Stability to light, heat, and hydrogen peroxide at different pH values and DPPH radical scavenging activity of acylated anthocyanins from red radish extract.

The stability of red radish extract to light, heat, and hydrogen peroxide at different pH values (3, 5, and 7) was examined, in which major anthocyanins were pelargonidin glycosides acylated with a combination of p-coumaric, ferulic, or caffeic acids. The light irradiation (fluorescence light, 5000 lx; at 25 degrees C) indicated that the red radish extract was more stable at lower pH than at higher pH. The HPLC analyses revealed that diacylated anthocyanins in the extract were more stable to light at pH 3 than monoacylated anthocyanins. No significant difference in degradation rates of acylated anthocyanins at pH 5 was observed, whereas anthocyanins acylated with p-coumaric or ferulic acids were more stable at pH 7 than ones with caffeic acids. The stability to heat (at 90-95 degrees C) showed a tendency similar to that for light. The number of intramolecular acyl units contributes to stability to light and heat at lower pH, whereas the characteristics of intramolecular acyl units influence the stability at higher pH. The degradation behavior of red radish extract to H2O2 were almost the same to those of light and heat, depending on the pH. However, HPLC analyses revealed that the stability of individual acylated anthocyanins were independent of the pH. These data suggest that the characteristics, the number, and the binding site of intramolecular acyl units affect the stability of anthocyanin to H2O2. DPPH radical scavenging activity of all acylated anthocyanins was higher than those of pelargonidin and perlargonidin-3-glucoside. The activity of acylated anthocyanins mostly depended on the activity of intramolecular acyl units (caffeic acid > ferulic acid > p-coumaric acid). However, the activity was highly affected by the binding site of intramolecular acyl units even if anthocyanins have common acyl units.

Effects of paprika pigments on oxidation of linoleic acid stored in the dark or exposed to light.

We examined the antioxidant effects of paprika pigments on oxidation of linoleic acid and on decoloration of the sample when stored at 37 degrees C in the dark or exposed to fluorescent light for 8 h per day. (1)H nuclear magnetic resonance with dioxane as an external proton reference was used to estimate the oxidative deterioration of linoleic acid. Oxidation was estimated by observing the ratio of the divinylmethylene proton signal area in linoleic acid vs the proton signal area in dioxane. The addition of paprika pigments suppressed the oxidation of linoleic acid during storage in the dark, and the effect was markedly increased with increasing concentrations (0.02, 0.2, and 2%). When the linoleic acid with added paprika pigments was exposed to light, only a slight suppression of oxidation was observed, and the color of the sample disappeared more rapidly than that in the dark. At the time of decoloration of the sample with added pigments, considerable oxidation of linoleic acid occurred. As the color change is due to degradation of the pigment, an increase in oxidation at the time of discoloration is consistent with the pigments functioning as antioxidants. The addition of alpha-tocopherol to paprika pigments stabilized degradation of the pigments by light. Although the addition of alpha-tocopherol to linoleic acid with added paprika pigments prolonged the decoloration of the sample under light, the prevention of oxidation under the light condition was not as effective as for the samples stored in the dark.

Identification and behavior of reaction products formed by chlorination of ethynylestradiol.

Six products were formed by reaction of ethynylestradiol (EE2) with sodium hypochlorite in buffered solutions. 4-Chloroethynylestradiol (4-ClEE2) and 2,4-dichloroethynylestradiol (2,4-diClEE2) were identified as the two major reaction products, using preparative HPLC, MS, and NMR. When EE2 reacted with chlorine at different pHs (pH 5, 7, and 9) or chlorine concentrations (0.2, 1, 2, and 5 mmol/l, corresponding to molar ratios to EE2, 1, 5, 10, and 25, respectively), the formation of 4-ClEE2 and 2,4-diClEE2 was observed under the above conditions, and the highest yields were 20 and 52 mol%, respectively. EE2 was consumed almost completely within 5 min of chlorination by addition of chlorine of more than 1 mmol/l (molar ratio to EE2, 5). On the other hand, the two products existed in highly chlorinated solutions after 60 min (4ClEE2, 1-6 mol%; 2,4-diClEE2, 3-25 mol%). The estrogenic activities of 4-ClEE2 by estrogen receptor alpha or beta binding assay were similar to those of the parent EE2, and the activities of 2,4-diClEE2 were lower about 10 times.

Identification of reaction products of acylated anthocyanins from red radish with peroxyl radicals.

Red radish anthocyanin extract, which consists of 12 known acylated anthocyanins, was reacted with 2,2'-azobis(2-amidinopropane) dihydrochloride (AAPH) to generate peroxyl radicals under acidic pH conditions at 37 degrees C. The reaction products were isolated using preparative HPLC, and their chemical structures were determined to be p-hydroxybenzoic acid (1), 6-O-(E)-p-coumaroyl-2-O-beta-d- glucopyranosyl-alpha-d-glucopyranoside (3), p-coumaric acid (4), 6-O-(E)-feruloyl-2-O-beta-d-glucopyranosyl-alpha-d-glucopyranoside (5), and ferulic acid (6). Some products were not identified. HPLC analyses of the mixture of acylated pelargonidin isolated from red radish and AAPH revealed that the acylated pelargonidins possess the radical scavenging ability on some common sites even if the characteristics of the intramolecular acyl units are different. Degradation rates of acylated pelargonidins and the formation rates of the resulting reaction products were found to be quite different.

Acylated anthocyanins from red radish (Raphanus sativus L.).

Twelve acylated anthocyanins were isolated from the red radish (Raphanus sativus L.) and their structures were determined by spectroscopic analyses. Six of these were identified as pelargonidin 3-O-[6-O-(E)-feruloyl-2-O-beta-D-glucopyranosyl]-(1-->2)-beta-D-glucopyranoside]-5-O-(beta-D-glucopyranoside), pelargonidin 3-O-[6-O-(E)-caffeoyl-2-O-(6-(E)-feruloyl-beta-D-glucopyranosyl)-(1-->2)-beta-D-glucopyranoside]-5-O-(beta-D-glucopyranoside), pelargonidin 3-O-[6-O-(E)-p-coumaroyl-2-O-(6-(E)-caffeoyl-beta-D-glucopyranosyl)-(1-->2)-beta-D-glucopyranoside]-5-O-(beta-D-glucopyranoside), pelargonidin 3-O-[6-O-(E)-feruloyl-2-O-(6-(E)-caffeoyl-beta-D-glucopyranosyl)-(1-->2)-beta-D-glucopyranoside]-5-O-(beta-D-glucopyranoside), pelargonidin 3-O-[6-O-(E)-p-coumaroyl-2-O-(6-(E)-feruloyl-beta-D-glucopyranosyl)-(1-->2)-beta-D-glucopyranoside]-5-O-(beta-D-glucopyranoside), and pelargonidin 3-O-[6-O-(E)-feruloyl-2-O-(2-(E)-feruloyl-beta-D-glucopyranosyl)-(1-->2)-beta-D-glucopyranoside]-5-O-(beta-D-glucopyranoside).