Concentrations of vitellogenin (VTG) in serum of wild male flounder (Pleuronectes yokohamae) were investigated, as a biomarker for environmental estrogens in marine waters, from January 1997 to May 1998 in Tokyo Bay, Japan (n = 130) and from a reference site in Hokkaido, Japan, far from urban areas (about 40 km from Hakodate city) from March 1997 to June 1998 (n = 62). Statistically higher concentrations of serum VTG were detected in wild male flounder collected in Tokyo Bay (range: 25-2200 ng ml-1) compared to fish from Hokkaido (range: 31-91 ng ml-1) throughout the entire sampling period. Three out of 20 males collected off Haneda, in the inner part of Tokyo Bay, showed testes containing small numbers of oocytes. This intersex condition was not found among males collected from the reference site. This study suggests that the abnormally high levels of VTG and the presence of oocytes in the testis of male flounder collected from Tokyo Bay could be the effect of environmental estrogens.