J. Jensen, C. Cornett, C. Olsen
Sep 1, 1993
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Journal
European Journal of Pharmaceutical Sciences
Abstract
Abstract The ability of 5-aminosalicylic acid (5-ASA) to be oxidase to a quinone monoimine compound capable of conjugating with nucleophilic compounds such as N-acetyl-cysteine (NAC) and gluthione (GSH) has been investigated in vitro . There isomeric conjugates of 5-ASA and NAC as well as three isomeric conjugates of 5-ASA and GSH were found to be formed. 5-ASA was initially oxidized by PbO 2 in a solution of TRIS-HC1 buffer pH 9.3 followed by the in situ addition of N-acetyl-cysteine or glutathione to the oxidized 5-ASA at pH 7.5. The resulting conjugates were N-acetylated at the aromatic amino group in order to avoid autooxidation of the products formed. The N-acetylated conjugates were isolated by preparative HPLC and the structures were characterized by nuclear magnetics resonance ( 1 H-NMR and 13 C-NMR) spectroscopy as well as by mass spectrometry (MS) and the data obtained confirmed the formation of thioether linkages between 5-ASA and the SH compounds. The chemical nature of the reactive intermediate capable of adding SH compounds was verified to be the 2-carboxy-quinone monoamine by 1 H-NMR spectroscopy. The N-acetylated conjugates of 5-ASA and NAC were used as reference standards in order to investigate whether such conjugates are excreted in the urine from persons treated with 5-ASA. The N-acetyl-cysteine conjugates could be detected by fluorescense, which resulted in low detection limits ranging from 0.02 μg to 0.06 μg per ml corresponding to the transformation of about 0.003% of the daily dose of 5-ASA into mercapturic acids of 5-ASA, when 1 g of 5-ASA was ingested. In spite of the low detection limits, none of the mercapturic acid conjugates was detected in the urine from persons treated with 5-ASA.