Vol. 30, Issue 9, 1005-1012, September 2002

Glucuronidation of Dihydroartemisinin in Vivo and by Human Liver Microsomes and Expressed UDP-Glucuronosyltransferases

Kenneth F. Ilett, Brian T. Ethell, James L. Maggs, Timothy M. E. Davis, Kevin T. Batty, Brian Burchell, Tran Quang Binh, Le Thi Anh Thu, Nguyen Canh Hung, Munir Pirmohamed, B. Kevin Park, and Geoffrey Edwards

Department of Pharmacology, University of Western Australia, Crawley, Western Australia (K.F.I.); Clinical Pharmacology and Toxicology Laboratory, Western Australian Centre for Pathology and Medical Research, Nedlands, Western Australia (K.F.I.); Department of Medicine, University of Western Australia, Fremantle Hospital, Fremantle, Western Australia (T.M.E.D.); School of Pharmacy, Curtin University, Perth, Western Australia (K.T.B.); Department of Biochemical Medicine, University of Dundee, Ninewells Hospital, Dundee, United Kingdom (B.B., B.E.); Tropical Diseases Research Centre, Cho Ray Hospital, Ho Chi Minh City, Vietnam (T.Q.B., L.T.A.T.); Bao Loc Hospital, Lam Dong Province, Vietnam (N.C.H.); Department of Pharmacology and Therapeutics, the University of Liverpool, Liverpool, United Kingdom (M.P., J.L.M., B.K.P., G.E.); and Division of Parasite and Vector Biology, Liverpool School of Tropical Medicine, Liverpool (G.E.)

The aim of this study was to elucidate the metabolic pathways for dihydroartemisinin (DHA), the active metabolite of the artemisinin derivative artesunate (ARTS). Urine was collected from 17 Vietnamese adults with falciparum malaria who had received 120 mg of ARTS i.v., and metabolites were analyzed by high-performance liquid chromatography-mass spectrometry (HPLC-MS). Human liver microsomes were incubated with [12-³H]DHA and cofactors for either glucuronidation or cytochrome P450-catalyzed oxidation. Human liver cytosol was incubated with cofactor for sulfation. Metabolites were detected by HPLC-MS and/or HPLC with radiochemical detection. Metabolism of DHA by recombinant human UDP-glucuronosyltransferases (UGTs) was studied. HPLC-MS analysis of urine identified -DHA--glucuronide (-DHA-G) and a product characterized as the tetrahydrofuran isomer of -DHA-G. DHA was present only in very small amounts. The ratio of the tetrahydrofuran isomer, -DHA-G, was highly variable (median 0.75; range 0.09-64). Nevertheless, -DHA-G was generally the major urinary product of DHA glucuronidation in patients. The tetrahydrofuran isomer appeared to be at least partly a product of nonenzymic reactions occurring in urine and was readily formed from -DHA-G by iron-mediated isomerization. In human liver microsomal incubations, DHA-G (diastereomer unspecified) was the only metabolite found (V_max 177 ± 47 pmol min¹ mg¹, K_m 90 ± 16 µM). -DHA-G was formed in incubations of DHA with expressed UGT1A9 (K_m 32 µM, V_max 8.9 pmol min¹ mg¹) or UGT2B7 (K_m 438 µM, V_max 10.9 pmol mg¹ min¹) but not with UGT1A1 or UGT1A6. There was no significant metabolism of DHA by cytochrome-P450 oxidation or by cytosolic sulfotransferases. We conclude that -DHA-G is an important metabolite of DHA in humans and that its formation is catalyzed by UGT1A9 and UGT2B7.