Abstract
This paper reported development, validation and stability evaluations of a simple, selective, sensitive, precise, accurate, and rapid high-performance liquid chromatographic method for the determination of levofloxacin in human plasma. The method was validated according to US FDA norms. Extraction procedure employed a simple and rapid single-step protein precipitation to isolate levofloxacin from the plasma. Mobile phase consisted of 4 volumes of acetonitrile and 96 volumes of aqueous phase containing 1.5% v/v tetrabutylammonium hydroxide, 0.3% v/v of triethylamine as ion-pairing agents, adjusted at 0.8 ml/min for isocratic separation. Levofloxacin and norfloxacin (internal standard) were separated on Zorbax SB-CN, a reversed-phase C18 HPLC column kept at 40 degrees C. The quantification was done by fluorescence detector set at an excitation and emission wavelengths of 290 and 456 nm, respectively. The method was found linear within 03-2039 mu g/mL of levofloxacin in plasma with a coefficient of correlation over 0.998. The overall precision and accuracy ranged between 1-7%. The stability of levofloxacin in plasma at different working conditions and long term storage condition were found similar to 98-103% and 93-95%, respectively. The utility of the method is reflected in its simplicity, rapidity, cost-effectiveness such as a high-throughput sample processing and analysis (a batch of 18 samples for a particular time period was processed within 20 min followed by a short run time of 12 min). The mean pharmacokinetic parameters of 18 subjects such as C-max, T-max & AUC were 550 +/- 1.36 mu g/mL, 1306 +/- 0.6836 h, and 4038 +/- 8.1751 mu gh/mL, respectively.