Cytochrome P450 (CYP) enzymes are a large and ubiquitous family of hemecontaining proteins found in vertebrates, invertebrates, plants, and microorganisms that catalyze the oxidative biotransformation of diverse lipophilic xenobiotic and endogenous compounds including steroids, fatty acids, drugs, and organic pollutants. CYP enzymes play a role both in the detoxification and activation of halogenated and non-halogenated hydrocarbons found in the food chain. Because CYP enzymes are central to metabolism, levels of individual CYP enzymes are important determinants of bioaccumulation and ultimately of susceptibility to environmental contaminant exposure. A toxicologically relevant aspect of CYP regulation is the observation that several substrates can increase expression of specific CYP enzymes involved in their metabolism. For example, induction of hepatic CYP1A enzymes is a common and characteristic biochemical response in vertebrates of exposure to certain planar halogenated and non-halogenated hydrocarbons, such as polychlorinated dibenzo-p-dioxins (PCDDs), polychlorinated biphenyls (PCBs) and polynuclear aromatic hydrocarbons (PAHs), that often precedes toxicity. In mammals, the CYP1A subfamily contains two related forms, CYP1A1 and CYP1A2, that can be separated by chromatographic, electrophoretic and immunochemical means. Expression of both CYP1A1 and CYP1A2 is regulated in a differential and tissue-specific manner by the aryl hydrocarbon (Ah) receptor. Although several different CYP enzymes have been identified in fish, CYP1A has received the most attention as the major hydrocarbon inducible form and as a possible biomarker of exposure to marine hydrocarbon contaminants. CYP1A enzyme induction in fish populations is generally recognized as a useful and sensitive biochemical marker of contaminant exposure, and by inference, of marine ecosystem health. Usually, induction of CYP1A was determined by measurement of an associated enzyme activity, namely ethoxyresorufin O-deethylase (EROD). The EROD activity is measured by following the increment in fluorescence of the reaction product resorufin. But the optimal conditions for the EROD activity vary from marine fish species. According to ecological features, Sebastiscus marmoratus, Sparus latus and Sparus macrocephalus were selected as experimental animals in the present study. A systematic study had been performed on optimal conditions of hepatic microsomal EROD kinetic reaction system for the control and the induced groups of Sebastiscus marmoratus with the water soluble fraction (WSF) of 0# diesel fuel. The optimal conditions for the microsomal EROD activity of the Sebastiscus marmoratus were pH 7.60, 15℃, 0.5 μmol/L 7-ethoxyresorufin (substrate), 100-800 μg microsomal protein and 70 nmol/mL NADPH, which showed no difference between the control and the induced groups. However, the EROD activity was apparently higher in the induced groups than the control ones. The optimal conditions for the hepatic microsomal EROD activity from Sebastiscus marmoratus were firstly established in this study. Moreover, K_m values for the control and the induced groups were 0.144 μmol/L and 0.357 μmol/L, and V_max values for the two groups were 0.466 nmol min^-1 mg^-1protein and 2.360 nmol min^-1 mg^-1protein, respectively. Furthermore, the comparison of optimal pH values and temperatures (two major factors) for the hepatic microsomal EROD activity among the three species of fish were conducted in the study. The results indicated that optimal pH values and temperatures for the EROD activity of both Sparus latus and Sparus macrocephalus were pH 7.80 and 25℃, which were different from those of Sebastiscus marmoratus.