Rapid chromatographic determination of cefotaxime and its metabolite in biological fluids

A reversed-phase high-performance liquid chromatographic assay for the simultaneous determination of cefotaxime and its metabolite desacetylcefotaxime in plasma and urine was developed. Plasma was deproteinized with small amounts of acetonitrile. After separation of the proteins the supernatant was extracted with a mixture of chloroform and 1-butanol. A phase separation was obtained leaving the cephalosporin and its metabolite in the aqueous part and extracting most of the interfering endogenous material. The aqueous phase was injected directly into the chromatograph. As part of the plasma water was dissolved in the acetonitrile--1-butanol--chloroform layer, the concentration of the cephalosporin in the aqueous phase was significantly higher than in the original plasma sample. Therefore, the usual diluting effect of the deproteinization could be avoided. In a similar way the assay was applicable to measure cefotaxime and its metabolite in urine. Calibration curves were set up and were linear up to 25 micrograms/ml for desacetylcefotaxime and 250 micrograms/ml for cefotaxime. The assay was applied to study the pharmacokinetics of cefotaxime and its metabolite in a healthy volunteer. In a similar way this deproteinization and extraction method was also applied to assay for ceftazidime, cephalexin, cephazolin and cefoxitin.