A cross-sectional study was conducted to assess whether urinary 1-hydroxypyrene (1-OHP) could serve as a biomarker to assess the effect of PAHs on cellular and molecular changes of sperm. Urine and semen samples were collected from a total of 65 healthy coke oven workers. Sperm quality parameters (concentration, motility, vitality, and morphology) and semen integrity (DNA fragmentation, 8-oxodGuo, bulky DNA adducts) were analyzed. Sixteen (16) targeted PAHs at the personal breathing zone area were monitored and quantified. Results showed that urinary 1-OHP positively correlated with measured levels of 16 targeted PAHs. Urinary 1-OHP did not significantly correlate with semen quality; however, PAHs with heavy molecular weight, e.g., benzo(g,h,i)perylene and benzo(k)fluoranthene, negatively correlated with morphology and motility of sperms (p = 0.02 and 0.002, p = 0.04 and 0.04, respectively). Urinary 1-OHP positively correlated with the level of 8-oxodGuo and bulky DNA adducts, but not DNA fragmentation. Urinary 1-OHP was a suitable biomarker for an estimate of biologically effective doses of PAH exposure. However, urinary 1-OHP may not be sufficient as a biomarker to assess both cellular and molecular changes of sperm induced by PAHs.