Phytic Acid Hydrolysis and Soluble Zinc and Iron in Whole Wheat Bread As Affected by Calcium Containing Additives

The effects of nonfat dry milk, CaCO₃, CaCl₂ and MgCl₂ on phytate hydrolysis and on Zn and Fe availability was evaluated in whole wheat bread and in a model fermentation system. CaCO₃ and CaCl₂ both depressed phytate hydrolysis to an equivalent degree, but the milk had a greater effect than could be accounted for by its Ca content. MgCl₂ had a less pronounced effect than the Ca salts in both the bread and the model system. Increasing fermentation time in the model system increased phytate hydrolysis, but the effect was delayed when milk or CaCl₂ was added. Supplementing the bread dough with Ca equivalent to that typically contributed by Ca-containing additives caused 50% reductions in the available quantities of soluble (“free”) zinc and iron; the decreases observed with varying added amounts of Ca or milk exhibited a high degree of correlation with the observed increases in residual phytate P.     

Lactic Fermentation of Non-Tannin and High-Tannin Cereals: Effects on In Vitro Estimation of Iron Availability and Phytate Hydrolysis

The effect on iron availability estimated in vitro and phytate hydrolysis was investigated in non-tannin and high-tannin cereals, lactic fermented as flour/water slurries or gruels. A natural starter culture initiated fermentation and addition of germinated flour and phytase in the fermentation process was tested. Lactic fermentation of nontannin cereals with added flour germinated sorghum seeds or wheat phytase increased iron solubility from about 4% up to 9 and 50%, respectively. Soaking flour in water before adding starter culture had a similar effect. The increase in soluble iron was strongly related to enzymatic deeradation of phytate (p<0.001). The reduction of inositol hexa- and pentaphosphates was about 50% with added germinated flour. Reduction was > 90% after soaking the flour prior to fermentation and almost complete with 50 mg phytase added. High-tannin cereals showed a minor increase in soluble iron after fermentation, ascribed to the inhibitory effect of tannins (both on iron solubility and on enzymatic hydrolysis of phytate). Lactic-fermented cereal foods have a potential in developing countries to improve iron nutrition.     

The degradation of phytate by microbial and wheat phytases is dependent on the phytate matrix and the phytase origin

BACKGROUND: Phytases increase utilization of phytate phosphorus in feed. Since wheat is rich in endogenous phytase activity it was examined whether wheat phytases could improve phytate degradation compared to microbial phytases. Moreover, it was investigated whether enzymatic degradation of phytate is influenced by the matrix surrounding it. Phytate degradation was defined as the decrease in the sum of InsP₆ + InsP₅. RESULTS: Endogenous wheat phytase effectively degraded wheat InsP₆ + InsP₅ at pH 4 and pH 5, while this was not true for a recombinant wheat phytase or phytase extracted from wheat bran. Only microbial phytases were able to degrade InsP₆ + InsP₅ in the entire pH range from 3 to 5, which is relevant for feed applications. A microbial phytase was efficient towards InsP₆ + InsP₅ in different phytate samples, whereas the ability to degrade InsP₆ + InsP₅ in the different phytate samples ranged from 12% to 70% for the recombinant wheat phytase. CONCLUSION: Wheat phytase appeared to have an interesting potential. However, the wheat phytases studied could not improve phytate degradation compared to microbial phytases. The ability to degrade phytate in different phytate samples varied greatly for some phytases, indicating that phytase efficacy may be affected by the phytate matrix. Copyright © 2011 Society of Chemical Industry     

Application of Bifidobacteria as Starter Culture in Whole Wheat Sourdough Breadmaking

