Phytase activity in brown rice during steeping and sprouting.

Phytase in brown rice will be activated and accumulated during seed germination. Changes of phytase activity in brown rice during two stages of germination (steeping and sprouting) affected by process conditions were studied. It was shown that steeping led to significant decrease of phytase activity (p < 0.01), varying with steeping temperature and steeping medium composition. Steeping respectively in demineralized water, 0.5 mM CaCl2 and 0.1 M H2O2 at 20 °C for 24 h led to the lowest phytase activity in brown rice, which was only 25% of that in raw rice. At steeping stage, steeping media had no significant effect. During the sprouting stage, phytase activity increased with prolonged time and gradually reached stable levels, and with higher temperature in the range of 15-25 °C. Phytase activity in brown rice reached 320-340 U kg-(1) after 5 d sprouting. The evolution pattern of phytase activity during sprouting differed with the solutes previously used during steeping. Steeping either with CaCl2 or H2O2 caused a delay followed by a rapid activation of phytase, while for control, a gradual accumulation of phytase activity was observed. Compared with acidic and alkaline steeping solutions, demineralized water at neutral (6.8) pH provided the best pre-treatment prior to sprouting at 25 °C, to activate maximum levels of phytase. Extreme conditions, either strong acidic or alkaline inhibited activation of phytase, and changed appearance of brown rice kernel as well.

Depolymerisation optimisation of cranberry procyanidins and transport of resultant oligomers on monolayers of human intestinal epithelial Caco-2 cells.

Procyanidins in cranberries are predominantly polymers (>85%). The objective of this study was to optimise the depolymerisation of polymers and to investigate the absorption of resultant oligomers on Caco-2 cell monolayers. Depolymerisation conditions were optimised using response surface methodology. Depolymerisation, with or without added epicatechin, yielded 644 µg and 202 µg of oligomers (monomer through tetramers) per mg of partially purified polymers (PP), respectively. Oligomers (yielded from both methods) were transported through Caco-2 cell monolayer despite absorption rates being low. With the aid of response surface methodology, the optimum depolymerisation conditions were determined to be 60°C, 0.1M HCl in methanol and 3h without added epicatechin. The predicted maximum yield was 364 µg oligomers per mg of PP. The optimum depolymerisation condition with added epicatechin shared the same temperature, acid concentration and reaction time, in addition to an epicatechin/PP mass ratio of 2.19. Its predicted maximum oligomer yield was 1,089 µg/mg. The predicted yields were verified by experimental data.

Microbial catabolism of procyanidins by human gut microbiota.

SCOPE: A major portion of ingested procyanidins is degraded by human microbiota in the colon into various phenolic compounds. These microbial metabolites are thought to contribute to the health benefits of procyanidins in vivo. The objective of this study was to identify and quantify the microbial metabolites of procyanidins after anaerobic fermentation with human microbiota. METHODS AND RESULTS: (-)-Epicatechin, (+)-catechin, procyanidin B2, procyanidin A2, partially purified apple and cranberry procyanidins were incubated with human microbiota at a concentration equivalent to 0.5 mM epicatechin. GC-MS analysis showed that common metabolites of all six substrates were benzoic acid, 2-phenylacetic acid, 3-phenylpropionic acid, 2-(3'-hydroxyphenyl)acetic acid, 2-(4'-hydroxyphenyl)acetic acid, 3-(3'-hydroxyphenyl)propionic acid, and hydroxyphenylvaleric acid. 5-(3',4'-Dihydroxyphenyl)-γ-valerolactones and 5-(3'-hydroxyphenyl)-γ-valerolactones were identified as the microbial metabolites of epicatechin, catechin, procyanidin B2, and apple procyanidins but not from the procyanidin A2 or cranberry procyanidin ferments. 2-(3',4'-Dihydroxyphenyl)acetic acid was only found in the fermented broth of procyanidin B2, A2, apple, and cranberry procyanidins. The mass recoveries of microbial metabolites range from 20.0 to 56.9% for the six substrates after 24 h of fermentation. CONCLUSION: Procyanidins, both B-type and A-type can be degraded by human gut microbiota. The microbial metabolites may contribute to the bioactivities of procyanidins.

