Phytic acid. A natural antioxidant

The catalysis by iron of radical formation and subsequent oxidative damage has been well documented. Although many iron-chelating agents potentiate reactive oxygen formation and lipid peroxidation, phytic acid (abundant in edible legumes, cereals, and seeds) forms an iron chelate which greatly accelerates Fe2+-mediated oxygen reduction yet blocks iron-driven hydroxyl radical generation and suppresses lipid peroxidation. Furthermore, high concentrations of phytic acid prevent browning and putrefaction of various fruits and vegetables by inhibiting polyphenol oxidase. These observations indicate an important antioxidant function for phytate in seeds during dormancy and suggest that phytate may be a substitute for presently employed preservatives, many of which pose potential health hazards.     

Usefulness of the dietary phytic acid/ zinc molar ratio as an index of zinc bioavailability to rats and humans

Evidence suggests that rats can tolerate a dietary phytate/Zn molar ratio greater than 15 if the dietary Zn concentration is high. High dietary Ca exacerbates the effect of phytic acid on Zn utilization by rats. In a short term (15 d) balance trial with adult men, we observed slightly greater Zn balance when whole compared to dephytinized wheat bran was consumed (molar ratios 12 and 1.2, respectively). There was, however, greater fecal excretion of Zn during the first 5 d whole bran was consumed. In a second study, Na phytate was the major source of phytic acid and Zn balance was less when the phytate/Zn molar ratio was greater than 16 compared to 4. The difference was not significant, however, and there was evidence of physiological adjustments to maintain homeostasis when the high ratio diet was consumed. Mean Zn intake averaged 17 mg (0.26 mmole) and 11 mg (0.17 mmole) daily for the bran and Na phytate studies, respectively. The level of Zn intake may influence the response of humans to varying phytate/Zn ratios. Comparison of isotope retention studies and the balance data is discussed. Some information on the relationship of dietary Ca to the phytate/Zn effect in human diets is gathered from current literature. The phytate/Zn molar ratio is a useful index of Zn bioavailability.     

The role of phytic acid in legumes: antinutrient or beneficial function?

This review describes the present state of knowledge about phytic acid (phytate), which is often present in legume seeds. The antinutritional effects of phytic acid primarily relate to the strong chelating associated with its six reactive phosphate groups. Its ability to complex with proteins and particularly with minerals has been a subject of investigation from chemical and nutritional viewpoints. The hydrolysis of phytate into inositol and phosphates or phosphoric acid occurs as a result of phytase or nonenzymatic cleavage. Enzymes capable of hydrolysing phytates are widely distributed in micro-organisms, plants and animals. Phytases act in a stepwise manner to catalyse the hydrolysis of phytic acid. To reduce or eliminate the chelating ability of phytate, dephosphorylation of hexa- and penta-phosphate forms is essential since a high degree of phosphorylation is necessary to bind minerals. There are several methods of decreasing the inhibitory effect of phytic acid on mineral absorption (cooking, germination, fermentation, soaking, autolysis). Nevertheless, inositol hexaphosphate is receiving increased attention owing to its role in cancer prevention and/or therapy and its hypocholesterolaemic effect.     

Biodiversity of human faecal bacteria isolated from phytic acid enriched chemostat fermenters

BACKGROUND: Myo-inositol hexaphosphate (IP6) or phytic acid is found mostly in cereals and legumes and is thought to possess anti-carcinogenic properties. AIM: To isolate and identify faecal bacteria capable of phytic acid metabolism and to assess the effectiveness of prebiotics (dietary oligosaccharides, metabolised by selective colonic bacteria) in preserving the integrity of phytic acid. METHODS: Faecal samples from three volunteers were used in continuous culture experiments under varying conditions of pH, substrate concentration and dilution rates, seventy three different isolates cultured at steady state were then screened for phytic acid metabolism and identified through partial sequencing of their 16S rRNA genes (16S ribosomal ribonucleic acid). Utilisation of phytic acid was also assessed in a continuous culture system enriched with prebiotic fructooligosaccharides (FOS). RESULTS: Bacteroides spp., Clostridium spp. and facultatively anaerobic bacteria generally appeared to maintain viable counts in the presence of phytic acid. Bifidobacterium spp. and Lactobacillus spp. appeared less able to maintain viable counts in the presence of phytic acid. These results were confirmed by an increase in viable counts of Bacteroides spp., Clostridium spp. and a decrease in viable counts of Bifidobacterium spp. and Lactobacillus spp. once phytic acid was introduced to a FOS enriched continuous culture. CONCLUSIONS: The phytate metabolising biodiversity from the human large intestine does not appear to encompass major bacterial genera associated with beneficial or benign health effects (e.g. Lactobacillus spp. and Bifidobacterium spp).     

