A comparative study on mixed-function oxygenase (MFO) systems was carried out on four echinoderm species: the asteroids Asterias rubens and Marthasterias glacialis, a holothurian Holothuria forskali and an echinoid Echinus esculentus. Cytochromes P-450 and b5 and the MFO system-associated NADH-ferricyanide reductases NADH-cytochrome c reductase, NADPH-cytochrome c reductase and benzo(a)pyrene (BP) hydroxylase activities were present in microsomal fractions of pyloric caeca of A rubens and M. glacialis and in the haemal plexus of H. forskali. In contrast, cytochrome P-450 and BP hydroxylase activity were not detectable in the gonads of E. esculentus. The tissue and subcellular distribution of the MFO system was studied in A. rubens. MFO system components were found in the stomachs and gonads, although detection of cytochrome P-450 in the latter tissue was difficult. Sex-related differences were not significant. The contents of the MFO system components and BP hydroxylase activities were highest in the microsomal (100 000 ×g) fractions, but MFO system components were also found in the mitochondrial (12 000 ×g) and cytosolic fractions. The BP hydroxylase activity in pyloric caeca microsomes of A. rubens was NADPH-dependent and was inhibited by several agents known to be inhibitors of vertebrate cytochromes P-450. In the former respect, the characteristics of the MFO system were more like those of vertebrates and crustaceans than that of molluscs. © 1990 Springer-Verlag.