This investigation is aimed at developing a new cereal-based product, with increased nutritional quality, by using Bifidobacterium pseudocatenulatum ATCC 27919 as starter in whole wheat sourdough fermentation and evaluating its performance. Four different sourdough levels (5%, 10%, 15%, and 20% on flour basis) in bread dough formulation were analysed. The effects of the use of bifidobacteria in sourdough bread were comparatively evaluated with controls (yeast and/or chemically acidified sourdough with antibiotics). The sourdough and dough fermentative parameters analysed were pH, total titratable acidity, d/l-lactic and acetic acids. Bread performance was evaluated by specific volume, slice shape, crumb structure and firmness, crust and crumb colour, pH, total titratable acidity, and d/l-lactic and acetic acids, phytate, and lower myo-inositol phosphate contents. The sourdough breads showed similar technological quality to the control sample, with the exception of specific bread volume (decreased from 2.46 to 2.22 mL/g) and crumb firmness (increased from 2.61 to 3.18 N). Sourdough inoculated with bifidobacteria significantly increased the levels of organic acids in fermented dough and bread. The Bifidobacterium strain contributed to the fermentation process, increasing phytate hydrolysis during fermentation owing to the activation of endogenous cereal phytase and its own phytase, resulting in bread with significantly lower phytate levels (from 7.62 to 1.45 μmol/g of bread in dry matter). The inclusion of sourdough inoculated with bifidobacteria made possible the formulation of whole wheat bread with positive changes in starch thermal properties and a delay and decrease in amylopectin retrogradation.     

Peniophora lycii phytase is stabile and degrades phytate and solubilises minerals in vitro during simulation of gastrointestinal digestion in the pig

BACKGROUND: Microbial phytases (EC 3.1.3) are widely used in diets for monogastric animals to hydrolyse phytate present in the feed and thereby increase phosphorus and mineral availability. Previous work has shown that phytate solubility is strongly affected by calcium in the feed and by pH in the gastrointestinal (GI) tract, which may have an effect on phytase efficacy. An in vitro model simulating the GI tract of pigs was used to study the survival of Peniophora lycii phytase and the effect of the phytase on phytate degradation, inositol phosphate formation and mineral solubilisation during in vitro digestion of a 30:70 soybean meal/maize meal blend with different calcium levels. RESULTS: The phytase retained 76 and 80% of its initial activity throughout the gastric in vitro digestion. Total phytate hydrolysis by P. lycii phytase was in the same range at total calcium levels of 1.2 and 6.2 mg g^?1 dry matter (DM), despite very large differences in phytate solubility at these calcium levels. However, at 11.2 and 21.2 mg Ca g^?1 DM, phytate hydrolysis was significantly lower. The amount of soluble mineral was generally increased by P. lycii phytase. CONCLUSION: Stability of P. lycii phytase during gastric digestion was not found to be critical for phytate hydrolysis. Furthermore, original phytate solubility was not an absolute requirement for phytate degradation; phytate solubility seemed to be in a steady state, allowing insoluble phytate to solubilise as soluble phytate was degraded. This is new and interesting knowledge that adds to the current understanding of phytate–phytase interaction. Copyright © 2007 Society of Chemical Industry     

Enzymatic Reduction of Phytate in Whole Wheat Breads

The presence of phytate in flour may be responsible for reduced bioavailability of iron, magnesium, zinc, and calcium from bread. The effect of various concentrations of commercial phytase or phosphatase added to whole wheat flour-yeast doughs on their phytate and nonphytate phosphorus content has been investigated. By using 2.0% (flour basis) of phytase and 0.11% phosphatase the initial phytate phosphorus concentration of the dough was reduced to 1/8 and 1/12 of its initial values, respectively. Storage of the whole wheat breads for up to 96 hr at room temperature showed further significant reduction of phytate phosphorus. The phytate phosphorus content of yeast leavened whole wheat breads decreased during 2 hours of dough fermentation, baking and the subsequent 48 hours of storage at room temperature from 24 mg/100g dough (dry matter) to 1.7 mg/100g bread (dry matter); the phytate phosphorus continued to decrease and after 96 hours storage it was 0.6 mg/100g bread.     

