysis to localise and quantify acid phosphatase activity in the rhizosphere of two plant species, one with cluster roots (Dryandra sessilis (Knight) Domin) and another with ectomycorrhizal roots (Pinus taeda L.). Membranes were placed in contact with roots and then treated with a solution of x, α-naphthyl phosphate and Fast Red TR. Acid phosphatase activity was visualised as a red imprint on the membrane. We quantified acid phosphatase activity by image analysis of scanned imprints. The method was used to estimate the spatial distribution of acid phosphatase activity within particular root classes (lateral roots, mycorrhizal roots, root clusters). Over 95% of the acid phosphatase activity of the root system of D. sessilis was associated with cluster roots, and between 20 and 32% of the root surface active. About 26 % of the acid phosphatase activity of the root system of P. taeda was associated with mycorrhizal roots and unsuberised white root tips and less than 10% of the root surface was active, irrespective of root type. This non-destructive method can be used for rapid, semi-quantitative assessment of acid phosphatase activity in the laboratory and in situ. This revised version was published online in August 2006 with corrections to the Cover Date.     

Non-destructive methods for demonstrating chemical changes in the rhizosphere II. Application of methods

Chemical changes in the rhizosphere of soil-grown plants are demonstrated by non-destructive techniques based on colour reactions. The following examples are given: FeIII reduction in the rhizosphere of a Hakea species, MnIV reduction in the rhizosphere of chikpea, complexation of Al in the rhizosphere of Norway spruce, and the activity of acid phosphatase in the rhizosphere of maize.     

Effect of nitrogen form and phosphorus source on the growth,nutrient uptake and rhizosphere soil property of Camellia sinensis L.

The effects of nitrogen form and phosphorus source on the growth, nutrient uptake and rhizosphere soil property of tea (Camellia sinensis L.) were investigated in a pot experiment. The experiment was performed with a compartmental cropping device, which enables the collection of rhizosphere soil at defined distances from the root of tea plant. Nitrogen was supplied as nitrate or ammonium in combination with soluble phosphorus as Ca(H₂PO₄)₂ or insoluble P as rock phosphate. The leaf dry matter production of tea was significantly greater in the treatments with NH₄ ⁺ than NO₃ ^-, whereas dry matter production of root and stem was not significantly affected. Addition of phosphorus as either source did not influence the dry matter production. The concentrations of K in root, Mg and Ca in both the shoot and root supplied with NO₃ ^- were significantly higher than in NH₄ ⁺ and influence of P sources was minor. On the contrary, Al and Mn concentrations were significantly larger in NH₄ ^--fed plants which could be attributed to remarkably increased availability of Al and Mn caused by acidification of the rhizosphere soil (the first 1-mm soil section from the root surface) with NH₄–N nutrition. The concentration of N in shoot was also significantly higher in NH₄- than in NO₃-fed plants, indicating higher use efficiency of NH₄–N. Whatever the phosphate source, rhizosphere pH declined in ammonium compared to in nitrate treatment. The pH decrease was much larger when no P or soluble P were applied and reached 0.85–1.30 units which extended to 3–5 mm away from the root surface. Exchangeable acidity, content of exchangeable Al and Mn were also considerably higher in the rhizosphere soils of NH₄ ⁺ fed tea plants. Significant amounts of P dissolved from rock phosphate accumulated in rhizosphere of NH₄ ⁺, not NO₃ ^-, suggesting that the dissolution of rock phosphate was induced by the proton excreted by tea root fed with ammonium. With soluble P addition, shoot and root P concentrations were greater in NH₄ ⁺ than in NO₃ ^- treatment and it appeared that this difference could not be sufficiently explained by the available P content in soil which was only slightly higher in NH₄ ⁺ treatment. With rock phosphate addition, the shoot and root P concentrations were hardly affected by nitrogen form, although the available P content was much higher and accumulated in the rhizosphere soil supplied with ammonium. The reason for this was discussed with regard to the inter-relationship of Al with P uptake. This revised version was published online in June 2006 with corrections to the Cover Date.     

