Determination of Phenacetin and Its Matabolites in Human Urine and Plasma by High-Performance Liquid Chromatography

Abstract

A method for quantifying phenacetin and its major metabolites has been developed. Metabolites that can be separated include glucuronide and sulphate, cysteine and mercapturic acid conjugates of acetaminophen as well as acetaminophen. The method used high-performance liquid chromatographic separation of these compounds on a C18 reversed-phase column. The mobile phases were methanol, 1% glacial acetic acid and 0.1 M KH₂PO₄ (33: 3: 64) for the unchanged phenacetin and (0: 3: 97) for the separation of metabolites.
4-Fluorophenol was used as an internal standard. The minimum detectable limits (signal to noise ratio≥2) were 20-50ng injected onto column using UV detection at 254nm.