Crystalline ovomucoid and soybean trypsin inhibitors were used to inhibit in vitro fertilization of rabbit ova by capacitated rabbit sperm. Cleavage was determined in 222 control ova and in 244 ova treated with an inhibitor. The data demonstrate that there is good correlation between the percentage inhibition of the trypsin-like enzyme of the rabbit sperm acrosome by the inhibitor and the percentage inhibition of in vitro fertilization by the inhibitor. These results provide additional evidence that the trypsin-like component of the rabbit sperm acrosome is essential for dissolution and penetration of the zona pellucida by the spermatozoon.