A simple, convenient, and quantitative method for the preparation of methylglyoxal (pyruvaldehyde) solutions is described. The method involved acid hydrolysis (5% HZS04) of pyruvaldehyde dimethyl acetal at 100Β°C for 25 min. The hydrolysis method is quite reproducible and does not require standardization ofthe methylglyoxal solutions. The preparations of methylglyoxal by the procedure described have been used in the study of glyoxalase I activity.
The glyoxalase system, which has been known since 1913 (1,2), is widely distributed in nature and catalyzes the conversion of methylglyoxal (MG) to lactate with reduced glutathione (GSH) as cofactor (3-6). The system consists of two enzymes, glyoxalase I and glyoxalase II (7). Glyoxalase I acts upon the equilibrium adduct of MG and GSH, a hemimercaptal, with the resultant formation of the thioester, S-D-lactoylglutathione.
Glyoxalase II hydrolyzes the thioester to regenerate GSH and liberate free D-lactic acid.
Methylglyoxal, for use in enzymatic studies, is usually obtained by vacuum (8) or steam distillation (9) of a commercial 40% solution in order to eliminate polymerized MG. The pale green distillate may then be passed through an anion exchange resin (8, 10) to obtain acid-free MG. The MG is then standardized by Friedemann's titration method (11) after oxidation with H202, or enzymatically (12), or by the semicarbazide method of Alexander and Boyer (13).
This tedious method of distillation, removal of acidic contaminants, and quantitation is time consuming. Furthermore, MG is very unstable and readily undergoes polymerization and oxidation reactions (15). The MG content of solutions stored for even short time periods is questionable at best. For those engaged in a continuing study of glyoxalase I, a simple, convenient, and reproducible method of MG preparation is needed.
We hereby report a rapid and direct method for the conversion of pyruvaldehyde dimethylacetal to MG (pyruvaldehyde). This hydrolysis method, which does not require a quantitation analysis, serves as a convenient procedure for the preparation of MG for use in glyoxalase I assays.