The lipid class, fatty acid, and sterol composition of eight species of ichthyotoxic marine gymnodinioid dinoflagellate (Karenia, Karlodinium, and Takayama) species was examined. The major lipid class in all species was phospholipid (78%-95%), with low levels of triacylglycerol (TAG; 0%-16%) and free fatty acid (FFA; 1%-11%). The common dinoflagellate polyunsaturated fatty acids (PUFA), octadecapentaenoic acid (OPA 18:5x3), and docosahexaenoic acid (DHA 22:6x3), were present in all species in varying amounts (14%-35% and 8%-23%, respectively). The verylong-chain PUFA (VLC-PUFA) 28:7x6 and 28:8x3 were present at low levels (<1%), and the ratio of these fatty acids may be a useful chemotaxonomic marker at the species level. The typical dinoflagellate sterol dinosterol was absent from all species tested. A predominance of the 4-methyl and 4-desmethyl D 8 (14) sterols in all dinoflagellate species included 23-methyl-27-norergosta-8(14),22-dien-3b-ol (Karenia papilionacea A. J. Haywood et Steid, 59%-66%); 27-nor-(24R)-4a-methyl-5a-ergosta-8(14),22-dien-3b-ol, brevesterol, (Takayama tasmanica de Salas, Bolch et Hallegraeff 84%, Takayama helix de Salas, Bolch, Botes et Hallegraeff 71%, Karenia brevis (C. C. Davis) G. Hansen et Moestrup 45%, Karlodinium KDSB01 40%, Karenia mikimotoi (Miyake et Kominami ex Oda) G. Hansen et Moestrup 38%); and (24R)-4a-methyl-5a-ergosta-8(14),22-dien-3b-ol, gymnodinosterol, (K. mikimotoi 48%, Karenia umbella de Salas, Bolch et Hallegraeff 59%, Karlodinium veneficum (D. L. Ballant.) J.Larsen 71%-83%). In Takayama species, five steroid ketones were identified, including for the first time the 3-keto form of brevesterol and gymnodinosterol. These results indicate a biochemical link between sterol and steroid ketone biosynthesis, suggesting that selected dinoflagellates can make a significant contribution to ketones in marine sediments. The presence of steroid ketones, specific sterols, and fatty acids, and the ratio of VLC-PUFA may prove to be a useful chemotaxonomic tool for distinguishing between morphologically similar species. The relative levels of the PUFA, OPA, and DHA, coupled with the potential inhibitory action of D 8(14) sterols, may provide an insight into the ichthyotoxicity of these bloom-forming dinoflagellates.