低植酸作物突变体研究进展

植酸是玉米(Zea mays)、小麦(Triticum aestivum)、大麦(Hordeum vulgare)、水稻(Oryza sativa)和大豆(Glvcine max)等籽粒中广泛存在的一种有机酸(6-肌醇磷酸), 其与K+、Ca2+、Mg2+和Fe3+等金属离子形成的植酸盐是微量营养元素的重要贮存形式。植酸及植酸盐不能被人和非反刍动物所吸收利用; 植酸摄入体内后还会和其他来源的微量营养元素结合形成植酸盐, 造成这些营养元素的生物有效性下降, 从而造成微量元素缺乏症。此外, 大量的植酸及植酸盐随粪便排出, 造成严重的环境污染, 尤其是水体富营养化。由于土壤中缺乏分解微生物, 即使畜禽粪便作有机肥还田仍不能被作物吸收利用。近年来, 利用理化诱变与转基因技术已成功地获得了玉米、大麦、水稻和大豆等作物的低植酸突变体。本文对植酸的生物合成过程、低植酸突变体的诱发与研究、低植酸突变体的遗传特征与可能机理及营养评价进行了综述, 并对低植酸作物的应用前景进行了简要分析。     

Enumerating the phytic acid content in maize germplasm and formulation of reference set to enhance the breeding for low phytic acid

Phytic acid (Myoinositol 1, 2, 3, 4, 5, 6 hexakisphosphate) is a ubiquitous compound present in plants. It is an important constituent in seed reducing the bioavailability of phosphorous and mineral nutrients when fed to monogastric animals like swine, poultry, fish etc. Hence, identification of maize germplasm with reduced phytic acid content is imperative to formulate the breeding programs to evolve low phytate lines. Towards this, three hundred and thirty-eight maize germplasm accessions available at Department of Millets, TNAU, were raised and screened for phytic acid content which varied from 2.77 to 16.70 mg/g of seed. Based on the variability present, a reference set with fifty-eight genotypes for phytic acid was formulated. The reference set was formed with random genotypes selected from the base population to follow a normal distribution (skewness; 0.17, kurtosis; 0.61 and K–S test for normality Dn = 0.70) for phytic acid. The non-significant difference between the means of the base and the reference ensured the entire representation of the base in the formulated reference for phytic acid. Among all the lines in the reference set, the lowest phytic acid content were observed in the lines UMI-113 (2.77 mg/g) followed by UMI-300-1 (3.17 mg/g), UMI-467 (5.50 mg/g) and UMI-158 (6.58 mg/g) could be used as donors for low phytic acid in breeding programs. The principal component analysis for studying the extent of variability in the reference, revealed six major principal components that exhibited 80.40% of variation with flowering traits, ear height and phytic acid as a major contributor for variability. The characters namely plant stand, germination percentage, kernel yield, ear length, ear diameter and number of kernels per row were found to be positively correlated with the phytic acid and this emphasizes the negative pleiotropic effects of low phytic acid lines in germination and seed set. Thus this formulated reference set enables the breeders to handle minimum population for further grouping the genotypes to analyse their heterotic potential combined with low phytic acid.     

Phytic acid-metal ion interactions. II. The effect of pH on Ca(II) binding

The interaction of phytic acid with Ca(II) has been studied by potentiometric titration and by measurement of free Ca(II) concentrations using an ion Selective electrode. With increasing Ca(II) concentration, the titration curve of phytic acid is displaced to regions of lower pH. In the binding of calcium ions to phytic acid, there is no evidence that significant binding occurs below approximately pH 5. Above this pH, the extent of binding is dependent upon both pH and the calcium to phytic acid ratios. Maximum binding obtains at a Ca(II):phytate ratio of 6 with 4.8 mol of Ca(II) bound per mol of phytate above pH ca. 8. Binding constants are apparently very large since binding isotherms at any Ca(II):phytate ratio are a linear function of the total calcium ion concentration. In all cases, binding occurs only when one or more phosphate groups have been converted to the oxo dianion form. The apparent pK' values (curve-fit parameters) that describe the potentiometric titration data are in good agreement with the constants evaluated from the binding of Ca(II) to phytate as a function of pH. Using CPK space-filling models, structures containing six metal ions in coordinate linkage to pairs of oxo dianions have been constructed and discussed within the framework of the axial conformation of phytic acid and the order of proton removal with an increase in pH based upon NMR studies.     

