The Fate of Glutathione in Grape Berries Upon Crushing and the Expression of a Grape Glutathione Reductase During Berry Development

Reduced glutathione (GSH) is the most abundant free sulfhydryl-containing compound in grape berries (Vitis vinifera L.) at the onset of ripening. Upon crushing, GSH is incorporated into 2-S-glutathionyl caftaric acid, a process thought to limit browning of grape juice. Using 35S-GSH, the fate of GSH upon crushing was studied. In addition to forming the expected conjugate, nearly 40% of the GSH was incorporated into polar metabolites, which contained at least a small amount of oxidized glutathione (GSSG). A procedure was developed for the production of 35S-GSH conjugates using mushroom tyrosinase; the products were identical to those made using grape berry polyphenol oxidase. Yeast uptake of 35S-GSH and 35S-2-S-glutathionyl chlorogenic acid (GCQ) was studied in a model must. The results indicate that neither GSH nor its conjugate were taken up by yeast over a five-hour period. Following incubation of labeled GSH and GCQ in a white wine for 55 days, 35S-GCQ appeared unchanged, but 35S-GSH gave rise to six metabolites in addition to those polar ones also observed upon crushing. GSSG has not been previously reported in wines but this work suggests its presence. Because several reduced-sulfur thiols and disulfides have been associated with sulfur defects in wines, an enzymatic method was adapted for the determination of glutathione in wines.