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Human glutathione S-transferase-mediated glutathione conjugation of curcumin and efflux of these conjugates in caco-2 cells

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Author: Usta, M. · Wortelboer, H.M. · Vervoort, J. · Boersma, M.G. · Rietjens, I.M.C.M. · Bladeren, P.J. van · Cnubben, N.H.P.
Institution: TNO Kwaliteit van Leven
Source:Chemical Research in Toxicology, 12, 20, 1895-1902
Identifier: 240418
doi: doi:10.1021/tx7002245
Keywords: Biology · Toxicology and Applied Pharmacology · curcumin · glutathione transferase · adult · aged · article · atom · catalysis · cell strain CACO 2 · conjugate · drug degradation · drug excretion · drug structure · female · glutathione metabolism · human · human cell · human tissue · intestine · intestine epithelium cell · liquid chromatography · liver cytosol · male · mass spectrometry · monolayer culture · proton nuclear magnetic resonance · Adult · Aged · Caco-2 Cells · Curcumin · Cytosol · Female · Glutathione · Glutathione Transferase · Humans · Intestines · Liver · Magnetic Resonance Spectroscopy · Male · Metabolic Detoxication, Phase II · Middle Aged · Molecular Structure


Curcumin, an α,β-unsaturated carbonyl compound, reacts with glutathione, leading to the formation of two monoglutathionyl curcumin conjugates. In the present study, the structures of both glutathione conjugates of curcumin were identified by LC-MS and one- and two-dimensional 1H NMR analysis, and their formation in incubations with human intestinal and liver cytosol and purified human glutathione S-transferases and also in human Caco-2 cells was characterized. The results obtained demonstrate the site for glutathione conjugation to be the C1 atom, leading to two diastereoisomeric monoglutathionyl curcumin conjugates (CURSG-1 and CURSG-2). The formation of both glutathionyl conjugates appeared to be reversible. The monoglutathionyl curcumin conjugates decompose with a t1/2 of about 4 h to curcumin and other unidentified degradation products. Both human intestinal and liver cytosol catalyzed curcumin glutathione conjugation. At saturating substrate concentrations, human GSTM1a-1a and GSTA1-1 are shown to be especially active in the formation of CURSG-1, whereas GSTP1-1 and GSTA2-2 have no preference for the formation of CURSG-1 or CURSG-2. GSTT1-1 hardly catalyzes the glutathione conjugation of curcumin. In the Caco-2 human intestinal monolayer transwell model, CURSG-1 and CURSG-2 were formed at a ratio of about 2:1 followed by their excretion, which appeared to be three times higher to the apical (lumen) side than to the basolateral (blood) side. Given that GSTM1a-1a and GSTP1-1 are present in the intestinal epithelial cells, it can be concluded that efficient glutathione conjugation of curcumin may already occur in the enterocytes, followed by an efficient excretion of these glutathione conjugates to the lumen, thereby reducing the bioavailability of (unconjugated) curcumin. In conclusion, the present study identifies the nature of the diastereoisomeric monoglutathionyl curcumin conjugates, CURSG-1 and CURSG-2 formed in biological systems, and reveals that conjugate formation is catalyzed by GSTM1a-1a, GSTA1-1, and/or GSTP1-1 with different stereoselective preference. The formation of glutathione conjugates can already occur during intestinal transport, after which the monoglutathionyl conjugates are efficiently excreted to the intestinal lumen, thereby influencing the bioavailability of curcumin and, as a result, its beneficial biological effects. © 2007 American Chemical Society.