Phytic acid in green leaves

Phytic acid or phytate, the free‐acid form of myo‐inositolhexakiphosphate, is abundant in many seeds and fruits, where it represents the major storage form of phosphorus. Although also known from other plant tissues, available reports on the occurrence of phytic acid, e.g. in leaves, have never been compiled, nor have they been critically reviewed. We found 45 published studies with information on phytic acid content in leaves. Phytic acid was almost always detected when studies specifically tried to detect it, and accounted for up to 98% of total P. However, we argue that such extreme values, which rival findings from storage organs, are dubious and probably result from measurement errors. Excluding these high values from further quantitative analysis, foliar phytic acid‐P averaged 2.3 mg·g^?1, and represented, on average, 7.6% of total P. Remarkably, the ratio of phytic acid‐P to total P did not increase with total P, we even detected a negative correlation of the two variables within one species, Manihot esculenta. This enigmatic finding warrants further attention.     

Independent and mutual interactions of copper(II) and zinc(II) ions with phytic acid

The interactions of phytic acid with Cu(II) and Zn(II) ions were examined as functions of metal ion concentrations and pH. Cu(II) ion-selective potentiometric and electron spin resonance (ESR) experiments provide strong evidence for the binding of Cu(II) ions to the phytic acid molecule at low pH (2.4–3.4) values. The relative stabilities of the copper and zinc phytates at low pH values were found to be very similar. For systems with metal ion:phytic acid molar ratios of 1:1–4:1 and 5:1–6:1 and pH values in the 3.4–5.9 and 3.4–5.0 ranges, respectively, Zn(II) ions were found to form complexes with phytic acid that were more stable than those of Cu(II) ions with phytic acid. The phytic acid molecule, however, was found to accommodate Cu(II) ions more readily than Zn(II) ions. For example, in systems containing equal amounts of Cu(II) and Zn(II) ions, 2 Zn(II) ions and 2, 3, 4, or 4.5 Cu(II) ions were found per phytic acid molecule depending upon metal ion:phytic acid molar ratios in the systems and pH. Total metal ion:phytic acid molar ratios and pH affected resultant metal ion solubilities and were factors influencing the effects of Zn(II) and Cu(II) ions on the binding of each other by phytic acid. Zn(II) and Cu(II) ions were observed to potentiate the binding of each other by phytic acid in some systems and compete with each other for phytate binding sites in others.     

Heats of precipitation of calcium phytate

The heat of precipitation has been determined for the reaction, in aqueous solution, of calcium ion with the phytate anion. The reaction is endothermic. From the value of the equilibrium constant associated with the reaction, the entropy change has been calculated. The large, positive entropy change is consistent with the view that when calcium ion reacts with the phytate anion there is a decrease in the hydrophilic character of the phytate anion. The quantitative aspects of the interaction of phytic acid with calcium provides a rational basis for the understanding of the mechanism of metal deprivation by phytic acid.     

Dietary Fiber Intake Increases the Risk of Zinc Deficiency in Healthy and Diabetic Women

