Enzymatic preparation of silybin phase II metabolites: sulfation using aryl sulfotransferase from rat liver
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Aryl sulfotransferase IV (AstIV) from rat liver was overexpressed in Escherichia coli and purified to homogeneity. Using the produced mammalian liver enzyme, sulfation—the Phase II conjugation reaction—of optically pure silybin diastereoisomers (silybin A and B) was tested. As a result, silybin B was sulfated yielding 20-O-silybin B sulfate, whereas silybin A was completely resistant to the sulfation reaction. Milligram-scale sulfation of silybin B was optimized employing resting E. coli cells producing AstIV, thus avoiding the use of expensive 3′-phosphoadenosine-5′-phosphate cofactor and laborious enzyme purification. Using this approach, we were able to reach 48 % conversion of silybin B into its 20-sulfate within 24 h. The sulfated product was isolated by solid phase extraction and its structure was characterized by HRMS and NMR. Sulfation reaction of silybin appeared strictly stereoselective; only silybin B was sulfated by AstIV.
KeywordsSilybin Biotransformation Diastereoisomers Sulfation Aryl sulfotransferase Rat liver
This work was supported by the grant from the Czech Science Foundation P301/11/0767; and projects M200201204 of the Academy of Sciences of the Czech Republic, RV06138897 of the Institute of Microbiology, and by EU ESF COST project MultiGlycoNano CM1102.
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