OBJECTIVE: To explore the effect of occupational wireless fidelity(Wi-Fi) microwave radiation on testosterone synthesis in male workers. METHODS: A total of 51 male workers exposed to microwave radiation in Wi-Fi test station of a mobile phone manufacturer were selected as exposure group by judgment sampling method. They were divided into <2.0 years subgroup and ≥2.0 years subgroup according to the length of work years. At the same time, 30 male workers who were not exposed to occupational hazards in the same factory were selected as the control group. Serum total cholesterol level was detected by colorimetry. Serum testosterone, cyclic adenosine monophosphate(cAMP), cytochrome P450 17 A1(P450 c17), cytochrome P450 cholesterol side chain cleavage enzyme(P450 scc), levels were detected by enzyme-linked immunosorbent assay. The relative expression of P450 scc and P450 c17 mRNA in whole blood was measured by real-time quantitative polymerase chain reaction. RESULTS: The levels of serum testosterone, P450 c17 and the relative expression of P450 c17 mRNA in workers of the exposure group were lower than that in the control group(P≤0.05), and the above indexes in the sub-exposure group with work age ≥2.0 years was lower than that in the control group(P<0.05). There was no significant difference in levels of serum total cholesterol, cAMP, P450 scc and relative expression of P450 scc in whole blood among the exposed group,two subgroups and the control group(P>0.05). CONCLUSION: Long-term exposure to Wi-Fi microwave radiation can inhibit the expression of P450 c17 mRNA and the synthesis of P450 c17 protein, both are key enzymes for testosterone synthesis in male workers, thereby affecting the synthesis and secretion of testosterone.