The development of HPLC dual wavelength fluorescence detection method for the simultaneous determination of four fluoroquinolones in human plasma

Abstract

A high-performance liquid chromatography-fluorescence detection (HPLC-FLD) method for determination of the fluoroquinolones norfloxacin (NOR), ciprofloxacin (CIP), levofloxacin (LEVO), and moxifloxacin (MOXI) in human plasma has been successfully developed. The detection was optimized with a dual wavelength excitation and emission fluorescence detector. Analytical wavelengths of λex/λem=290/500 nm (for LEVO and MOXI) and λex/λem=280/445 nm (for NOR and CIP) were found when measuring fluoroquinolones in the mixture. The plasma sample was pre-treated during the deproteinization step using methanol. This method was validated by linearity in the ranges of 0.05-15; 0.05-10; 0.05-15; and 0.10-10 µg ml-1 with limits of detection of 0.0071; 0.0080; 0.0074; and 0.0171 µg ml-1 for NOR, CIP, LEVO, and MOXI, respectively. Further, the HPLC-FLD method was proven to be precise and accurate with relative standard deviations lower than 6% and recoveries ranging from 92.5-105.4% for all four fluoroquinolones. Therefore, the proposed HPLC-FLD method provides an alternative approach for the simultaneous analysis of fluoroquinolones in plasma.A high-performance liquid chromatography-fluorescence detection (HPLC-FLD) method for determination of the fluoroquinolones norfloxacin (NOR), ciprofloxacin (CIP), levofloxacin (LEVO), and moxifloxacin (MOXI) in human plasma has been successfully developed. The detection was optimized with a dual wavelength excitation and emission fluorescence detector. Analytical wavelengths of λex/λem=290/500 nm (for LEVO and MOXI) and λex/λem=280/445 nm (for NOR and CIP) were found when measuring fluoroquinolones in the mixture. The plasma sample was pre-treated during the deproteinization step using methanol. This method was validated by linearity in the ranges of 0.05-15; 0.05-10; 0.05-15; and 0.10-10 µg ml-1 with limits of detection of 0.0071; 0.0080; 0.0074; and 0.0171 µg ml-1 for NOR, CIP, LEVO, and MOXI, respectively. Further, the HPLC-FLD method was proven to be precise and accurate with relative standard deviations lower than 6% and recoveries ranging from 92.5-105.4% for all four fluoroquinolones. Therefore, the proposed HPLC-FLD method provides an alternative approach for the simultaneous analysis of fluoroquinolones in plasma.

Keywords

• dual wavelength excitation and emission
• fluoroquinolones
• HPLC-FLD, plasma