The aim of this study was to describe a high-performance liquid chromatography (HPLC) assay for the determination of cefepime and cefpirome in human serum without changing chromatographic conditions. The assay consisted to measure cefepime and cefpirome which were unbound to proteins having a molecular mass of 10,000 or more by ultrafiltration followed by HPLC with a Supelcosil ABZ+ column and UV detection at a wavelength of 263 nm. The assay was been found to be linear and has been validated over the concentration range 200 to 0.50 microg/ml for both cefepime and cefpirome, from 200 microl serum, extracted. In future, the assay will support therapeutic drug monitoring for cefepime and cefpirome in neutropenic patients in correlation with microbiological parameters such as MIC90 (minimal inhibitory concentration of antibiotic which kills 90% of the initial bacterial inoculum) and clinical efficacy.