title: Development and validation of a high-performance liquid chromatography-fluorescence detection method for the accurate quantification of colistin in human plasma.
abstract: Recently, colistin has become one of the most important drugs for treating infections caused by multidrug-resistant Gram-negative bacteria. Therapeutic drug monitoring is recommended to ensure the safety and efficacy of colistin and to improve clinical outcomes. This study developed an accurate and sensitive high-performance liquid chromatography-fluorescence detection (HPLC-FLD) method for the quantification of colistin in human plasma. The sample preparation included protein precipitation using trichloroacetic acid (TCA) and methanol, followed by in-solid phase extraction (In-SPE) derivatization with 9-fluorenylmethyl chloroformate (FMOC-Cl). A Poroshell 120 EC-C18 2.1×100mm (2.7μm) column was used in the HPLC method with a mobile phase composed of acetonitrile (ACN), tetrahydrofuran (THF), and deionized (DI) water (82%, 2%, 16% (v/v), respectively). Polymyxin B1 was used as the internal standard. The total analysis time was 22min under optimal separation conditions. The HPLC-FLD method was validated over a therapeutic range of 0.3-6.0μgmL(-1). The intra-day and inter-day precisions for colistin A and colistin B were below 9.9% and 4.5% relative standard deviations, respectively. The accuracy test results were between 100.2 and 118.4%. The extraction recoveries were between 81.6 and 94.1%. The method was linear over the test range, with a 0.9991 coefficient of determination. The limit of detection was 0.1μgmL(-1). The validated HPLC-FLD method was successfully applied to quantify the colistin concentrations in 2 patient samples for therapeutic drug monitoring.
authors.affiliation: School of Pharmacy, College of Medicine, National Taiwan University, Taipei, Taiwan.
authors.forename: Divyabharathi
authors.initials: D
authors.lastname: Chepyala
authors.affiliation: School of Pharmacy, College of Medicine, National Taiwan University, Taipei, Taiwan.
authors.forename: I-Lin
authors.initials: IL
authors.lastname: Tsai
authors.affiliation: Department of Internal Medicine, National Taiwan University Hospital, Taipei, Taiwan.
authors.forename: Hsin-Yun
authors.initials: HY
authors.lastname: Sun
authors.affiliation: School of Pharmacy, College of Medicine, National Taiwan University, Taipei, Taiwan; Graduate Institute of Clinical Pharmacy, College of Medicine, National Taiwan University, Taiwan; Department of Pharmacy, National Taiwan University Hospital, Taipei, Taiwan.
authors.forename: Shu-Wen
authors.initials: SW
authors.lastname: Lin
authors.affiliation: School of Pharmacy, College of Medicine, National Taiwan University, Taipei, Taiwan; Department of Internal Medicine, National Taiwan University Hospital, Taipei, Taiwan. Electronic address: kuoch@ntu.edu.tw.
authors.forename: Ching-Hua
authors.initials: CH
authors.lastname: Kuo
mesh_terms: D000097:Acetonitriles / Q000737:chemistry; D002851:Chromatography, High Pressure Liquid / Q000379:methods*; D003091:Colistin / Q000097:blood; D005453:Fluorescence; D005663:Furans / Q000737:chemistry; D006801:Humans; D015203:Reproducibility of Results
chemical_list: D000097:Acetonitriles; D005663:Furans; C018674:tetrahydrofuran; C032159:acetonitrile; D003091:Colistin
keywords: Colistin; FMOC-Cl; HPLC-FLD; Human plasma; Therapeutic drug monitoring