Endocrine disrupting activities of three isomers of monohydroxylated 1-nitropyrene (1-NP) [3-, 6-, and 8-hydroxy-1-nitropyrenes (OHNPs)] were evaluated for the first time by yeast two-hybrid assay. OHNPs, which are not only metabolites of 1-NP but are also found in airborne particles, did not exhibit androgenic activity but exhibited estrogenic, antiestrogenic, and antiandrogenic activities. 6-OHNP showed the strongest estrogenic activity among the three OHNP isomers examined in this study. Concentrations of the OHNP isomers that gave 10% of activity of 1.0×10^-6M 17β-estradiol (E₂) were as follows: 3-OHNP, 6.0×10^-7M; 6-OHNP, 6.0×10^-8M; 8-OHNP, 9.0×10^-7M. On the contrary, 8-OHNP exhibited the strongest antiestrogenic and antiandrogenic activities of the three isomers. 1.0×10^-6M of 8-OHNP inhibited 32 and 90% of β-galactosidase activity induced by 1.0×10^-9M of E₂ and 1.0×10^-8M of 5α-dihydrotestosterone (DHT), respectively. These findings point out the necessity for detailed investigation of environmental sources and distributions of OHNPs as well as the parent 1-NP.