Abstract
The dietary polyphenol resveratrol (RES) exists as cis- and trans-isomers with known stereospecific and stereoselective glucuronidation at the 3 and 4′ positions by distinct UGT1A isoforms. We examined cis-RES glucuronidation in various protein sources. UGT1A6 or UGT1A1 genotype-dependent cis-or trans-RES glucuronidation, respectively, was further determined. cis-RES exhibited partial substrate inhibition in UGT1A6 Supersomes and human embryonic kidney 293 cells overexpressing genetically variant UGT1A6 alleles. Cells expressing UGT1A6*4 had the highest activity with a Vmax of 612 ± 27.36 nmol/min/mg, followed by UGT1A6*3. The *2 allozyme had a higher Vmax (1.6-fold) and Km (1.9-fold) than *1. In 51 human liver samples genotyped for UGT1A6, four alleles (frequencies) were identified as *1 (0.58), *2 (0.36), *3 (0.01), and *4 (0.05), leading to assignment of the following genotypes (frequencies): *1/*1 (0.29), *1/*2 (0.45), *1/*3 (0.02), *1/*4 (0.10), and *2/*2 (0.14). Up to 5-fold variability in trans-RES glucuronidation was observed in individual liver samples. In livers stratified by UGT1A6 genotype, a significant difference in cis-RES glucuronidation activity (p < 0.05) was seen between the *2 variants compared with homozygous *1 livers. The trans-RES glucuronidation was quantitated in a human liver bank genotyped for the UGT1A1 TATA box repeat polymorphism. There was no significant difference for formation of trans-RES 3-O-glucuronide. We were surprised to find that trans-RES 4′-O-glucuronide formation was higher in livers with the 7/7 genotype compared with 6/6 and 6/7 (p < 0.05). In conclusion, cis-RES glucuronidation exhibited atypical partial substrate inhibition kinetics in vitro. Whereas cis-RES glucuronidation varied with UGT1A6 genotypes, the UGT1A1*28 polymorphism did not explain variability in trans-RES glucuronidation.
Footnotes
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Part of this work was accepted for poster presentation as follows: Iwuchukwu OF, Ajetunmobi J, Ung D, and Nagar S (2008) Characterizing the effects of common polymorphisms in UGT1A1 and UGT1A6 on the glucuronidation of cis and trans resveratrol. Annual Meeting of the American Association of Pharmaceutical Scientists; 2008 Nov 16–20; Atlanta, GA. American Association of Pharmaceutical Scientists, Arlington, VA.
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J.A. and D.U. contributed equally to this work.
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Article, publication date, and citation information can be found at http://dmd.aspetjournals.org.
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doi:10.1124/dmd.109.027391.
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ABBREVIATIONS: RES, resveratrol; UGT, UDP glucuronosyltransferase; R3G, trans-resveratrol 3-O-glucuronide; R4′G, trans-resveratrol 4′-O glucuronide; cis-R3G, cis-resveratrol 3-O-glucuronide; SNP, single nucleotide polymorphism; DMSO, dimethyl sulfoxide; PCR, polymerase chain reaction; HEK, human embryonic kidney; TBST, Tris-buffered saline/Tween 20; HLM, human liver microsome; HPLC, high-performance liquid chromatography; cis-R4′G, cis-resveratrol 4′-O-glucuronide; E-H, Eadie-Hofstee plots; ANOVA, analysis of variance; PSI, partial substrate inhibition.
- Accepted April 29, 2009.
- Received March 3, 2009.
- The American Society for Pharmacology and Experimental Therapeutics
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