A high-performance liquid chromatographic method was devised for the qualitative and quantitative analysis of lupine alkaloids in plants. The separation of 22 naturally occurring lupine alkaloids was performed by adsorption chromatography (silica gel) with three solvent systems consisting of diethyl ether, methanol and ammonia solution and reversed-phase chromatography (octadecylsilica) with a buffered aqueous solution containing acetonitrile. The elution of alkaloids was monitored by UV absorption at 220 and 310 nm. Lupine alkaloids containing a 2-pyridone ring were detected by UV absorption at both wavelengths, while the alkaloids that do not possess this unsaturared heterocyclic ring lack UV absorption at 310 nm. The determination of lupine alkaloids was carried out by an external standard method using (-)-cytisine as the standard. The peak area for 1 μg of each alkaloid detected at 220 nm was measured and normalized for that of (-)-cytisine, and these relative factors were used for the determination of the alkaloids. The qualitative and quantitative analysis of lupine alkaloids in plants of the genus Thermopsis was performed by this method. © 1989.