Analysis of solutions containing glutathione and inorganic nitrite: Application to nitroglycerin metabolism studies

Nitroglycerin (GTN) is metabolized to 1,2-dinitroglycerin (1,2-GDN) and 1,3-dinitroglycerin (1,3-GDN) in vivo and in liver homogenates. 1,2-GDN and 1,3-GDN are converted to isomers of glyceryl mononitrate (GMN) in vivo. The denitration reactions yield inorganic nitrite (NO − 2) which is oxidized to inorganic nitrate (NO − 3). Denitration involves utilization of glutathione (GSH). In attempting to use the Bratton—Marshall assay for NO − 2 in studies of GTN metabolism in vitro, and in attempting to use Ellman's reagent for GSH in the same research, apparent concentrations of both NO − 2 and GSH were noticed lower than anticipated. Apparent mutual interference by NO − 2 and GSH in their respective assays was then found. Development of a specific liquid chromatographic method for measurement of NO − 2, NO − 3, GSH and oxidized glutathione (GSSG) permitted the study of the interaction of NO − 2 and GSH, which yielded NO − 3 and GSSG.