Background: Oxidative stress caused by free radicals can cause reproductive disorders. Considering the low cytoplasm and limited activity of antioxidant defense in sperm cells, researchers are looking for suitable antioxidant compounds to protect sperm. N-Acetylcysteine (NAC) is one of the antioxidant compounds that can remove free radicals and protect sperm. In numerous studies, NAC with antioxidant properties has been studied to treat infertile men, but its mechanisms on normal sperm cells have not been fully evaluated. Therefore, it is necessary to understand the direct mechanism of NAC's effect on sperm cells before considering its use in clinical conditions. Objective: Considering that NAC easily passes through the cell membrane and can quickly increase the amount of intracellular glutathione using intracellular enzymes, therefore, our objective was to investigate the effect of NAC on motility, viability, and mitochondrial membrane potential (MMP) in vitro. Materials and Methods: Normal samples of fertile men were collected based on WHO criteria. Then, each sample was divided into 3 groups: fresh, control (untreated after 1 hr of incubation), and NAC (treated with 50 µM NAC antioxidant after 1 hr of incubation). Then, the samples were examined with an optical microscope regarding motility. Also, the percentage of viability and MMP were evaluated using eosin-nigrosin and rhodamine 123 stainings, respectively. Results: A significant increase in the mean sperm motility and mitochondrial membrane potential was observed in the NAC group compared to the fresh (p ≤ 0.001) and control (p ≤ 0.01) groups. Also, NAC significantly increased the mean sperm viability compared to the control group (p ≤ 0.01). Conclusion: Based on the results obtained in this study, NAC can protect sperm against oxidative stress damage and improve sperm motility, viability, and MMP.
