Glutathione (GSH) conjugation is important in the detoxification of carcinogens and other exogenous electrophilic drugs and chemicals. The conjugation is catalyzed by GSH S-transferase. Neutrophil GSH content was 12.3 +/- 2.5 (mean +/- SD) nmol/10(7) cells, and transferase activity of cytosol preparations was 0.187 +/- 0.035 nmol/min/mg protein with 1-chloro-2,4-dinitrobenzene (CDNB) as substrate. Maximal activity was found at a pH of 7.0 and a temperature of 40 degrees C. Apparent Km of transferase was 1.25 +/- 0.18 mmol/L CDNB, and apparent Vmax was 0.621 +/- 0.22 nmol/min/mg. GSH-CDNB conjugate was quantitated by HPLC in cells and in medium after CDNB exposure. Transport of conjugate from cells to medium increased with CDNB concentration to 50 mumol/L, and kinetic data showed two saturable transport mechanisms with apparent Km of 5.90 mumol/L and 0.265 mumol/L, respectively. Cellular GSH content fell rapidly with CDNB concentration greater than 2.5 mumol/L, and was depleted in a 10-minute incubation at a concentration of greater than 50 mumol/L CDNB. Neutrophils have a significant content of GSH and a significant amount of transferase activity, and transport of GSH conjugate involves two distinct saturable pathways. GSH depletion can be accomplished with a relatively low concentration of an exogenous chemical and could impair the ability of the cells to carry out their phagocytic functions.