Determination and stability assessment of clarithromycin in human plasma using RP-LC with electrochemical detection

A selective, sensitive and stability-indicating reversed-phase high-performance liquid chromatography method was developed and validated for the determination of clarithromycin antibiotic in human plasma. Liquid chromatography was performed on a 5-µm (100 × 4.6 mm) C8 column at 40°C. The mobile phase consisted of acetonitrile with 0.045M H(3)PO(4) (37:63, v/v) adjusted to pH 6.7 and pumped at a flow rate of 1.2 mL/min. Detections were monitored on an electrochemical detector operated at a potential of 0.85 V with glassy carbon electrode against Ag/AgCl reference electrode. Each analysis required 13 min and quantification over the range of 0.05-5.0 µg/mL of plasma was linear, as indicated by a correlation coefficient (R(2)), 0.9999. The method was validated according to international guidelines. Data with respect to accuracy, within-run and between run, were close to 100% with 4% precision. Absolute recovery was 95%. The limit of quantification was 0.05 µg/mL. Neither endogenous substances nor commonly used drugs were found to interfere with the retention times of analytes. Stock solutions and calibration standards of the drug and quality control preparations were demonstrated to be stable at room temperature and -20°C for long and short periods of time. Eventually, the proposed method was successfully applied to quantify clarithromycin in spiked human plasma and real samples from healthy volunteers, indicating the utility and throughput of this method for clinical and bioavailability studies.