Phytic acid is a major determinant of zinc bioavailability. Little is known about phytic acid intakes or indices of zinc bioavailability in type 2 diabetes mellitus (DM), a condition that predisposes to zinc deficiency. The aim of this cross-sectional study was to measure and explore the relationships among phytic acid intake, zinc bioavailability, and molecular markers of zinc homeostasis in 20 women with DM compared to 20 healthy women. The phytate/zinc, (calcium)(phytate)/zinc, and (calcium + magnesium)(phytate)/zinc molar ratios were used to indicate zinc bioavailability. Plasma zinc concentrations and zinc transporter (ZnT1, ZnT8, and Zip1) gene expression in mononuclear cells were measured. Participants with DM consumed 1,194?±?824?mg/day (mean?±?SD) phytic acid, an amount similar to the intake of healthy women (1,316?±?708?mg/day). Bread products and breakfast cereals contributed more than 40?% of the phytic acid intake in each group. A positive relationship was observed in all participants between phytic acid and dietary fiber (r?=?0.6, P?<?0.001) and between dietary fiber and the (calcium)(phytate)/zinc ratio (r?=?0.5, P?<?0.001). Compared to the healthy group, the messenger RNA ratio of ZnT1 (zinc export) to Zip1 (zinc import) was lower in participants with DM, which may indicate perturbed zinc homeostasis in the disorder. The plasma zinc concentration was not predicted by age, body mass index, health status, zinc bioavailability, or zinc transporter expression. Healthy and diabetic women consume phytic acid in amounts that are likely to decrease the bioavailability of dietary zinc. Recommendations to consume greater amounts of dietary fiber, much of which is associated with phytate, increase the risk of zinc deficiency.     

The heat of complex formation of copper(II) with phytic acid

The heats of complex formation of Cu(II) with phytic acid to form soluble complexes in the absence of precipitation at acid pH have been measured. The reaction was examined over a wide range of mol ratios of Cu(II):phytate. In all cases the heats of reaction were endothermic. Measurements of the uncombined copper by use of a copper electrode allowed calculation of the combined copper and hence the enthalpies. These latter values varied to some extent, depending upon the Cu(II):phytate ratio and the pH region where the reaction was examined. Factors which could contribute to the variation in the enthalpy terms include changes in the heats of ionization and possible structural changes with Cu(II) bound.     

Molecular marker development and linkage analysis in three low phytic acid barley (<Emphasis Type="Italic">Hordeum vulgare</Emphasis>) mutant lines

Phytate is the primary form of phosphorus found in mature cereal grain. This form of phosphorus is not available to monogastric animals due to a lack of the enzyme phytase in their digestive tract. Several barley low phytic acid (lpa) mutants have been identified that contain substantial decreases in seed phytate accompanied by concomitant increases in inorganic phosphorus. Seed homozygous for low phytic acid 1-1 (lpa1-1) or low phytic acid 2-1 (lpa2-1) has a 50% and 70% decrease in seed phytate respectively. These mutations were previously mapped to chromosomes 2HL and 7HL respectively. The RFLP marker ABC153 located in the same region of 2H was converted to a sequence-characterized-amplified-region (SCAR) marker. Segregation analysis of the CDC McGwire × Lp422 doubled haploid population confirmed linkage between the SCAR marker and the lpa1-1 locus with 15% recombination. A third low phytic acid mutant, M635, has a 75% decrease in phytate. This mutation was located to chromosome 1HL by linkage with an inter-simple sequence repeat (ISSR) based marker (LP75) identified through bulked-segregant analysis, and has been designated lpa3-1. Based on analysis of recombination between marker LP75 and low phytic acid in an additional mutant line M955 (95% phytate decrease), lpa3-1 and the mutation in M955 are in the same region on chromosome 1HL, and may be allelic.     

Inhibition of PCR amplification by phytic acid, and treatment of bovine fecal specimens with phytase to reduce inhibition

Development of effective polymerase chain reaction (PCR)-based diagnostic tests using ruminant fecal specimens has been thwarted by excessive inhibition. A PCR system based on amplification of 1000 copies of bacteriophage lambda-DNA was used as a model to evaluate inhibition levels in bovine feces. Dilution experiments using a bovine fecal specimen suggested that as little as 40 microg of feces (in a 100-microl PCR) affected the efficiency of amplification. It was discovered that phytic acid (the hexaphosphoric ester of inositol) is a powerful inhibitor of PCR. Above 0.3 mM phytate, the PCR is completely inhibited. In a very narrow range around 0.2 mM target-specific amplification proceeds efficiently. At concentrations between 10 and 100 microM, phytate nonspecific amplification (e.g., primer-dimer formation) is dominant. Below 10 microM, phytate target-specific amplification proceeds efficiently. A simple processing procedure using 50 units/ml of Aspergillus niger 3-phytase [E.C. 3.1.3.8] was developed that reduced PCR inhibition levels in bovine fecal specimens by approximately 500-fold.     

