Toxicological and epidemiologic evidence has suggested that exposure to disinfection by-products (DBPs) impairs semen quality, while the underlying biological mechanisms remain unclear. This study aimed to examine the mediating role of oxidative stress in association between DBP exposure and semen quality. We measured a urinary biomarker of DBP exposure [trichloroacetic acid (TCAA)] and three urinary biomarkers of oxidative stress [8-hydroxy-2-deoxyguanosine (8-OHdG), 8-iso-prostaglandin F2(alpha) (8-isoPGF2(alpha)) and 4-hydroxy-2-nonenal-mercapturic acid (HNE-MA)] among men from an infertility clinic (n = 299). The associations of oxidative stress biomarkers with urinary TCAA and semen quality were evaluated using multivariable linear regression models, and the mediating role of oxidative stress biomarkers was assessed by a mediation analysis. Urinary TCAA was positively associated with urinary 8-OHdG and 8-isoPGF2(alpha) in a dose-response manner (both P for trend < 0.001). Significantly inverse dose-response associations were observed between urinary 8-isoPGF2(alpha) and sperm concentration and between urinary 8-OHdG and sperm motility (both P for trend < 0.05). The mediation analysis indicated a significant indirect effect of urinary 8-isoPGF2(alpha) in the association between urinary TCAA and decreased sperm concentration (P = 0.01). Our results suggest that lipid peroxidation may be an intermediate mechanism by which DBP exposure impairs semen quality. (C) 2020 Elsevier Ltd. All rights reserved.