The antifungal properties of 515 synthetic and semi-synthetic protoberberines were investigated. HWY-289 was chosen for further study because it exhibited the most significant anti-Candida activity (MICs were 1.56 mg/L for Candida albicans and Candida krusei; 6.25 mg/L for Candida guilliermondii) but did not demonstrate toxicity in rats. HWY-289 inhibited the incorporation of L-[methyl-(14)C]methionine into the C-24 of ergosterol in whole cells of C. albicans (IC(50) 20 microM). However, HWY-289 (100 microM) had no effect on mammalian cholesterol biosynthesis in rat microsomes while miconazole (100 microM) was a potent inhibitor of cholesterol biosynthesis under identical assay conditions. A second major target site for HWY-289 was identified that involves cell wall biosynthesis in C. albicans. HWY-289 was a potent inhibitor of the chitin synthase isozymes CaCHS1 and CaCHS2, with IC(50) values of 22 microM for each enzyme. The effect was highly specific in that HWY-289 had no significant effect on C. albicans CaCHS3 (IC(50) > 200 microM). Thus, HWY-289 compared favourably with well-established antifungal agents as an inhibitor of the growth of Candida species in vitro, and may have considerable potential as a new class of antifungal agent that lacks toxic side effects in the human host.