Polychlorinated biphenyls (PCBs) are endocrine-disrupting chemicals that mimic estrogen in fish. Among the various PCBs, 3,3',4,4',5-pentachlorobiphenyl (PCB126), considered an estrogen antagonist, has been reported to elicit estrogenic activity in fish. We investigated the estrogenic potency of PCB126 in in vitro oocyte maturation in dusky tripletooth goby, Tridentiger obscurus. In this study, we quantified steroid metabolites following exposure to PCB126 and estradiol-17 beta (E-2). Vitellogenic ovarian follicles were incubated, in vitro, with 10 and 100 ng/mL PCB126 or E-2 with [H-3]17 alpha-hydroxyprogesterone as a precursor. Testosterone (T) and E-2,E- were identified using thin-layer chromatography, high-performance liquid chromatography, and gas chromatography-mass spectrometry. Both PCB126 and E-2 increased T and E-2 metabolite production. Further, vitellogenic ovarian follicles were exposed to PCB126 (1, 10 and 100 ng/mL) or E-2 (0.01, 0.1 and 1 ng/mL) in vitro, and T and E-2 from the incubation media were measured. PCB126 (100 ng/mL) inhibited T production and increased E-2 production. Furthermore, we investigated the effects of PCB126 on the final oocyte maturation process. Germinal vesicle migration ovarian follicles were in vitro incubated with 0.1, 1, 10 and 100 ng/mL of PCB126 or E-2. High doses of PCB126 (10 and 100 ng/mL) inhibited germinal vesicle breakdown. These results suggest that PCB126 has an estrogenic potency in oocyte maturation in T. obscurus.