Supplementation of alkaline phytase (Ds11) in whole-wheat bread reduces phytate content and improves mineral solubility

Abstract: In this study, alkaline phytase was added to whole‐wheat bread and the phytate content and mineral profiles were compared to commercially available acidic phytase. At neutral pH, some phytate (approximately 20%) was degraded by endogenous phytase in wheat flour, while 40% of phytate was hydrolyzed by alkaline phytase DS11 and a 35% reduction was observed with acidic phytase. Most of the enzymatic activity occurred during the proofing stage, and the rate of reaction depended on pH. DS11 phytase effectively degraded the phytate level within a 30 min treatment at pH 7; however, at least 60 min was needed with acidic phytase to achieve the same hydrolysis level. Mineral profiles were also dramatically affected by the phytate reduction. The biggest increase was observed in Fe²⁺ by the phytase treatment. The Fe²⁺ content increased 10‐fold at pH 7 and 8‐fold at pH 5 with alkaline phytase DS11. Alkaline phytase DS11 was shown to be effective at phytate reduction in whole‐wheat bread preparation. Additionally, phytate degradation enhanced the mineral availability of bread.     

Sourdough fermentation of wheat fractions rich in fibres before their use in processed food

Fibre‐rich fractions of wheat are an important source of minerals but also contain considerable amounts of phytic acid, known to impair mineral absorption. This study explores the efficiency of wheat bran sourdough fermentation on phytate hydrolysis and mineral solubility, in comparison with whole‐wheat flour. In vitro trials were performed to assess the consequences of the addition of calcium carbonate (CaCO₃), an alkalinising salt, on phytic acid breakdown and mineral bioavailability during sourdough fermentation. Sourdough fermentation was found effective for solubilising minerals in whole‐wheat flours but was less effective with bran. In addition, sourdough acidity was blunted by the addition of CaCO₃, whereas degradation of phytic acid remained effective. Despite extensive breakdown of phytic acid (almost 70%), the addition of calcium exerted a very negative effect on zinc solubility. In conclusion, a pre‐fermentation process of whole cereals or bran, in suitable conditions of hydration, allows degradation of the major part of phytic acid and optimal mineral bioavailability. Copyright © 2007 Society of Chemical Industry     

Fermentation and enzyme treatment of tannin sorghum gruels: effects on phenolic compounds,phytate and in vitro accessible iron

The presence of polyphenols and phytate in cereal products has been shown to interfere with the bioavailability of minerals such as iron. In the present study, we added enzymes (wheat phytase and mushroom polyphenol oxidase) during fermentation of tannin sorghum gruels prepared from flour with or without addition of 5% flour of germinated tannin-free sorghum grains (power flour), and investigated the effects on phenolic compounds, phytate and in vitro accessible iron. Assayable phenolic compounds were significantly reduced by fermentation, with high reductions observed in gruels with added enzymes. Fermentation of the gruels with addition of enzymes reduced (on average) total phenols by 57%, catechols by 59%, galloyls by 70% and resorcinols by 73%. The phytate content was significantly reduced by fermentation (39%), with an even greater effect after addition of power flour (72%). The largest reduction of phytate (88%) was, however, obtained after addition of phytase. The in vitro accessible iron was 1.0% in the sorghum flour and it increased after fermentation with power flour and/or with enzymes. The highest in vitro accessibility of iron (3.1%) was obtained when sorghum was fermented with addition of power flour and incubated with phytase and polyphenol oxidase after the fermentation process.     

植酸钙的提取及固定化细胞植酸酶酶解

本文重点研究了以麦麸为原料提取植酸钙的工艺条件,最佳工艺条件为60℃水浴浸泡6h,通过浸泡方式的改进,以二次浸泡和动态摇床浸泡两种方式进行实验,结果发现这两种方式均使植酸钙得率提高。另外,利用在细胞壁表面产植酸酶的克鲁维假丝酵母,采用细胞固定化技术,进行植酸酶酶解。细胞固定化和酶解反应的方法都比较简单,而且多次反应后酶解率损失较低。此法为肌醇、磷酸肌醇的酶法制取提供了可行性方案。     

