LC-MS/MS based quantitation of ciprofloxacin and its application to antimicrobial resistance study in Balb/c mouse plasma, urine, bladder and kidneys

Abstract

The authors proposed a simple, high throughput liquid chromatography-tandem mass spectrometric (LC-MS/MS) method for the determination of ciprofloxacin in small sample volumes of plasma, urine, bladder and kidneys of mice. d₈-Ciprofloxacin was used as an internal standard (IS) which efficiently tracked the matrix effect in different matrices. The method employed 20 μL of plasma/30 μL of urine or tissue homogenate for sample processing using a simple protein precipitation technique. The processed samples were chromatographed on a C₁₈ column by using a mixture of 5 mM ammonium formate (0.1% formic acid)–acetonitrile as the mobile phase at a flow rate of 0.6 mL min⁻¹ with a gradient elution for 3 min. The calibration curve obtained was linear over the concentration range of 100–5000 ng mL⁻¹ with r² ≥ 0.99 in all matrices. Method validation was performed as per FDA guidelines and the results met the acceptance criteria. The multiple reaction-monitoring mode (MRM) was used for the quantification of ion transitions at m/z 332.1/230.8 and 340.1/296.1 for the analyte and the IS, respectively. The validated method was used for preclinical studies of antimicrobial resistance in mice.