低植酸作物突变体研究进展

植酸是玉米(Zea mays)、小麦(Triticum aestivum)、大麦(Hordeum vulgare)、水稻(Oryza sativa)和大豆(Glvcine max)等籽粒中广泛存在的一种有机酸(6-肌醇磷酸),其与K 、Ca2 、Mg2 和Fe3 等金属离子形成的植酸盐是微量营养元素的重要贮存形式.植酸及植酸盐不能被人和非反刍动物所吸收利用;植酸摄入体内后还会和其他来源的微量营养元素结合形成植酸盐,造成这些营养元素的生物有效性下降,从而造成微量元素缺乏症.此外,大量的植酸及植酸盐随粪便排出,造成严重的环境污染,尤其是水体富营养化.由于土壤中缺乏分解微生物,即使畜禽粪便作有机肥还田仍不能被作物吸收利用.近年来,利用理化诱变与转基因技术已成功地获得了玉米、大麦、水稻和大豆等作物的低植酸突变体.本文对植酸的生物合成过程、低植酸突变体的诱发与研究、低植酸突变体的遗传特征与可能机理及营养评价进行了综述,并对低植酸作物的应用前景进行了简要分析.     

植物种子植酸研究进展

磷是植物生长发育所必需的大量营养元素。在种子发育过程中,植酸是磷的贮存库,对维持植物体内磷平衡有重要的作用。在种子萌发过程中,植酸酶分解植酸盐,释放磷、矿质营养和肌醇供幼苗生长。本文综述了近年来植物(作物)种子中植酸的生物合成途径、种子植酸含量的遗传、低植酸作物的育种等研究进展。首先,植酸生物合成途径中最初的反应底物为葡萄糖-6-磷酸,形成肌醇后,以肌醇为底物合成植酸共有两条路径:依赖脂类与不依赖脂类,目前,已分离鉴定若干植酸合成所需的关键酶及其编码基因,包括肌醇-3-磷酸合成酶、肌醇激酶、肌醇多磷酸盐激酶,以及参与植酸运输的ATP结合盒转运子。其次,利用作图群体及关联分析群体,分别在水稻(Oryza sativa L.)、白菜(Brassica rapa L.)、菜豆(Phaseolus vulgaris L.)等植物中鉴定出多个与种子植酸磷含量相关的遗传位点。第三,筛选获得有价值的低植酸突变体是培育低植酸作物的主要途径。当把低植酸作为育种目标时,可能会忽略种子植酸含量的降低给植物带来的不利影响,如何消除低植酸造成的不利影响,成为科学家们亟需解决的问题。     

Phytic acid: divalent cation interactions: III. A calorimetric and titrimetric study of the pH dependence of copper(II) binding

The interaction of the cupric ion with phytic acid as a function of pH has been studied by potentiometric and thermal titration and by the determination of ligand binding. As has been found for the reaction of zinc and calcium cations with phytate, the presence of the Cu(II) ion results in a displacement of the titration curves to more acid values. Evaluation of the parameters that describe such changes in ionization behavior by curve-fit analysis showed that as the Cu(II):phytate mol ratio was increased from one to eight, the pK' values of the ionizable group sets of phytic acid (ranging from 1.59 to 9.79) were consolidated into just two sets with curve-fit (CP) values ca. 1.5 and 3.7. Marked pH hysteresis effects are seen in such systems because of the pronounced acid strength of the Cu(II):aqua ion and the Cu(II) ligand aqua ion complex. The combined heat of binding and precipitation (plus solvation changes, etc.) of Cu(II) to phytate is endothermic (21.8–22.2 kcal mol⁻¹). This is similar in magnitude to that reported for the binding of either Zn(II) or Ca(II) to phytate. In the titration of Cu(NO₃)₂ with KOH, presumably to form Cu(OH)₂, ΔH° was exothermic (−12.5 kcal mol⁻¹). From measurements of free Cu(II) cation concentration in the presence of phytate the binding reaction was found to be stoichiometric with 6 mols Cu(II) bound at pH 6. Binding occurs within the pH range 2–6. An apparent necessary requirement for binding is the availability of the oxo dianion structure formed from the second dissociation step of a phosphoryl group. Curve-fit analysis of the binding data as a function of pH showed that a group or group set with CP value ca. 4 governs the binding reaction(s) at all mol ratios of Cu(II) to phytate examined. It is suggested that the binding of cupric ions to phytate may occur to the equatorial rather than the axial configuration as suggested for Ca(II) binding. A space-filling molecular model to illustrate this has been constructed. Soluble Cu(II):phytate complexes are formed within the pH range from 2 to ca. 3.4. This is supported by the results of difference absorption spectrometry.     

