[Objective] To investigate the effect of the secretory proteins of the ventral prostate glands on the sperm membrane proteins in golden hamsters. [Methods] The sperm was collected from female hamsters uteri after mated with the males with or without ventral prostate gland. Male golden hamsters were divided into four experimental groups: (i) all accessory sex glands (ASG) removed; (ii) ventral prostate gland removed; (iii) all ASG removed except ventral prostate gland and (iv) sham-operated controls. Each group contained 6 hamsters. The sperm membrane proteins were extracted from uterine sperm and analyzed by SDS-PAGE and two dimension electrophoresis. [Results] The SDS-PAGE results of sperm membrane showed that the ventral prostate glands added up some proteins or increased the quantity of some proteins, which contained molecular mass (MM)15 k, 29 k, 38 k, 55 k, and 91 k proteins, to the sperm membrane. 2D-electrophoresis of sperm membrane showed that some extra protein spots were appeared in with ventral prostate gland groups, their MM and isoelectric point (IP) corresponding to 16 k/8.60, 16.6 k/9.20, 28 k/5.88, 28 k/6.10, 29 k/5.98, 32 k/6.35, 32 k/6.50, 32 k/7.20, 61 k/5.90,and 83 k/6.40, respectively. The quantity of some protein spots increased in sperm membrane of ventral prostate glands group, their MM/IP coresponding to 17 k/5.95, 17.5 k/6.50, 24 k/7.20, 26 k/5.40, 26 k/5.60, 27 k/7.20, 27 k/7.50, 28 k/5.70, 29 k/8.50, 42 k/6.50, and 42 k/7.00, respectively. [Conclusion] The ventral prostate gland secretion plays a role in regulating the sperm membrane proteins and the latter may in turn affect the male fertility or embryo development.