In 1976, it has been reported that phenylglyoxal (C6H5COCHO) selectively inhibits endocytosis in phagocytes of rabbit and mouse. We have tested the specificity of this compound by measuring its effect on human neutrophil chemotaxis, respiration and release of lysosomal enzymes. Pretreatment of human neutrophils with 100 microgram phenylglyoxal/ml for 30 min at 37 degrees C resulted in almost complete inhibition of phagocytosis of opsonized zymosan. However, after treatment with phenylglyoxal, spontaneous mobility as well as chemotaxis of these cells towards casein, rosette formation with opsonized zymosan, stimulation of the oxidative metabolism and release of lysosomal enzymes were also severely decreased. Most of these functions were only partially restored by resuspension of the cells in a medium without phenylglyoxal. The intracellular level of ATP was not affected by phenylglyoxal, but the level of reduced glutathione was decreased. We conclude from the inhibitory action of phenylglyoxal on the stimulated oxygen consumption and its reaction with intracellular glutathione that phenylglyoxal does not necessarily act exclusively on the outside of the plasma membrane. From our studies, it follows that phenylglyoxal is not a specific inhibitor of endo- or exocytosis in human neutrophils. Phenylglyoxal can be used effectively in the bacterial-killing test of phagocytes to inhibit intracellular killing after an initial period of ingestion.