INSERM, ERI-23, Poitiers, France.
Antimicrob Agents Chemother. 2010 May;54(5):1941-8. doi: 10.1128/AAC.01367-09. Epub 2010 Feb 22.
A rapid high-performance liquid chromatography-tandem mass spectrometry (LC-MS/MS) assay was developed for the routine quantification of colistins A and B and their prodrugs, colistin methanesulfonate (CMS) A and CMS B, respectively, in human plasma and urine by using polymyxin B1 as the internal standard (IS). CMS concentrations were determined indirectly by subtracting the colistin concentrations determined in biological samples from the whole colistin concentrations determined after sample treatment with sulfuric acid in order to hydrolyze CMS into colistin. After extraction on a solid-phase extraction column, the colistins were separated on an XBrigde C(18) column with isocratic elution (run time, 3.8 min). The mobile phase was 0.1% (vol/vol) formic acid in acetonitrile-0.1% (vol/vol) formic acid in water (20:80, vol/vol), run at a 0.2-ml/min flow rate. Ions were detected in the turbo-ion-spray-positive and multiple-reaction-monitoring modes. The ions monitored (precursor M + 2H to product ions) were m/z 585.5/101.2 for colistin A, m/z 578.5/101.2 for colistin B, and m/z 602.5/241.2 for IS. Prevalidation studies demonstrated the stability of CMS in biological samples and extracts, a key point for the reliable quantification of colistin and CMS. The assay was accurate and reproducible for the quantification of colistins A and B and CMSs A and B in plasma samples over concentration ranges appropriate for pharmacokinetic studies: 0.024 to 6.144, 0.015 to 3.856, 0.029 to 7.492, and 0.010 to 2.508 microg/ml, respectively. In urine samples, the assay was validated over the same concentration ranges for colistins and over concentration ranges of 0.058 to 7.492 microg/ml and 0.020 to 2.508 microg/ml for CMSs A and B, respectively.
建立了一种快速高效液相色谱-串联质谱（LC-MS/MS）测定法，用于常规定量人血浆和尿液中的黏菌素 A 和 B 及其前体药物硫酸黏菌素甲磺酸盐（CMS）A 和 CMS B，以多黏菌素 B1 为内标（IS）。通过在样品处理后用硫酸将 CMS 水解为黏菌素，从生物样品中测定的黏菌素浓度中减去 CMS 浓度，间接测定 CMS 浓度。经过固相萃取柱萃取后，用 XBrigde C（18）柱等度洗脱（运行时间 3.8 分钟）分离黏菌素。流动相为乙腈-水（20：80，体积/体积）中的 0.1%（体积/体积）甲酸-0.1%（体积/体积）甲酸，流速为 0.2ml/min。离子在涡轮离子喷雾正模式和多重反应监测模式下检测。监测的离子（前体 [M + 2H]（2+）到产物离子）为 m/z 585.5/101.2 的黏菌素 A、m/z 578.5/101.2 的黏菌素 B 和 m/z 602.5/241.2 的 IS。预验证研究表明 CMS 在生物样品和提取物中的稳定性，这是可靠定量黏菌素和 CMS 的关键。该测定法准确且可重现，可用于定量血浆样品中黏菌素 A 和 B 以及 CMSs A 和 B 的浓度范围：0.024 至 6.144、0.015 至 3.856、0.029 至 7.492 和 0.010 至 2.508 µg/ml。在尿液样品中，该测定法在相同的浓度范围内验证了黏菌素，并且在 CMSs A 和 B 的浓度范围为 0.058 至 7.492 µg/ml 和 0.020 至 2.508 µg/ml 时进行了验证。
