ABSTRACT:We have identified several novel metabolites of ticlopidine, a well known antiplatelet agent and have revealed its metabolic route in rats. The main biliary metabolite of ticlopidine was characterized as a glutathione (GSH) conjugate of ticlopidine S-oxide, in which conjugation had occurred at carbon 7a in the thienopyridine moiety. Quantitative analysis revealed that 29% of the dose was subjected to the formation of reactive intermediates followed by conjugation with GSH after oral administration of ticlopidine (22 mg/ kg) to rats. In vitro incubation of ticlopidine with rat liver 9000g supernatant fraction (S9) fractions led to the formation of multiple metabolites, including 2-oxo-ticlopidine, the precursor for the pharmacologically active ticlopidine metabolite, [1-(2-chlorobenzyl)-4-mercaptopiperidin-(3Z)-ylidene] acetic acid. A novel thiophene ring-opened metabolite with a thioketone group and a carboxylic acid moiety has also been detected after incubation of 2-oxo-ticlopidine with rat liver microsomes or upon incubation of ticlopidine with rat liver S9 fractions.Ticlopidine is a well known antiplatelet agent (Quinn and Fitzgerald, 1999) and has been widely used for the secondary prevention of atherothrombosis (Jacobson, 2004). The metabolism of ticlopidine is complex because of extensive oxidation in the liver (Saltiel and Ward, 1987). For example, less than 1% of the parent compound was detected in urine, whereas approximately 60 and 25% of radioactivity was recovered in urine after a single oral administration of [ 14 C]ticlopidine to humans (Noble and Goa, 1996) and rats (Tuong et al., 1981), respectively. Furthermore, a large difference was observed between the total radioactivity and unchanged ticlopidine level in the plasma (Panak et al., 1983). In the 1980s, metabolic studies suggested that N-dealkylation, N-oxidation, and oxidation of the thiophene ring followed by ring opening appeared to be the main routes, but numerous highly polar urinary and biliary metabolites in both humans and animals remained unidentified (Tuong et al., 1981;Panak et al., 1983). More recently, novel ticlopidine metabolites such as the S-oxide form (Ha-Duong et al., 2001) and the pharmacologically active metabolite (Yoneda et al., 2004) have been detected. However, the whole metabolism of ticlopidine has remained unclarified.In the present study, we have identified several novel metabolites of ticlopidine and revealed the metabolic pathways of ticlopidine quantitatively. Moreover, an in vitro experiment was conducted to elucidate the whole metabolic route of ticlopidine including the biotransformation to the pharmacologically active metabolite.