Phytate Hydrolysis by Phytase in Cereals; Effects on In Vitro Estimation of Iron Availability

Phytate (inositol hexaphosphate) hydrolysis by endogenous and exogenous phytases was studied for their effect on increasing iron availability in cereals. Wheat bran and whole meal flours of rye and oats were soaked at optimal conditions for phytase activity (55°C, pH 5) for different time intervals. Phytate and its degradation products were determined by HPLC and related to iron solubility under simulated physiological conditions. Small amounts of phytate (< lμmol/g) had a strong negative effect on iron solubility. When inositol hexa- and pentaphosphates of wheat bran and rye flour were completely hydrolyzed by activating endogenous phytase, iron solubility under simulated physiological conditions increased from 3 to 53% (wheat) and 5 to 21% (rye). Addition of wheat phytase to uncooked oatmeal increased iron solubility from 4 to 11 and in precooked to 18%, while endogenous phytase of uncooked oatmeal had less effect on phytate digestion and iron solubility.     

Bioavailability of Calcium from Sesame Seeds, Almond Powder, Whole Wheat Bread, Spinach and Nonfat Dry Milk in Rats

Calcium bioavailability (BV) from sesame seeds, almond powder, whole wheat bread, spinach, and nonfat dry milk (NFDM) was compared to calcium BV from a calcium carbonate (CaCO₃)-supplemented control diet using a rat model. When comparing different calcium sources, the relative BV of the products was CaCO₃ (100%), NFDM (100%) whole wheat bread (95%), almond powder (66%), sesame seeds (65%), NFDM and spinach mixture (52%), and spinach (47%). Separate almond, NFDM, and CaCO₃ diets were supplemented with 0.4% ascorbic acid; vitamin C addition had no significant effect on calcium BV.     

Simultaneous application of phytase and xylanase to broiler feeds based on wheat: in vitro measurements of phosphorus and pentose release from wheats and wheat-based feeds†

An in vitro procedure that simulated digestion in growing broilers was tested to predict phosphorus availability and arabinoxylan hydrolysis in samples of nine wheat varieties and in a wheat-based diet. Amounts of dialysable phosphorus freed from wheat samples correlated with activities of endogenous phytase (R = 0.913; p < 0.0001), whereas amounts of pentoses released were correlated with viscosities of the digested samples (R = 0.899; p < 0.0001). Differences in phosphorus release resulting from graded levels of microbial phytase added to feeds that were either autoclaved or not autoclaved revealed a decreasing role of endogenous phytase in dephosphorylation as levels of microbial phytase supplementation grew. Amounts of pentoses released from feeds containing two different xylanase preparations reflected literature data on different in vivo efficacies of those preparations. Simultaneous addition of phytase and xylanase affected phosphorus release in a manner that depended upon the form of xylanase preparation used (liquid or powder). There was a positive influence of acid protease on both phytate and arabinoxylan hydrolysis in feeds supplemented with phytase. Effects observed by the in vitro procedures corresponded to in vivo phenomena described in the literature. © 1999 Society of Chemical Industry     

PURIFICATION AND CHARACTERIZATION OF A PHYTASE FROM SPELT

Four soluble phytases were identified in germinating spelt. Although numerous purification strategies were applied, none of the four phytases could be purified to homogeneity. The purest phytase preparation, called D21, contained a phytase (major component) and an acid phosphatase (APH) (minor component). The phytase behaves like a monomeric protein of a molecular mass of about 68 kDa and shows a broad substrate specificity. Optimal pH for degradation of phytate was 6.0 and the optimal temperature 45C. Kinetic parameters for the hydrolysis of Na-phytate were K_M 400 μmoll^?1 and k_cat 368s^?1 at pH 6.0. The spelt phytase D21 degrades phytate stepwise.     