Effect of enriched rhizosphere carbon dioxide on nitrate and ammonium uptake in hydroponically grown tomato plants

Previous reports have indicated positive effects of enriched rhizosphere dissolved inorganic carbon on the growth of salinity-stressed tomato (Lycopersicon esculentum L. Mill. cv. F144) plants. In the present work we tested whether a supply of CO₂ enriched air to the roots of hydroponically grown tomato plants had an effect on nitrogen uptake in these plants. Uptake was followed over periods of 6 to 12 hours and measured as the depletion of nitrogen from the nutrient solution aerated with either ambient or CO₂ enriched air. Enriched rhizosphere CO₂ treatments (5000 μmol mol^-1) increased NO₃ ^- uptake up to 30% at pH 5.8 in hydroponically grown tomato plants compared to control treatments aerated with ambient CO₂ (360 μmol mol^-1). Enriched rhizosphere CO₂ treatments had no effect on NH₃ ⁺ uptake. Acetazolamide, an inhibitor of apoplastic carbonic anhydrase, increased NO₃ ^- uptake in ambient rhizosphere CO₂ treatments, but had no effect on NO₃ ^- uptake in enriched rhizosphere CO₂ treatments. Ethoxyzolamide, an inhibitor of both cytoplasmic and extracellular carbonic anhydrase, decreased NO₃ ^- uptake in ambient rhizosphere CO₂ treatments. In contrast, a CO₂ enriched rhizosphere increased NO₃ ^- uptake with ethoxyzolamide, although not to the same extent as in plants without ethoxyzolamide. It is suggested that a supply of enriched CO₂ to the rhizosphere influenced NO₃ ^- uptake through the formation of increased amounts of HCO₃ ^- in the cytosol. This revised version was published online in June 2006 with corrections to the Cover Date.     

The effects of root-induced pH changes on the depletion of inorganic and organic phosphorus in the rhizosphere

A new method allowing control of rhizosphere pH and mineral nutrition was applied to study depletion of various organic and inorganic phosphorus fractions extractable sequentially with 0.5M KHCO₃ (pH 8.5), 0.1M NaOH and residual P extractable with 6M H₂SO₄ from the rhizosphere soil.Soil pH was affected about 2 mm from the root mat. Depletion zones of inorganic P (KHCO₃-P_i) extractable with 0.5M KHCO₃ extended up to about 4 mm but the depletion zones of all other P fractions were about 1 mm only. The root-induced decrease of soil pH from 6.7 to 5.5 increased the depletion of total P from all fractions by 20% and depletion of KHCO₃-P_i and residual P by 34% and 43%, respectively. Depletion of organic P (KHCO₃-P_o) extractable with 0.5M KHCO₃ was not affected by a change in rhizosphere pH. With constant or increased pH, depletion of inorganic P (NaOH-P_i) was 17% and organic P (NaOH-P_o) was 22% higher than with decreased pH. Only 54–60% of total P withdrawn from all fractions was from KHCO₃-P_i. Substantial amounts of KHCO₃-P_o and NaOH-P_o were mineralized and withdrawn from the rhizosphere within 1 mm from the root mat, as 11–15% of total P withdrawn originated from the organic P fractions. A remaining 11–16% was derived from NaOH-P_i, and 15–18% from residual P fractions likely to be rather immobile. Thus, 40–46% of the P withdrawn near the root mat of rape originated from non-mobile P fractions normally not included in 0.5M NaHCO₃ extraction used to obtain an index of plant-available soil P.     

Effect of nitrogen form and root-zone pH on growth and nitrogen uptake of tea (Camellia sinensis) plants

BACKGROUND AND AIMS: Tea (Camellia sinensis) is considered to be acid tolerant and prefers ammonium nutrition, but the interaction between root zone acidity and N form is not properly understood. The present study was performed to characterize their interaction with respect to growth and mineral nutrition. METHODS: Tea plants were hydroponically cultured with NH4+, NO3- and NH(4+) + NO3-, at pH 4.0, 5.0 and 6.0, which were maintained by pH stat systems. KEY RESULTS: Plants supplied with NO3- showed yellowish leaves resembling nitrogen deficiency and grew much slower than those receiving NH4+ or NH(4+) + NO3- irrespective of root-zone pH. Absorption of NH4+ was 2- to 3.4-fold faster than NO3- when supplied separately, and 6- to 16-fold faster when supplied simultaneously. Nitrate-grown plants had significantly reduced glutamine synthetase activity, and lower concentrations of total N, free amino acids and glucose in the roots, but higher concentrations of cations and carboxylates (mainly oxalate) than those grown with NH4+ or NH(4+) + NO3-. Biomass production was largest at pH 5.0 regardless of N form, and was drastically reduced by a combination of high root-zone pH and NO3-. Low root-zone pH reduced root growth only in NO(3-)-fed plants. Absorption of N followed a similar pattern as root-zone pH changed, showing highest uptake rates at pH 5.0. The concentrations of total N, free amino acids, sugars and the activity of GS were generally not influenced by pH, whereas the concentrations of cations and carboxylates were generally increased with increasing root-zone pH. CONCLUSIONS: Tea plants are well-adapted to NH(4+)-rich environments by exhibiting a high capacity for NH4+ assimilation in their roots, reflected in strongly increased key enzyme activities and improved carbohydrate status. The poor plant growth with NO3- was largely associated with inefficient absorption of this N source. Decreased growth caused by inappropriate external pH corresponded well with the declining absorption of nitrogen.     

