Development and validation of an LC-ESI-QTOF-MS method to measure cefepime in the plasma and peritoneal fluid of rats...

Cefepime (CEF) is a cephalosporin and can be administered in secondary peritonitis together with metronidazole to treat sepsis. This study aimed to develop and validate an LC-ESI-QTOF-MS method for the quantification of cefepime in the plasma and peritoneal microdialysate of healthy Wistar rats. Chromatographic separation was performed using a CLC-ODS C₁₈ column (250 × 4.6 mm), a C₁₈ pre-column (4 mm, 5 μm) and isocratic elution. Gallic acid was used as the internal standard. The mobile phase consisted of (A) ultrapure water (pH adjusted to 3.5) and (B) acetonitrile (80:20, v/v) at 0.8 ml/min. Quantification was performed using a mass spectrometer with electrospray ionization in positive mode to monitor ions with m/z 481.1322 (CEF) and m/z 171.0288 (IS). The method was validated for selectivity, precision, accuracy, linearity, stability, lower limit of quantification, carryover, recovery and matrix effect. Calibration was done in the ranges 1-40 and 1-100 μg/ml for the peritoneal microdialysate and plasma, respectively. Plasma extraction recovery ranged from 93.9 to 99.9%. The technique was validated and successfully applied in a pilot pharmacokinetic study for estimating the free concentration of CEF in the peritoneal microdialysate of rats for the first time.