Phytate degradation by Bifidobacterium on whole wheat fermentation

Whole cereal-based products have a beneficial effect on health, but they also contain high levels of phytate, defined as anti-nutrient. The possible use of different strains of bifidobacterial species (B. catenulatum, B. longum and B. breve) with phytate degrading activity as starters in the fermentation process was investigated. Fermentative parameters of doughs (pH, volume, total titrable acidity [TTA] and acetic and lactic acids production) and the hydrolysis profile of the phytic acid were determined during whole wheat dough fermentation. During fermentation in the presence of different bifidobacterial strains, the concentration of phytic acid showed a progressive decrease, leading to a higher release of hydrolysis products than in the control, within short fermentation time. According to the fermentative parameters bifidobacterial strains showed a good adaptation to the dough ecosystem. The range of TTA and lactic acid production was dependent on the strain. Among the tested bifidobacterial strains, those belonging to the species B. breve and B. longum induced a higher hydrolysis of phytic acid with simultaneous production of lower inositol phosphates. In addition, dough containing those strains had high pH and minor acidity than those containing a commercial starter (L. plantarum). Therefore, the tested bifidobacterial strains could be used as breadmaking starters contributing to different acidification degrees and promoting simultaneously the degradation of phytic acid in the whole wheat dough.     

体外消化/Caco-2细胞模型分析发酵对面粉铁生物利用率的影响

中国居民膳食以谷物为主食,谷物中铁吸收率低是造成营养性贫血的主要原因,适当的加工方式可缓解谷物中植酸、多酚等物质对铁生物利用率(Fe bioavailability,FeBV)的影响。为考察酵母发酵对面粉FeBV的影响,采用体外消化/Caco-2细胞模型。结果表明,面团在发酵后pH值呈下降趋势,酸度呈上升趋势;多酚、植酸含量降低,植酸酶活性升高,以上各指标在发酵前后差异均具有统计学意义(P0.05)。发酵后面粉样品的FeBV增加约5%~38%,大多数面粉FeBV发酵前后比较,差异具有统计学意义(P0.05)。结论:发酵可降低面粉的pH值,增加酸度,有利于面粉多酚和植酸的降解以及植酸酶活力的升高,可有效提高面粉FeBV。     

Monitoring the stepwise phytate degradation in the upper gastrointestinal tract of pigs

The degradation and formation of inositol phosphates as affected by microbial phytase and gastrointestinal enzyme activities during the passage of phytate through the stomach and small intestine were studied in two experiments with four barrows and three collection periods. The degradation and formation of inositol phosphates were measured at the duodenal and ileal sites using Cr‐NDR, TiO₂ and Co‐EDTA as indigestible markers. In experiment 1, the effect of graded doses of Aspergillus niger phytase (0, 150 and 900 FTU Natuphos^® kg^?1), added to a maize–soybean meal‐based diet with very low intrinsic phytase activity on the degradation of phytate and the formation of inositol phosphates during digestion in the stomach and small intestine was investigated. In experiment 2, three different mixtures of inositol phosphates, produced by Aspergillus niger phytase, containing mainly high, intermediate and low phosphorylated inositol phosphates, were added to the same maize–soybean meal‐based diet as used in experiment I. The fate of the inositol phosphates during digestion in the stomach and small intestine was studied. Experiment 1 showed that the extent of phytate degradation was dependent of the graded dietary phytase activities. At high phytase activity (900 FTU kg^?1 of diet), strong phytate degradation occurred and the once hydrolysed phytate was rapidly dephosphorylated to lower inositol phosphates (mainly inositol di‐ and triphosphates). Intermediate inositol phosphates, such as inositol tetraphosphates, were quantitatively unimportant in duodenal and ileal digesta. At a phytase activity of 150 FTU kg^?1 of diet, a broader spectrum of intermediate inositol phosphates was determined, which was probably due to a slower breakdown of phytate. Experiment 2 showed as a predominant result that lower inositol phosphates such InsP₄ and InsP₃ were degraded, whereas InsP₂ accumulated in the duodenal and ileal digesta. No substantial disappearance of phytate from the stomach and small intestine was found when high concentrations of soluble phytate were added to the diet, which indicates that no substantial phytate absorption occurs in the upper part of the pig gut. Copyright © 2005 Society of Chemical Industry     

