The contribution of the toxicity of glycerol egg yolk citrate (GEYC) cryopreservative medium to the loss of function of human spermatozoa during cryopreservation was determined by investigating the effect of mixing semen with the medium on sperm motility. The percentage of progressively motile spermatozoa, velocity (μm s-1) and lateral head displacement (μm) (mean ± SEM, n = 28) were 55 ± 4.1, 47 ± 2.7, 4.4 ± 0.2, and 32 ± 3.8, 40 ± 2.5, 3.6 ± 0.25 and 15 ± 2.5, 28 ± 1.1, 2.8 ± 0.15 in suspensions of washed spermatozoa prepared from fresh, GEYC-treated and frozen-thawed semen, respectively. The variables changed only slightly after incubation for 3h. The toxicity of GEYC did not vary significantly between samples which survived the complete freeze-thaw cycle well or very poorly. The toxicity of GEYC is responsible for about 50% of the loss of progressively motile spermatozoa during the complete cryopreservation process, but has little effect on the quality of motility. Susceptibility to GEYC does not explain observed differences in the ability of semen samples to survive freezing.