Fertility of men depends on the quality of semen. The aim of the present paper is to determine both the acrosin activity by radioimmunoassay and ATP concentration by bioluminescence in human spermatozoa, and evaluate these results in those samples with normal or low sperm penetration according to SPA test. Ejaculates obtained from 42 untreated men, were studied one hour after the obtention. These materials were divided into two groups:20 human semen samples with "in vitro" potentiality to penetrate zone-free hamster ova, between 15% to 98% and 22 human semen with SPA test between 0% to 14%. When we compare the group with normal penetration response vs that group with low or absent penetration ones, a significant decrease of ATP and acrosin concentrations was observed (P less than 0.001). Nevertheless no significant difference was observed in relation with percentage of motility, volume (ml), sperm concentration (10(6)/ml), percent of quick progressive spermatozoa and number of gametes capable of migrating into the medium layer (10(6)), between the group with low or absent penetration test against that one with normal zona-free hamster egg test.