The exposure and uptake of environmental estrogenic compounds have been reported in previouss studies of demersal flatfish species in the central Southern California Bight (SCB), USA. The objective of this study was to evaluate the estrogenic or feminizing activity of marine sediments from the SCB by using in vivo vitellogenin (VTG) assays in male or juvenile fish. In 2003, sediments were collected near wastewater outfalls serving the counties of Los Angeles (LACSD) and Orange (OCSD), and the city of San Diego (SD), California, USA. Cultured male California halibut (CH; Paralichthys californicus) were either directly exposed to sediments for 7 d or treated with two intraperitoneal injections of sediment extract over 7 d. The 17β-estradiol (E2) equivalent values ranged from 1 to 90 μg/kg with LACSD > SD > OCSD. Measurable concentrations of E2 were observed in all sediment extracts and ranged from 0.16 to 0.45 ng/g. Estrone (E1) was only observed in sediments near the LACSD outfall (0.6 ng/g). Alkylphenols and alkylphenol ethoxylates were observed in all sediment samples, but were highest near the OCSD outfall, where concentrations of nonylphenol were 3,200 ng/g. Fractionation studies of the LACSD sediment extract collected in 2004 failed to demonstrate relationships between VTG expression and 62 analytes, including E2, which was observed in the whole extract (2.9 ng/g). Oxybenzone (1.6 ng/g) was identified in bioactive fractions as well as unknown compounds of relatively high polarity. These results indicate that estrogen receptor-based assays may underestimate environmental estrogenic activity and estrogenic compounds other than classic natural and xenoestrogens may contribute to estrogenic activity of sediments from the SCB.