Transport of cranberry A-type procyanidin dimers, trimers, and tetramers across monolayers of human intestinal epithelial Caco-2 cells.

A-type procyanidin oligomers in cranberries are known to inhibit the adhesion of uropathogenic bacteria. B-type procyanidin dimers and trimers are absorbed by humans. The absorption of A-type procyanidins from cranberries in humans has not been demonstrated. This study examined the transport of A-type cranberry procyanidin dimers, trimers, and tetramers on differentiated human intestinal epithelial Caco-2 cell monolayers. Procyanidins were extracted from cranberries and purified using chromatographic methods. Fraction I contained predominantly A-type procyanidin dimer A2 [epicatechin-(2-O-7, 4-8)-epicatechin]. Fraction II contained primarily A-type trimers and tetramers, with B-type trimers, A-type pentamers, and A-type hexamers being minor components. Fraction I or II in solution was added onto the apical side of the Caco-2 cell membranes. The media at the basolateral side of the membranes were analyzed using HPLC-MS(n) after 2 h. Data indicated that procyanidin dimer A2 in fraction I and A-type trimers and tetramers in fraction II traversed across Caco-2 cell monolayers with transport ratio of 0.6%, 0.4%, and 0.2%, respectively. This study demonstrated that A-type dimers, trimers, and tetramers were transported across Caco-2 cells at low rates, suggesting that they could be absorbed by humans after cranberry consumption.

Effect of neutrase, alcalase, and papain hydrolysis of whey protein concentrates on iron uptake by Caco-2 cells.

Effects of enzymatic hydrolysates of whey protein concentrates (WPC) on iron absorption were studied using in vitro digestion combined with Caco-2 cell models for improved iron absorption. Neutrase- and papain-treated WPC could improve iron absorption; especially hydrolysates by Neutrase could significantly increase iron absorption to 12.8% compared to 3.8% in the control. Hydrolysates by alcalase had negative effects to the lowest at 0.57%. Two new bands at molecular weights (MW) around and below 10 kDa occurred at tricine-SDS-PAGE of hydrolysates by Neutrase, and one new band at MW below 10 kDa occurred in hydrolysates by papain. No new band was observed in hydrolysates by alcalase. Concentration of free amino acids indicated that, except for tyrosine and phenylalanine, amino acids in papain-treated hydrolysates were higher than that of alcalase, and no cysteine and proline were found in hydrolysates by alcalase. The results suggested that hydrolysate by Neutrase-treated WPC is a promising facilitator for iron absorption. Peptides of MW around and lower than 10 kDa and aspartic acid, serine, glutamic acid, glycin, cysteine, histidine, and proline may be contributors to enhancement.

In vitro solubility of calcium, iron, and zinc in rice bran treated with phytase, cellulase, and protease.

Absorption of minerals is inhibited by phytic acid, fiber, and protein because of the chelates formed. Response surface method (RSM) was used in this study to evaluate the effect of application of commercial phytase, protease, and cellulase in rice bran on the in vitro solubility of calcium (IVCa), iron (IVFe), and zinc (IVZn). It is shown that IVCa and IVZn were significantly improved by the application of phytase and cellulase, and the models of two second-order polynomials are recommended for prediction, with coefficients at R(2) = 0.86 and R(2) = 0.88, respectively. Interaction between protease and cellulase also significantly affected IVCa and IVZn. Cellulase had more efficiency than phytase on IVCa. Differing in its effect on Ca and Zn, phytase had a significant linear correlation with IVFe, and none of the other processing parameters exerted a significant effect. The largest increment for IVFe, IVCa, and IVZn occurred in the treatment with applications of phytase at 2.5 U g(-1) and protease and cellulase at 0.2% and 1%, respectively, which were 3.3, 3.6, and 4.3 times, respectively, that of the untreated material.