title: Enzymatic synthesis of substituted epicatechins for bioactivity studies in neurological disorders.
abstract: Glucuronidated and/or methylated metabolites of the proanthocyanidin (PA) monomer (-)-epicatechin are detected in both blood and brain following feeding of rodents with a monomeric grape seed PA extract shown to reduce symptoms in a mouse model of Alzheimer's disease. To generate metabolites for future mechanistic studies, we investigated the ability of recombinant human glucuronosyl transferases of the UGT1A and UGT2B families to glucuronidate epicatechin or 3'-O-methyl epicatechin in vitro. Of twelve enzymes tested, UGT1A9 was the most efficient, producing epicatechin 3'-O-glucuronide as the major product. Incubation of UGT1A9 with 3'-O-methyl-epicatechin resulted in two major products, one of which was identified as 3'-O-methyl-epicatechin 5-O-glucuronide, a major metabolite found in blood plasma and brain tissue of the rodents following feeding with a grape seed extract. We also investigated in vitro methylation of epicatechin and epicatechin glucuronides by human catechol O-methyltransferase. Enzymatic production of 3'-O-methyl-epicatechin 5-O-glucuronide was optimized to 50% overall yield. These studies form a basis for generation of mg quantities of pure epicatechin (methyl) glucuronides of biological significance, and provide clarification of structure of previously identified epicatechin metabolites.
authors.affiliation: Plant Biology Division, Samuel Roberts Noble Foundation, Ardmore, OK 73401, USA.
authors.forename: Jack W
authors.initials: JW
authors.lastname: Blount
authors.forename: Mario
authors.initials: M
authors.lastname: Ferruzzi
authors.forename: Dan
authors.initials: D
authors.lastname: Raftery
authors.forename: Giulio M
authors.initials: GM
authors.lastname: Pasinetti
authors.forename: Richard A
authors.initials: RA
authors.lastname: Dixon
mesh_terms: D000544:Alzheimer Disease / Q000097:blood / Q000378:metabolism; D000818:Animals; D002392:Catechin / Q000031:analogs & derivatives* / Q000096:biosynthesis / Q000737:chemistry / Q000302:isolation & purification; D002851:Chromatography, High Pressure Liquid; D005965:Glucuronates / Q000096:biosynthesis* / Q000737:chemistry / Q000302:isolation & purification; D014453:Glucuronosyltransferase / Q000737:chemistry*; D006801:Humans; D051379:Mice; D044945:Proanthocyanidins / Q000096:biosynthesis* / Q000097:blood / Q000737:chemistry; D011994:Recombinant Proteins / Q000737:chemistry*; D000090264:UDP-Glucuronosyltransferase 1A9; D000099148:UGT1A1 Enzyme
chemical_list: D002392:Catechin; D005965:Glucuronates; D014453:Glucuronosyltransferase; D044945:Proanthocyanidins; D011994:Recombinant Proteins; D000090264:UDP-Glucuronosyltransferase 1A9; D000099148:UGT1A1 Enzyme; C000715591:UGT1A9 protein, human; C000715592:Ugt1a9 protein, mouse; C480662:epicatechin-3'-O-glucuronide; C013221:proanthocyanidin