Enzyme-Assisted Synthesis and Structural Characterization of Nitrocatechol Glucuronides

Enzyme-assisted synthesis and characterization are described for 3-O-β-d-glucuronides 1b−4b of the aglycons E- and Z-2-cyano-N,N-diethyl-3-(3,4-dihydroxy-5-nitrophenyl)propenamide (entacapone), 1a and 2a, respectively, 3-(3,4-dihydroxy-5-nitrobenzylidene)-2,4-pentanedione (nitecapone) 3a and 4‘-methyl-3,4-dihydroxy-5-nitrobenzophenone (tolcapone) 4a, and 1-o- and 2-o-glucuronides 5b and 6b of the aglycon 1,2-dihydroxy-4-nitrobenzene 5a. Liver microsomes from rats pretreated with Aroclor 1254 were used as catalyst in the synthesis. Glucuronidation was regio- and stereoselective in the case of 1a−4a; only one product was observed by HPLC, HPTLC, and NMR. The glucuronidation of 1,2-dihydroxy-4-nitrobenzene 5a resulted in equal amounts of 1-O-β-d- and 2-O-β-d-glucuronides. Purification of the crude products by C18 solid-phase extraction and/or flash chromatography gave compounds 1b−6b in 38−98% yields (50−84 mg). The structures of the glucuronides were characterized on the basis of UV and IR spectra and confi...