Physiological dissection revealed that both uptake and assimilation are the major components regulating different growth responses of two tobacco cultivars to nitrogen nutrition

• K326 and HD represent major tobacco cultivars in China, which required large N fertiliser input but at different application rates. To understand primary components affecting tobacco N use physiology, we adopted these two varieties as valuable genetic material to assess their growth response to N nutrition.
• We established a hydroponic culture system to grow plants supplied with different N regimes. Plant biomass, N, ammonium, nitrate, arginine, GS and NR activity, N transfer and use efficiency as well as root uptake were examined.
• Our data revealed the preference of K326 and HD to utilise nitrate or ammonium nitrate but not ammonium alone, with 2 mm N supply probably sufficient and economical to achieve good biomass production at the vegetative stage. Moreover, both varieties were very sensitive to ammonium, perhaps due to lack of or abnormal signalling related to nitrate and/or arginine rather than impairment of N acquisition and initial assimilation; this was supported by measurements of the plant content of N, ammonium and activities of GS and NR. Notably, short‐term ¹⁵N root influx studies identified differential uptake kinetics of K326 and HD, with distinct affinities and transport rates for ammonium and nitrate.
• The data suggest that the growth adaptation of K326 or HD to higher or lower N may be ascribed to different competences for effective N uptake/translocation and assimilation. Thus, our work provides valuable information to prompt deeper investigation of the molecular basis controlling plant N use efficiency.

     

Influence of inorganic nitrogen and pH on the elongation of maize seminal roots

BACKGROUND AND AIMS: Root absorption and assimilation of inorganic nitrogen usually alters rhizosphere pH, but the immediate influence of such pH changes on root elongation as well as that of exogenous inorganic nitrogen itself has been uncertain. METHODS: A differential extensiometer that monitored on a real-time, continuous basis root elongation in an intact 3-d-old maize plant was developed. Treatments included root media at pH 6.5 or 5.6 that lacked nitrogen and ones at pH 6.5 that contained 100 mmol m(-3) NH(4)(+) or NO(3)(-). KEY RESULTS: Acidifying the root medium from pH 6.5 to 5.6 nearly doubled the elasticity of the seminal root, but slightly decreased its elongation. Plasticity of the root apex was not detectable in all treatments. The presence of ammonium or nitrate in the medium stimulated elongation by 29 % or 14 %, respectively. Addition of an osmoticum to the medium had no effect on root elongation in the absence of inorganic nitrogen, but diminished the stimulation of elongation in the presence of ammonium and nitrate. This indicates that these ions or their by-products serve partially as osmolytes. CONCLUSIONS: In nutrient solution, root elongation of a maize seedling--even one with ample nitrogen reserves--depended most strongly on exogenous inorganic nitrogen, and less so, if at all, on either the pH of the bulk nutrient solution or the mechanical properties of cell walls.     

Origins of root-mediated pH changes in the rhizosphere and their responses to environmental constraints: A review