Phytase Activity from Lactobacillus spp. in Calcium-Fortified Soymilk

Abstract: The presence of phytate in calcium-fortified soymilk may interfere with mineral absorption. Certain lactic acid bacteria (LAB) produce the enzyme phytase that degrades phytates and therefore may potentially improve mineral bioavailability and absorption. This study investigates the phytase activity and phytate degradation potential of 7 strains of LAB including: Lactobacillus acidophilus ATCC4962, ATCC33200, ATCC4356, ATCC4161, L. casei ASCC290, L. plantarum ASCC276, and L. fermentum VRI-003. Activity of these bacteria was examined both in screening media and in calcium-fortified soymilk supplemented with potassium phytate. Most strains produced phytase under both conditions with L. acidophilus ATCC4161 showing the highest activity. Phytase activity in fortified soymilk fermented with L. acidophilus ATCC4962 and L. acidophilus ATCC4161 increased by 85% and 91%, respectively, between 12 h and 24 h of fermentation. All strains expressed peak phytase activity at approximately pH 5. However, no phytate degradation could be observed.     

Effect of exogenous phytase on feed inositol phosphate hydrolysis in an in vitro rumen fluid buffer system

Three in vitro experiments using a rumen fluid buffer system were performed to investigate the effect of addition of 4 experimental phytases (Phy1, Phy2, Phy3, and Phy4) compared with no addition of phytase on feed inositol phosphate hydrolysis in wheat and rapeseed cake to determine which of the 4 phytases was most suitable under rumen-like conditions. The feedstuffs were incubated with a mixture of physiological buffer, ruminal fluid, and exogenous phytase at pH 6.2, after which the samples were incubated for different periods. Incubations were stopped using HCl, and the samples were analyzed for inositol phosphates via high performance ion chromatography. Addition of phytase (Phy1) resulted in enhanced degradation of myo-inositol hexakisphosphate (InsP₆) in rapeseed cake, whereas addition of exogenous phytase did not improve the degradation of InsP₆ in wheat. Only rapeseed cake was therefore used subsequently. All 4 phytases increased degradation of InsP₆ in rapeseed cake in the in vitro system, and degradability of InsP₆ increased with higher incubation time and higher phytase dosages, independent of phytase. Addition of 2 units of phytase per gram of substrate of the phytases Phy1, Phy2, Phy3, and Phy4 led to an undegraded InsP₆ content of 56, 49, 70, and 18%, respectively, when incubated with rapeseed cake for 6 h, indicating that Phy2 and Phy4 were the most effective phytases. However, Phy2 had a higher specific activity than Phy4, as 60% of the original InsP₆ content was remaining after 3 h when 5 mg of enzyme protein per gram of substrate of Phy2 was added to rapeseed cake, whereas 150 mg of enzyme protein per gram of substrate of Phy4 was necessary to achieve a similar result. Therefore, Phy2 appeared to be most applicable under rumen-like conditions.     

Purification and characterization of a phytate-degrading enzyme from germinated oat (Avena sativa)

A phytate-degrading enzyme (myo-inositol hexakisphosphate phosphohydrolase) has been purified about 5,400-fold from germinated oat seedlings to apparent homogeneity. The molecular mass of the native monomeric enzyme was estimated to be about 67 kDa. Optimal pH for degradation of phytate was 5.0 and the optimal temperature 38 °C. Kinetic parameters for the hydrolysis of Na-phytate are K_M 30 µM and k_cat 356 s⁻¹ at 35 °C and pH 5.0. The oat phytase exhibits a broad affinity for various phosphorylated compounds and hydrolyses phytate in a stepwise manner. The first hydrolysis product was identified as D /L -l(1,2,3,4,5) P₅. © 1999 Society of Chemical Industry