Phenotypic,genetic and molecular characterization of a maize low phytic acid mutant (lpa241)

Phytic acid, myo-inositol 1,2,3,4,5,6-hexakisphosphate, is the major storage compound of phosphorous (P) in plants, predominantly accumulating in seeds (up to 4–5% of dry weight) and pollen. In cereals, phytic acid is deposited in embryo and aleurone grain tissues as a mixed "phytate" salt of potassium and magnesium, although phytates contain other mineral cations such as iron and zinc. During germination, phytates are broken down by the action of phytases, releasing their P, minerals and myo-inositol which become available to the growing seedling. Phytic acid represents an anti-nutritional factor for animals, and isolation of maize low phytic acid (lpa) mutants provides a novel approach to study its biochemical pathway and to tackle the nutritional problems associated with it. Following chemical mutagenesis of pollen, we have isolated a viable recessive mutant named lpa 241 showing about 90% reduction of phytic acid and about a tenfold increase in seed-free phosphate content. Although germination rate was decreased by about 30% compared to wild-type, developement of mutant plants was apparentely unaffected. The results of the genetic, biochemical and molecular characterization experiments carried out by SSR mapping, MDD-HPLC and RT-PCR are consistent with a mutation affecting the MIPS1S gene, coding for the first enzyme of the phytic acid biosynthetic pathway.Communicated by F. Salamini     

Phytic acid-zinc ion interactions: a calorimetric and titrimetric study

Using an isoperibolic titration microcalorimeter, the ionization characteristics and associated heat changes of phytic acid (myo-inositol hexaphosphate) and phytic acid in the presence of varying Zn(II) concentrations have been examined over the pH range 2.5–11 at 25°C in 0.2 M KCl. In the absence of Zn(II), ca. 7 of the 12 ionizable protons in phytic acid are titrated in this pH range with ionization heats varying from ca. 2 to −3 kcal-mol^-1. At Zn(II): phytate mol ratios of 4:1 and greater, the dissociation of all protons and complex formation of phytate with Zn(II) occurs below pH 6. From the difference titration curves of phytic acid plus Zn(II) versus Zn(II) alone, ca. 3.5 mol Zn(II) bind per mol phytate. Since Zn(II):phytate complexes are insoluble, the observed heat changes contain contributions not only from heats of precipitation but also from binding, ionization, neutralization, and hydration effects. From the heat change for the titration of (a) phytic acid, pH 2.6–10.4; (b) phytic acid + Zn(II), pH 2.6–6.1; and (c) Zn(II), pH 2.6–6.1 at Zn(II): phytate ratios of 4 to 10, the value of 24.7 ± 0.5 kcal mol⁻¹ phytate has been obtained for the binding of 3.5 mols Zn(II). This figure also includes the heat of precipitation of the complex. In pH-drop experiments, with the initial pH at 8.65, the value of 23.9 kcal mol^-1 was obtained for ΔH°. Hysteresis effects are prevalent in these reaction solutions. Time-dependent changes in pH occur with a change in pH. For the phytate-Zn(II) reactions, the time-course curves are biphasic and fit a rate equation for two simultaneous first order reactions. Hysteresis effects seen in the titration of Zn(II) fit simple first-order kinetics. These effects most probably arise from the ejection of a proton from the aqua ion or aqua ion ligand complex(es).     

Phytate hydrolysis by germfree and conventional rats.

Phytic acid is naturally occurring compound that reduces intestinal absorption of many metals. Early work suggests that some dietary phytate may be hydrolyzed in the large intestines by bacteria, but more recently nutritionists have suggested that a mucosal enzyme is responsible. This paper reports a study intended to resolve this controversy. The hydrolysis of dietary phytic acid was measured in germfree and conventional rats fed either of two diets that differed in their calcium content. Negligible phytate hydrolysis occurred in the germfree rats, whereas 22 and 56% of the phytic acid was hydrolyzed by conventional rats fed high- and low-calcium diets, respectively. We concluded that bacteria were responsible for the hydrolysis of phytate in these diets and that any activity of endogenous enzyme was negligible.