Protein interactions with cyanidin‐3‐glucoside and its influence on α‐amylase activity

BACKGROUND: Recent studies indicate that the bioavailability of anthocyanins is extremely low. One of the possible reasons could be their binding to proteins. Therefore, the binding affinity of cyanidin‐3‐glucoside (Cy3glc) to HSA and α‐amylase was investigated by the quenching of protein tryptophan fluorescence. From data obtained, the binding constants and the free Gibbs energy were calculated. The changes in conformation of the proteins tested were studied with circular dichroism and the influence of binding on α‐amylase activity determined. RESULTS: Cy3glc quenched the tryptophan fluorescence and upon ligand binding a change in protein structure was observed related to the corresponding decrease in the α‐amylase activity. The association constants of 25 to 77 × 10³ L mol^?1 were calculated for different proteins, indicating weak interactions of non‐covalent nature. Competitive binding with HSA in the presence of 8‐anilino‐1‐naphthalene sulfonic acid suggest involvement of hydrophobic interactions, in the case of HSA the possible site being subdomain IIA. CONCLUSION: The strongest affinity of Cy3glc for HSA being at pH 7 underlines its potential in transport and distribution of the phenolic compounds in organisms. An influence on salivary amylase activity is possible when drinking berry juices with high anthocyanins content. Copyright © 2008 Society of Chemical Industry     

Absorption and twenty‐four‐hour metabolism time‐course of quercetin‐3‐O‐glucoside in rats,in vivo

Rats were meal‐fed a semi‐synthetic diet, with or without quercetin 3‐O‐glucoside (Q3G; 100 mg per meal) and groups of three were killed either fasting, or at 2, 5 and 24 post‐feeding. Flavonoids and their metabolites in the diet, stomach contents, small intestinal lumen and mucosa, caecal contents and plasma were determined by LC/MS. Q3G was not hydrolysed in the stomach, but deglycosylation and further metabolism occurred in the small intestinal mucosa. At least 17 flavonoid glucuronides were identified in the lumen and mucosa, with evidence of time‐dependent changes such as de‐ and re‐glucuronidation. Quercetin mono‐sulphate was also detected in the small intestinal contents. Metabolites were still present in tissue and plasma 24 h after feeding. There was also evidence of complex microbial processing of Q3G in the caecal lumen with the appearance of at least one methylquercetin‐mono‐glucuronide, mono‐sulphate unique to this site in the gut, together with phenolic acid derivatives. Intestinal flavonoid metabolism is thus a very complex process in mammals, involving both enterocytes and bacteria. Copyright © 2004 Society of Chemical Industry     

Adsorption of Shiga Toxin to Poly‐γ‐Glutamate Precipitated

We screened foods containing indigestible ingredients in the ability to adsorb Shiga toxin (Stx). When 5 mg of foods and dietary fibers such as dry vegetables and inulin were mixed and incubated with 0.5 mL of Stx solution (100 ng/mL) containing 0.5% bovine serum albumin, both Stx1 and Stx2 seemed to be adsorbed by only a fermented food, natto (a traditional Japanese food prepared from steamed soybeans by the biological action of Bacillus subtilis). We purified the Stx‐adsorbing substance from natto by extraction with H₂O, acid treatment, Proteinase K treatment, and an ion exchange chromatography. The purified substance showed an average molecular mass of about 600 kDa. We identified it as poly‐γ‐glutamate (PGA) by amino acid analysis of its hydrolysate and peptide analysis after its treatment with Proteinase K. Purified PGA (MW: molecular weight = about 600 kDa) was considered to adsorb both Stx1 and Stx2 when we separated adsorbed and unadsorbed Stxs (MW = about 72 kDa) by an ultrafiltration method with a centrifugal filter unit (MWCO: molecular weight cut‐off = 100 K). However, PGA with the ability to adsorb Stx was an insoluble form precipitated in the filter unit during centrifugation. PGA precipitated beyond the saturated density was also confirmed to well adsorb both Stx1 and Stx2 by an equilibrated dialysis method. To the best of our knowledge, this is the 1st report on food‐adsorbing Stx.