Yoshihiko Hatano

Superoxide radical- and peroxynitrite-scavenging activity of anthocyanins; structure-activity relationship and their synergism.

Antioxidant activities of 15 purified bilberry anthocyanins together with pelargonidin 3-O-β-d-glucopyranoside and 4′-O-methyl delphinidin 3-O-β-d-glucopyranoside (MDp 3-glc), the major metabolite of delphinidin 3-O-β-d-glucopyranoside (Dp 3-glc), were evaluated in order to study the structure-antioxidant activity relationship and any synergism among them in the mixture. Both aglycone structure and the attached sugar moiety affected the - and ONOO− -scavenging activities, although the effect of the attached sugar moiety was smaller than that of the aglycone structure. The potency of activity toward the superoxide radical was in the following order: delphinidin > petunidin > malvidin ≈ cyanidin>(+)-catechin > peonidin > pelargonidin. The activity toward ONOO− was: delphinidin > cyanidin ≈ petunidin > malvidin ≈ (+)-catechin > peonidin > pelargonidin. It was confirmed that methylation of 4′-OH markedly reduced the antioxidant activity of anthocyanin. Further, it was revealed that synergism occurred in both - and ONOO− -scavenging activities among the anthocyanins in the mixture.

Simultaneous determination of membrane potential and pH gradient by photodiode array spectroscopy

Membrane potential (delta psi) and pH difference (delta pH) were simultaneously determined in liposomes using a photodiode array spectrophotometer. By the use of a cyanine dye (DiS-C3(5)) and 9-aminoacridine for delta psi and delta pH probes, respectively, both changes of delta psi and delta pH could be successfully determined by photodiode array spectrometry. Each dye did not disturb the fluorescence spectrum of the other probe when its concentration was lower than 5 microM. The K+-diffusion potential-driven, FCCP(protonophore)-mediated H+-influx process in the K+-loaded liposomes was analyzed by this method. Results indicate that the kinetic behavior of H+ influx changes at a FCCP concentration of approx. 30 nM. The rate of delta pH formation increased quantitatively with increasing concentrations of FCCP up to 30 nM, but was markedly enhanced at higher concentrations, although the maximal delta pH attained was about 3 pH units in any case when a K+-diffusion potential of -180 mV was applied.

Structural elucidation and biological fate of two glucuronyl metabolites of pelargonidin 3-O-β-D-glucopyranoside in rats.

A high proportion of pelargonidin 3-O-β-D-glucopyranoside (Pg3G) is metabolized to glucuronides and excreted in mammal urine after ingestion of strawberry fruit, suggesting that these metabolites play important functional roles in vivo. The aim of the present study was to elucidate the structures and determine the biological fate of the two dominant metabolites of Pg3G in rats to enable an accurate discussion of the biological properties of anthocyanins. Authentic Pg3G was orally administered to rats. One pelargonidin monoglucuronide and three Pg3G-monoglucuronides (glucuronides of the glucoside) were identified together with intact Pg3G in both blood plasma and urine samples. The structures of the two dominant metabolites were elucidated as pelargonidin 3-O-β-D-glucuronide (Pg3GlcA) and pelargonidin 3-O-β-D-glucuronyl-(1→2)-β-D-glucoside by means of (1)H and (13)C nuclear magnetic resonance spectroscopy and heteronuclear multiple-bond connective spectroscopy. The bioavailability of Pg3G in its intact form was 0.31% of the orally administered dose, and 0.65% was absorbed in the Pg3GlcA form.

Degradation mediated OH radical generation from synthetic cyclic peroxides: ESR studies

Hydroxy radical generation on thermal decomposition of synthetic cyclic peroxides was determined by ESR using DMPO as a spin‐trapping reagent. Both the 6‐ and 7‐membered cyclic peroxides, 4‐alkoxy‐2,3‐benzodioxan‐1‐ols and 4‐alkoxy‐2,3‐benzodioxepin‐1‐ols, respectively, which were synthesized according to our idea of a radical‐releasing drug, gave rise to DMPO‐OH signals on heat treatment, i.e. under their degradation conditions. The signals were completely abolished with higher concentrations of OH scavenger.