Blockage of Fertilization in Rana pipiens by Trypsin Inhibitors

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Abstract
Fertilization of Rana pipiens ova was blocked when spermatozoa were incubated with either naturally-occurring or synthetic trypsin inhibitors immediately prior to insemination. One synthetic inhibitor, 1-chloro-3-tosylamido-7-amino-2-heptanone hydrochloride (TLCK), had an ED50, of approximately 0.275 mg/ml. Washing TLCK-treated spermatozoa prior to insemination did not eliminate the block to fertilization. At effective doses, TLCK-treated spermatozoa had normal motility. These findings suggest that, in R. pipiens, sperm enzymes similar to trypsin are necessary for fertilization.