Bioanalytical development and validation of liquid chromatographic–tandem mass spectrometric methods for the quantification of total and free cefazolin in human plasma and cord blood

摘要

Objectives Cefazolin is a commonly prescribed β-lactam antibiotic for prophylaxis against skin infections following surgery, including caesarean sections. Assessment of maternal and neonatal exposure is important for correlating drug concentrations to clinical outcomes. Thus, bioanalytical methods for the quantification of both total and free cefazolin in maternal plasma and cord blood can assist in the comprehensive evaluation of cefazolin exposure. Design and methods Specimen preparation for the measurement of total cefazolin was performed via protein precipitation with acetonitrile containing the internal standard cloxacillin. Ultrafiltration was used to isolate free cefazolin. Processed samples were analyzed on a Prelude SPLC system coupled to a TSQ triple quadrupole Vantage mass spectrometer. Methods were validated following FDA bioanalytical guidelines. Results The analytical measuring ranges of these methods were 0.48–480μg/mL and 0.048–48μg/mL for total and free drug, respectively. Calibration curves were generated using 1/ x 2 weighted linear regression analysis. Total cefazolin demonstrated inter- and intra-assay precision of ≤20% at the LLOQ and ≤11.2% at other levels. Free cefazolin demonstrated inter- and intra-assay precision of ≤18.5% at the LLOQ and ≤12.6% at other levels, respectively. Accuracy (%DEV), carryover, matrix effects, recovery and stability studies were also acceptable based on FDA recommendations. Furthermore, it was demonstrated that samples prepared in cord blood can be accurately quantified from an adult plasma calibration curve, with recoveries ≤9.1% DIF and ≤11.9% DIF for total and free cefazolin, respectively. Conclusions The described LC–MS/MS methods allow for the measurement of total and free cefazolin in both plasma and cord blood. Highlights ? LC-MS/MS methods for quantification of total and free cefazolin were developed. ? Methods allow for cefazolin quantification in human plasma and cord blood. ? Validation studies were based on FDA guidelines. ? Methods and statistical graphics for carryover optimization are demonstrated.