The aldehyde 3-S-glutathionylhexanal is an intermediate which is produced during the formation of the wine aroma precursor 3-S-glutathionylhexanol, after the reaction of glutathione with trans-2-hexenal. This study was conducted to assess whether the chemical, as opposed to the enzymatic, production of 3-S-glutathionylhexanal could occur at a significant rate in grape juice. LC-MS/MS was used in low- and high-resolution modes, in combination with functional group derivatization, to identify and quantitate products. In comparison to cysteine, glutathione was found to induce less cyclized products on reaction with trans-2-alkanals and the glutathione-derived products were more reactive to hydrogen sulfite. The zero-order rates for 3-S-glutathionylhexanal formation in model grape juice were 1.08 ± 0.08 and 0.45 ± 0.05 mg/(L·day) glutathione equivalents at 25 and 13 °C, respectively, and the reaction rate increased 3-fold by increasing the pH from 3.2 to 3.8. 3-S-Glutathionylhexanal was detected in all five white grape juices examined. The concentration of the aldehyde could be increased by up to 10-fold after being released from hydrogen sulfite, demonstrating a potentially novel source for the production of varietal thiol aroma compounds in wine.