Tea Flavanols Block Advanced Glycation of Lens Crystallins Induced by Dehydroascorbic Acid.

Growing evidence has shown that ascorbic acid (ASA) can contribute to protein glycation and the formation of advanced glycation end products (AGEs), especially in the lens. The mechanism by which ascorbic acid can cause protein glycation probably originates from its oxidized form, dehydroascorbic acid (DASA), which is a reactive dicarbonyl species. In the present study, we demonstrated for the first time that four tea flavanols, (-)-epigallocatechin 3-O-gallate (EGCG), (-)-epigallocatechin (EGC), (-)-epicatechin 3-O-gallate (ECG), and (-)-epicatechin (EC), could significantly trap DASA and consequently form 6C- or 8C-ascorbyl conjugates. Among these four flavanols, EGCG exerted the strongest trapping efficacy by capturing approximate 80% of DASA within 60 min. We successfully purified and identified seven 6C- or 8C-ascorbyl conjugates of flavanols from the chemical reaction between tea flavanols and DASA under slightly basic conditions. Of which, five ascorbyl conjugates, EGCGDASA-2, EGCDASA-2, ECGDASA-1, ECGDASA-2 and ECDASA-1, were recognized as novel compounds. The NMR data showed that positions 6 and 8 of the ring A of flavanols were the major active sites for trapping DASA. We further demonstrated that tea flavanols could effectively inhibit the formation of DASA-induced AGEs via trapping DASA in the bovine lens crystallin-DASA assay. In this assay, 8C-ascorbyl conjugates of flavanols were detected as the major adducts using LC-MS. This study suggests that daily consumption of beverages containing tea flavanols may prevent protein glycation in the lens induced by ascorbic acid and its oxidized products.

Uptake and cytotoxicity of ascorbic acid and dehydroascorbic acid in neuroblastoma (SK-N-SH) and neuroectodermal (SK-N-LO) cells.

Ascorbic acid (AA) was found to be cytotoxic to neuroblastoma cells in vitro and in vivo. In the present, study we investigated whether the reduced--(AA) or oxidized form (dehydroascorbic acid, DhAA) and its rapidly formed metabolites were the main cytotoxic agents. In neuroblastoma SK-N-SH cells, AA was found to be more cytotoxic than DhAA, although considerably higher amounts of [14C]DhAA than of [14C]AA were incorporated. In contrast, SK-N-LO cells derived from neuroectodermal tissue in fact showed a similar uptake but were much less injured by both substances. We observed that uptake of [14C]AA and [14C]DhAA was impaired in the presence of dithiothreitol and glutathione. Once inside the cell, [14C]DhAA was partially reduced to [14C]AA. From these data we conclude first that at least part of AA is oxidized prior to its uptake, and second that the reduced form of AA and perhaps ascorbyl radicals but not DhAA or its metabolites are the most important forms in mediating cytotoxic reactions in neuroblastoma cells. Furthermore, the results strengthen the previous assumption that AA acts as a pro-oxidant in neuroblastoma cells and supports its use in the treatment of neuroblastoma, especially in combination with existing chemotherapeutics.