PROTECTIVE ACTIVITY OF HAMAMELITANNIN ON CELL-DAMAGE INDUCED BY SUPEROXIDE ANION-RADICALS IN MURINE DERMAL FIBROBLASTS

Previously we demonstrated that hamamelitannin (2',5-di-O-galloyl hama melose) in Hamamelis virginiana L. exhibits potent superoxide-anion sc avenging activity. We then examined the physiological and pharmacologi cal activities of hamamelitannin as well as its functional homologues, gallic acid and syringic acid. The following results were obtained: ( 1) Hamamelitannin has a higher protective activity against cell damage s induced by superoxide anions than gallic acid which is the functiona l moiety of hamamelitannin. The protective activity of hamamelitannin on murine fibroblast-damage induced by superoxide anions was found at a minimum concentration of 50 mu M, while the corresponding figure for gallic acid was 100 mu M. (2) Pre-treatment of fibroblasts with hamam elitannin enhances cell survival. (3) The superoxide-anion scavenging activity of the compound in terms of its IC50 value (50% inhibition co ncentration of superoxide anion radicals generated) was evaluated by E SR spin-trapping. Both hamamelitannin (IC50=1.31+/-0.06 mu M) and gall ic acid (IC50=1.01+/-0.03 mu M) exhibited high superoxide-anion scaven ging activity followed by syringic acid (IC50=13.90+/-2.38 mu M). (4) When hamamelitannin was treated with superoxide anions generated by a KO2-crown ether system, HPLC analysis showed the disappearance of hama melitannin and the concomitant formation of hamamelitannin-derived rad icals (g=2.005, Delta H-1=2.16 G, Delta H-2=4.69 G) was detected by ES R spectrometry. From these observations, we concluded that (i) a compo und,vith polyphenolic hydroxyl groups, especially the galloyl group (3 ,4,5-trihydroxy-), has potent scavenging activity against superoxide a nions, and (ii) hamamelitannin is superior to gallic acid in protectin g against cell damage induced by superoxide anions, suggesting that th e high affinity of hamamelitannin for cells or membranes may be an imp ortant factor for protecting cells against active oxygen species.