title: Overestimation of flavonoid aglycones as a result of the ex vivo deconjugation of glucuronides by the tissue β-glucuronidase.
abstract: Flavonoid glucuronides are the main circulating metabolites of flavonoids in humans and animals. There has been a growing interest in the biological function of glucuronides. In order to differentiate biological activity and to assess efficacy it is essential to accurately determine the levels of flavonoid aglycone and metabolic conjugate in vivo. Many organs and body fluids of humans and animals exhibit β-glucuronidase against flavonoid glucuronides. Studies have shown that β-glucuronidase within the tissues hydrolyzes glucuronides to their aglycones during the tissue extraction, leading to artificially higher reported tissue levels of aglycone than actual in vivo concentrations. The aims of this study were to estimate the extent by which the aglycones were overestimated and to investigate the use of saccharo-1,4-lactone, a β-glucuronidase inhibitor, to block the ex vivo hydrolysis of flavonoid glucuronides. Our data demonstrate that in mouse liver tissues and human tumor xenografts levels of quercetin and methylated quercetin aglycones could be over-estimated by 7-fold. The inhibition of deconjugation of quercetin and baicalein glucuronides by saccharo-1,4-lactone is dose-dependent. The amount of saccharo-1,4-lactone used to produce optimal inhibition of the enzyme activity is in the range of 15-24μmol per gram of liver tissue. The use of β-glucuronidase inhibitor blocks the ex vivo deconjugation resulting in an accurate estimation of tissue levels of aglycone and conjugate. Our study described here can be extended to other animal models and human studies with different types of substrates of β-glucuronidase.
authors.affiliation: Department of Medicine, David Geffen School of Medicine, University of California, Los Angeles, CA 90095, USA. Electronic address: qlu@mednet.ucla.edu.
authors.forename: Qing-Yi
authors.initials: QY
authors.lastname: Lu
authors.forename: Lifeng
authors.initials: L
authors.lastname: Zhang
authors.forename: Guido
authors.initials: G
authors.lastname: Eibl
authors.forename: Vay Liang W
authors.initials: VL
authors.lastname: Go
mesh_terms: D000818:Animals; D002851:Chromatography, High Pressure Liquid; D019587:Dietary Supplements; D004305:Dose-Response Relationship, Drug; D044950:Flavanones / Q000737:chemistry; D005419:Flavonoids / Q000037:antagonists & inhibitors / Q000737:chemistry*; D005937:Glucaric Acid / Q000737:chemistry; D005966:Glucuronidase / Q000737:chemistry*; D020719:Glucuronides / Q000737:chemistry; D006023:Glycoproteins / Q000737:chemistry; D006801:Humans; D007783:Lactones / Q000737:chemistry; D008099:Liver / Q000187:drug effects; D008113:Liver Neoplasms; D008745:Methylation; D051379:Mice; D010179:Pancreas / Q000187:drug effects; D011794:Quercetin / Q000737:chemistry; D023041:Xenograft Model Antitumor Assays
chemical_list: C571832:D-Glucaric acid, 1,4-lactone; D044950:Flavanones; D005419:Flavonoids; D020719:Glucuronides; D006023:Glycoproteins; D007783:Lactones; C011473:beta-glucuronidase inhibitor; C006680:baicalein; D011794:Quercetin; D005966:Glucuronidase; D005937:Glucaric Acid
keywords: Baicalin; Glucuronide conjugates; Quercetin; d-Saccharic acid 1,4-lactone; β-Glucuronidase inhibitor