This study has investigated the effect of elevating intracellular calcium levels, using an internal calcium ATPase inhibitor, 2,5-di(tert-butyl) hydroquinone (TBQ), on human sperm function. Isolated sperm samples from five fertile donors were incubated in a capacitating media for up to 6 hr. After 0, 3, and 6 hr incubation, sperm were exposed to a range of TBQ concentrations; 100 microM, 10 microM, and 1 microM, for a fixed incubation period of 5 min. Controls were run for each experiment where sperm were incubated for 5 min in the absence of TBQ. Sperm capacitation and the acrosome reaction were monitored prior to and after exposure to TBQ, using the Chlortetracycline assay. In addition, sperm motility was assessed at each time point and after sperm had been exposed to TBQ. The treatment of sperm with TBQ caused a significant increase in the number of capacitated sperm with an optimum response being achieved in the presence of 100 microM TBQ. However, sperm motility was found not to be effected by the addition of TBQ. The results from the present study suggest that elevating intracellular calcium levels in human sperm by short exposure to a high concentration of TBQ can rapidly accelerate the capacitation process. Furthermore, the observation that TBQ did not elicit a change in sperm motility suggests that TBQ may be highly specific in its mode of action by acting within the head region of human sperm.