The present study was undertaken to elucidate the mechanism of spermicidal action of NIM-76, a fraction isolated from neem oil. The spermicidal activity of NIM-76 was confirmed using a fluorescent staining technique. NIM-76 was found to affect the motility of the sperm in a dose-dependent manner. Supplementation of pentoxifylline, which is known to enhance the motility of the sperm, could not prevent the spermicidal action of NIM-76. There was a gradual leakage of cytosolic LDH from the sperm in the presence of NIM-76. Electron microscopic studies revealed the formation of pores and vesicles over the sperm head, indicating the damage to the cell membrane. Membrane fluidization studies did not reveal any significant change in the fluidity of sperm cell membrane structure.
PIP: Neem oil, an extract of a native plant of India, has been demonstrated to have anti-fertility, anti-implantation, and abortifacient properties. An active fraction, termed NIM-76, was extracted that eliminates its abortifacient properties while retaining spermicidal activity. This fraction kills all human sperm in vitro in under 20 seconds at a concentration of 25 mg/ml. With increases in NIM-76 concentrations from 10 to 1000 mcg/ml, there was a linear decrease in percentages of motile as well as progressively motile sperm with time; also recorded were decreases in percentages of rapid, medium, and slow moving sperm, mean track speed, progressive velocity, mean linearity, and lateral head displacement and an increase in the percentage of static sperm. Electron microscopy revealed the formation of pores and vesicles over the sperm head, indicating damage to the cell membrane. Membrane fluidization studies did not reveal any significant change in the fluidity of sperm cell membrane structure. Since calcium supplementation did not relieve the sperm from the spermicidal action, it was determined that NIM-76 does not cause any depletion of intracellular calcium. The capability of NIM-76 to selectively kill sperm without affecting normal cells makes it a highly desirable potential vaginal contraceptive agent.