Organic anthropogenic compounds such as polycyclic aromatic hydrocarbons, polychlorinated biphenyls, nitroaromatics, dioxins, various pesticides and natural compounds can enter the aquatic environment. These lipophilic compounds are readily taken up into the tissues of aquatic organisms where biotransformation via Phase I and Phase II metabolism can in part, determine the fate and toxicity of the xenobiotics. In this study on gilthead seabream (Sparus aurata), liver microsomal ethoxyresorufin O-deethylase (EROD), methoxyresorufin O-demethylase (MROD), penthyloxyresorufin O-depenthylase (PROD), benzyloxyresorufin O-debenzylase (BROD), aniline 4-hydroxylase (A4H), N-nitrosodimethylamine N-demethylase (NDMA-DE), aminopyrine N-demethylase (APND), caffeine N-demethylase (CN3D) and erythromycin N-demethylase (ERND) were determined. A4H, ERND, NDMA-ND, EROD and PROD activities increased in gilthead seabream (Sparus aurata) liver microsomes of different ages, while CN3D and APND activities decreased. MROD activities were barely detectable in mature fish whereas BROD activities were not detectable at all ages studied. Results were also recorded at the protein level by Western blotting using anti-CYP1A, CYP2E and CYP3A antibodies. In conclusion, elevated levels of gilthead seabream liver microsomal CYP450 enzyme activities might reveal possible exposure to various exogenous compounds, which might affect the desired responses to drugs, hormones and dietary supplements used during breeding.