title: In Vitro Evaluation of the Effect of 7-Methyl Substitution on Glucuronidation of Daphnetin: Metabolic Stability, Isoform Selectivity, and Bioactivity Analysis.
abstract: The C-8 phenol group is essential to exert the bioactivities of daphnetin, but it is readily conjugated with glucuronic acid prior to excretion. In this study, daphnetin-7-methylether (7M-DNP) was used to investigate the effect of 7-methyl substitution on daphnetin glucuronidation in human/rat liver (HLM/RLM) and intestine (HIM/RIM) microsomes, and recombinant UDP-glucuronosyltransferases (UGTs). Compared with daphnetin, the Vmax /Km values of 7M-DNP via 8-O-glucuronidation were 2.1-fold lower in HLM, 1.7-fold lower in HIM, and 2.4-fold lower in RLM, suggesting an improvement in metabolic stability. Different from daphnetin 8-O-glucuronidation exclusively catalyzed by UGT1A6 and UGT1A9, UGT1A1, -1A3, -1A7, -1A8, and -1A9 showed glucuronidation activity toward 7M-DNP. Kinetics studies, chemical inhibition, and the relative activity factor approach were used to demonstrate that UGT1A9 was mainly responsible for the reaction in HLM, whereas UGT1A1 was a primary contributor in HIM. The Vmax /Km values of 7M-DNP glucuronidation in HLM and HIM were 0.61-0.74-fold lower than those of rat, suggesting the differences between the two species. The bioactivity analysis demonstrated that 7M-DNP had an anti-inflammatory activity comparable to that of daphnetin. These findings indicated that the outcomes of 7-methyl substitution on daphnetin might be positive, but this should be confirmed in future in vivo studies.
authors.affiliation: Second Affiliated Hospital of Dalian Medical University, Dalian, China.|Laboratory of Pharmaceutical Resource Discovery, Dalian Institute of Chemical Physics, Chinese Academy of Sciences, Dalian, China.|University of Chinese Academy of Sciences, Beijing, China.
authors.forename: Si-Cheng
authors.initials: SC
authors.lastname: Liang
authors.affiliation: Laboratory of Pharmaceutical Resource Discovery, Dalian Institute of Chemical Physics, Chinese Academy of Sciences, Dalian, China.
authors.forename: Guang-Bo
authors.initials: GB
authors.lastname: Ge
authors.affiliation: Laboratory of Pharmaceutical Resource Discovery, Dalian Institute of Chemical Physics, Chinese Academy of Sciences, Dalian, China.|University of Chinese Academy of Sciences, Beijing, China.
authors.forename: Yang-Liu
authors.initials: YL
authors.lastname: Xia
authors.affiliation: Laboratory of Pharmaceutical Resource Discovery, Dalian Institute of Chemical Physics, Chinese Academy of Sciences, Dalian, China.
authors.forename: Jiang-Wei
authors.initials: JW
authors.lastname: Zhang
authors.affiliation: Second Affiliated Hospital of Dalian Medical University, Dalian, China.
authors.forename: Xiao-Yi
authors.initials: XY
authors.lastname: Qi
authors.affiliation: Second Affiliated Hospital of Dalian Medical University, Dalian, China.
authors.forename: Cai-Xia
authors.initials: CX
authors.lastname: Tu
authors.affiliation: Laboratory of Pharmaceutical Resource Discovery, Dalian Institute of Chemical Physics, Chinese Academy of Sciences, Dalian, China.
authors.forename: Ling
authors.initials: L
authors.lastname: Yang
mesh_terms: D000293:Adolescent; D000328:Adult; D000368:Aged; D000818:Animals; D005260:Female; D020719:Glucuronides; D014453:Glucuronosyltransferase / Q000037:antagonists & inhibitors / Q000378:metabolism; D006801:Humans; D066298:In Vitro Techniques; D007413:Intestinal Mucosa / Q000378:metabolism; D007527:Isoenzymes / Q000378:metabolism; D007536:Isomerism; D007700:Kinetics; D008297:Male; D008745:Methylation; D008862:Microsomes, Liver / Q000378:metabolism; D008875:Middle Aged; D051381:Rats; D013045:Species Specificity; D014468:Umbelliferones / Q000494:pharmacology*; D055815:Young Adult
chemical_list: D020719:Glucuronides; D007527:Isoenzymes; D014468:Umbelliferones; D014453:Glucuronosyltransferase; C039952:daphnetin
keywords: daphnetin; enzyme kinetics; glucuronidation; glucuronosyltransferases; human liver microsomes; metabolic clearance; metabolic stability; methyl substitution; phase II enzymes; structural modification