Simultaneous determination of trimethoprim, sulphamethoxazole and N4-acetylsulphamethoxazole in serum and urine by high-performance liquid chromatography.

The simultaneous determination of trimethoprim, sulphamethoxazole and N4-acetyl-sulphamethoxazole in serum and urine by high-performance liquid chromatography using sulphafurazole as internal standard is described. The separation was achieved on a reversed-phase column employing acetic acid-methanol as the mobile phase with spectrophotometric detection at 230 nm. Precise simultaneous quantitative analysis of the relative components has been achieved at levels of 0.1 microgram/ml for trimethoprim and 1.0 microgram/ml for both sulphamethoxazole and its N4-acetyl metabolite using 1 ml of serum of urine.