A colorimetric assay of glutathione S-transferases using o-dinitrobenzene as a substrate

Conditions have been examined for the use of o-dinitrobenzene as a substrate for colorimetric assay of glutathione S-transferases. Activities can be determined by measuring nitrite released enzymatically from the substrate using a diazo-coupling method with N-(1-naphthyl)ethylenediamine dihydrochloride and sulfanilamide. The assay can be done in the presence of large amounts of reduced glutathione (GSH), cysteine, and protein, and is capable of quantitating the enzyme activity in about 30 micrograms (wet weight) of monkey liver, corresponding to 0.1 microgram of purified glutathione S-transferase from the same source. This method is suitable for assaying simultaneously a large number of samples with reasonable sensitivity and speed.