The acrosomal extracts from rabbit spermatozoa were more effective in dissolving the zona pellucida of the rabbit ova in the absence than in the presence of Ca2+. The extracts had high proteolytic activity but low esterolytic activity at alkaline pH in which pH effective zona hydrolysis occurred. The results suggest that acrosomal proteinases in addition to acrosin may be involved in the zona penetration.

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Significant release of the acrosomal enzymes arylsulfatase, β‐N‐acetylhexosaminidase and hyaluronidase was observed following the treatment of ejaculated rabbit spermatozoa for 12 hours in 20% rabbit serum for inducing in vitro capacitation, and these sperm were capable of in vivo fertilization; however, the treatment of sperm for 15 minutes in high ionic strength (380 mOsm/kg) or low ionic strength medium (305 mOsm/kg) for in vitro capacitation did not result in any significant release of the above enzymes nor were the sperm capable of in vivo fertilization. Serum‐treated spermatozoa remained significantly motile following the 12 hour treatment, 51% underwent the acrosome reaction and were capable of fertilizing 66% of the ova in vivo. Identical serum treatment of lysosomes from rabbit liver resulted in a comparable release of the lysosomal enzymes. Serum treatment for in vitro capacitation resulted in vesiculation of the anterior margin of half the spermatozoa, but left their inner acrosomal membranes and equatorial segments intact. A biochemical relationship between the release of acrosomal enzymes and capacitation is suggested.