Abstract: AIMTo study the metabolites of penehyclidine hydrochloride (PH) raceme, a new anti-cholinerigic drug invented by the Institute of Pharmacology and Toxicology, Academy of Military Medical Sciences. METHODS Three healthy rat urine samples were collected within 24 h after a single im dose of PH raceme and PH-d₅ ［(5+5) mg·kg^-1］ simultaneously. The eight metabolites of PH raceme were identified by the methods of LC-MS/MS, GC-MS, FAB-MS and the stable isotope ion cluster. Mass spectrometry was operated in the positive mode for the method of LC-MS/MS. RESULTSM1 and M1^* were identified as the oxygenated products of PH in the cyclopentyl group; M2 and M2^* were as the hydroxylated products of PH in the cyclopentyl group; M3 and M3^* were as the oxygented and hydroxylated products of PH at the meta-position of cyclopentyl group; M4 and M4^* were identified as the dihydroxylated metabolites of PH, the hydroxylated position were at the cyclopentyl group and quiniuclidinol ring of PH. Among them, M1 and M1^*, M2 and M2^*, M3 and M3^*, M4 and M4^* were the isomers of eath other. CONCLUSIONThese characteristics can be used for future structure elucidation in studies of the metabolites of PH optical isomers. The structure data of PH metabolites provide important information for the clinical use and for developing better anti-cholinerigic drug.