Synergistic effect of gramicidin and EDTA in inhibiting sperm motility and cervical mucus penetration in vitro

Gramicidin, a linear polypeptide with antiviral and antimicrobial properties, was compared in vitro with a commonly used spermicidal detergent—nonoxynol-9 (N9). The inhibition of sperm functions was evaluated by computer-assisted semen analysis (CASA) for sperm motility, in cervical mucus penetration assay, and by colorimetric tetrazolium salt and lactate dehydrogenase release assays routinely employed for testing the toxicity of drugs. The effective 100% inhibitory concentration (IC₁₀₀) of gramicidin in a 2-min sperm immobilization assay by CASA was equal to 4 μg/ml, whereas IC₁₀₀ of N9 was equal to 200 μg/ml. The presence of 0.1% of chelating agent, EDTA, reduced IC₁₀₀ of gramicidin to 10 ng/ml, while less than a twofold enhancement in N9 activity was observed upon combination with EDTA. Likewise, the gramicidin/EDTA combination was 100,000 times more potent than N9/EDTA in the sperm penetration assay. Quantitative toxicity tests confirmed that gramicidin is a potent spermostatic rather than spermicidal agent. Further development of a gramicidin/EDTA formulation is warranted as a nontoxic topical contraceptive with activity against viral and microbial sexually transmitted diseases (STDs).