Esterase 6, a carboxylesterase (EC 3.1.1.1.) of Drosophila melanogaster, has been unambiguously shown to be synthesized in the anterior ejaculatory duct of the reproductive tract of adult males. Radiolabelled esterase 6 was immunoprecipitated from homogenates prepared from ducts cultured for 5 hr in an in vitro system containing [ 3H]leucine. The anti-allatotropin, precocene, was used to treat adult males to determine if juvenile hormone levels affect the activity of esterase 6. In treated males the level of esterase 6 was 37% of the control levels. This system may allow for further exploration of the hormonal regulation of reproductive processes in Drosophila.