The aim of the present review is to define the various origins of root-mediated changes of pH in the rhizosphere, i.e., the volume of soil around roots that is influenced by root activities. Root-mediated pH changes are of major relevance in an ecological perspective as soil pH is a critical parameter that influences the bioavailability of many nutrients and toxic elements and the physiology of the roots and rhizosphere microorganisms. A major process that contributes root-induced pH changes in the rhizosphere is the release of charges carried by H⁺ or OH^– to compensate for an unbalanced cation–anion uptake at the soil–root interface. In addition to the ions taken up by the plant, all the ions crossing the plasma membrane of root cells (e.g., organic anions exuded by plant roots) should be taken into account, since they all need to be balanced by an exchange of charges, i.e., by a release of either H⁺ or OH^–. Although poorly documented, root exudation and respiration can contribute some proportion of rhizosphere pH decrease as a result of a build-up of the CO₂ concentration. This will form carbonic acid in the rhizosphere that may dissociate in neutral to alkaline soils, and result in some pH decrease. Ultimately, plant roots and associated microorganisms can also alter rhizosphere pH via redox-coupled reactions. These various processes involved in root-mediated pH changes in the rhizosphere also depend on environmental constraints, especially nutritional constraints to which plants can respond. This is briefly addressed, with a special emphasis on the response of plant roots to deficiencies of P and Fe and to Al toxicity. Finally, soil pH itself and pH buffering capacity also have a dramatic influence on root-mediated pH changes.     

New approaches to studying chemical and physical changes in the rhizosphere: an overview

The past decade has seen the rapid development of new techniques that have revealed substantial changes in soil physical and chemical properties in the rhizosphere compared to the bulk soil. This brief overview focuses on some examples of recently developed, innovative techniques now available and indicates the technical developments required for the future. The development of non-invasive imaging allied with computed tomography has begun to allow the study of root systems in situ and the measurement of localized uptake of water. Further development is still required to disaggregate the simultaneous changes in bulk density and water content that may occur as roots occupy new soil volumes, but resolution of 0.1 mm is now feasible in scanning times of less than 1 h thereby allowing dynamic processes to be measured. Changes in surface tension and composition of solutions close to roots, and of pH, can now be measured with a variety of techniques. Temporal and spatial changes of pH can be measured with micro-electrodes and dye indicator/agar gel techniques have allowed quantitative estimates of H⁺ fluxes albeit in artificial systems. Novel micro-sampling techniques are under development to quantify rhizosphere changes. So far these techniques have rarely been applied in soils but innovative sampling and analytical techniques should soon allow such studies. This revised version was published online in June 2006 with corrections to the Cover Date.     

Ion currents and the nitrogen status of roots of Hordeum vulgare and non-nodulated Trifolium repens

Abstract. Profiles of self-generated ion currents associated with the growing primary root tips of intact Hordeum vulgare L. and Trifolium repens L. (nonnodulated) seedlings were measured using a highly sensitive vibrating electrode in media containing NH⁺₄ or NO^-₃, and compared to control roots growing in nitrogen free media. Under these three nutrient regimes, positive current entered the root at regions corresponding to the meristematic tissues and main elongation zones of root tips and left from the mature root tissues. Mapping the surface of the roots with a pH-sensitive microelectrode revealed regions of external alkalinity where positive electrical current entered the root, and external acidity where positive current exited. The correlation between pH-profile and the pattern of ion current generation in these experiments suggests that H⁺ ions were responsible for carrying the bulk of the root-generated current. Assimilation of NHJ results in net H⁺ extrusion while assimilation of NO^-₃, results in net OH^-₃ efflux. Growth on NH⁺₄, as compared to growth on NO^-₃, stimulated the magnitude of the electrical current but did not affect significantly the growth rate of the roots. However, despite the differing stresses on internal pH regulation that arise due to growth on the two exogenous forms of combined nitrogen, the current profiles were qualitatively similar under the different conditions that were examined. The role of the circulating proton current is not yet known; however, the constancy of the current profile under different nutrient regimes sustains the hypothesis that the current may have a role in the regulation of root polarity.     

Root Development and Absorption of Ammonium and Nitrate from the Rhizosphere

Plant roots operate in an environment that is extremely heterogeneous, both spatially and temporally. Nonetheless, under conditions of limited diffusion and against intense competition from soil microorganisms, plant roots locate and acquire vital nitrogen resources. Several factors influence the mechanisms by which roots respond to ammonium and nitrate. Nitrogen that is required for cell division and expansion derives primarily from the apex itself absorbing rhizosphere ammonium and nitrate. Root density and extension are greater in nutrient solutions containing ammonium than in those containing nitrate as the sole nitrogen source. Root nitrogen acquisition alters rhizosphere pH and redox potential, which in turn regulate root cell proliferation and mechanical properties. The net result is that roots proliferate in soil zones rich in nitrogen. Moreover, plants develop thinner and longer roots when ammonium is the primary nitrogen source, an appropriate strategy for a relatively immobile nitrogen form. Present address of Alison R. Taylor: The Marine Biological Association, The Laboratory, Citadel Hill, Plymouth PL1 2PB, UK.     

