Determination of paracetamol and its metabolites in urine by high-performance liquid chromatography using reversed-phase bonded supports.

Paracetamol and its four main metabolites, the sulphate (S), glucuronide (G), cysteine (C) and mercapturic acid (M) conjugates, are readily separated in a synthetic mixture using slightly acidic aqueous alcoholic eluents (e.g. water-methanol-formic acid, 85:15:0.15, v/v/v) on either octadecyl silica (ODS silica) or octadecyl silica which has been further silanized to remove residual hydroxyl groups (ODS/TMS silica). The dependences of k' upon alcohol, acid and added salt concentrations are reported for both materials. The latter material gives the higher plate efficiencies and is much superior when applied to analysis of urines taken after therapeutic doses and overdoses of paracetamol. At least four additional metabolites are reported in overdose urines. Mass spectrometric analysis (high and low resolution) has confirmed the identity of M and identified one of the additional metabolites as methoxyparacetamol. Mass spectra of the remaining additional metabolites enable major structural features to be deduced. One of these metabolites may be associated with liver damage.