Simple and simultaneous determination of sulphapyridine and acetylsulphapyridine in human serum by column-switching high-performance liquid chromatography.

Objective: A high-performance liquid chromatography (HPLC) with an automated on-line column-switching system was used for the simultaneous determination of sulphapyridine and acetylsulphapyridine, two major active metabolites related to the adverse effects of sulphasalazine, in human serum.
Methods: Serum samples were directly injected into the HPLC, with the valve automatically switched on to remove serum proteins and other hydrophilic components remaining in the pre-column after elution of sulphapyridine and acetylsulphapyridine to the analytical column.
Results: Serum proteins did not interfere with the analysis of either compound. The recoveries of SLP and Ac-SLP from drug-free human serum were 93.03-99.18% and CV were 2.88-4.34%. The within-run reproducibility of assays was excellent with relative standard deviations (RSD) of 1.01-3.90% (SLP) and 0.77-5.56% (Ac-SLP). The limit of quantification of sulphapyridine and acetylsulphapyridine was 3.13 microg/mL and 0.50 microg/mL, respectively. The serum concentrations in a patient with ulcerative colitis, who took 1.0 g sulphasalazine twice daily, were 31.20 microg/mL for sulphapyridine and 14.64 microg/mL for acetylsulphapyridine at 7 h after ingestion.
Conclusion: The present simple and reproducible assay was useful for the monitoring of serum sulphapyridine and acetylsulphapyridine.