根分泌物对根际矿物营养及根际微生物的效应

综述了根系分泌物对植物生长的生理生态学效应,并就根系分泌物的定义、产生机制、组成成分和影响因素等方面进行了讨论。指出根系分泌物在缓解低矿物营养胁迫对植株造成的伤害及决定根际微生物的种群密度和数量方面起着重要的作用;根系分泌物的产生机制多样,组成成分复杂,影响因素繁多。对根分泌物的深入研究有助于进一步了解植物体与土壤间进行的生理生化过程及其调控机制。     

Phosphorus efficiency and the forms of soil phosphorus utilized by upland rice cultivars

Experimental measurements of phosphorus (P) uptake and the forms of soil P depleted from an Ultisol by 6 upland rice cultivars are reported. In both P-fertilized and-unfertilized soil, the majority of P taken up was solubilized from a 0.1 M NaOH-soluble pool by root-induced changes. The soil pH within 4 mm of the roots was lowered by up to 0.5 units (from 4.6), but this by itself could not account for the P solubilized, and nor could increased phosphatase activity near the roots. The possible role of root-released low molecular weight organic acid anions in P solubilization is discussed. No significant differences in the extent of solubilization by a given root mass could be detected between cultivars. In P-unfertilized soil, but not in P-fertilized soil, there were significant differences between cultivars in internal P efficiency as measured by shoot dry weight per unit total plant P. In unfertilized soil, root growth and P uptake were strongly correlated with the P content of the seeds from which the plants were grown.     

Influence of zoysiagrass rhizosphere fungal isolates on growth and yield of soybean plants

Among 21 rhizosphere fungi tested, eight sterile fungi and oneTrichoderma isolate (GT2-1) from zoysiagrass rhizosphere promoted the overall growth of soybean varieties when grown in the greenhouse. Out of nine effective isolates, GS7-4, GS8-2, GS8-3, GU23-3 (all sterile fungi) and GT2-1 (Trichoderma sp.) promoted plant growth and increased yield of Toyosuzu (variety 1) significantly, while GS8-3, GS10-1, GS10-2 (sterile fungi), and GT2-1 significantly caused plant growth promotion and yield increase of Kitamusume (variety 2). Among these efficient isolates, GS8-3 and GS10-2 induced considerable and consistent increases in length, biomass and yield of plants of varieties 1 and 2, respectively. In the field, however, only GS8-3 and GU23-3 among seven selected isolates, induced consistent and significant increases in plant growth and yield of varieties 1 and 2, while the ability of other isolates decreased. The plant growth promotion by these isolates in the field followed a similar trend to that in the greenhouse, but the effect was less marked. Some isolates which were effective in the greenhouse were less effective in the field. The degree of growth promotion by different isolates depended on the variety of soybean. The nutrient condition of soils used in experiments also seemed to play a vital role, since notable growth promotion by these isolates was observed in nutrient-depleted soil.The author is grateful to Ministry of Education, Science and Culture, Japan, for financial assistance.     

Comparison of the chemical changes in the rhizosphere of the nickel hyperaccumulator Alyssum murale with the non-accumulator Raphanus sativus

Changes in pH and redox potential were studied in the rhizosphere soil of a nickel hyperaccumulator plant (Alyssum murale) and of a crop plant, radish (Raphanus sativus). Differences in rhizosphere pH and reducing activity were found between the lateral and the main roots of both species, but the pH changes in the rhizosphere were similar in both species. Changes in pH were associated with the relative uptakes of cations and anions; whether the concentrations of heavy metals in the growth medium did not have any effect on the rhizosphere pH. The source of nitrogen (ammonium or nitrate) was the major factor determining the pH of the rhizosphere of both species. The redox potential of the rhizosphere was influenced by both the N-source and the concentrations of heavy metals. When heavy metals were not present in the growth medium, and nitrate was the N-source, the reducing capacity of A. murale roots was enhanced. However, the reducing activity of A. murale was always smaller than that of radish. Therefore, the mechanism of metal solubilization by the hyperaccumulator plant does not involve either the reduction of pH in the rhizosphere or the release of reductants from roots. The acidification and reducing activity of the roots of A. murale was always smaller than that of R. sativus.