反相HPLC测定培养基中莫西沙星含量的方法学研究

 目的 建立特定培养基中莫西沙星的HPLC测定法。方法 取样品100μL,以加替沙星作内标,采用乙腈沉淀蛋白;常温下氮气吹干,用0.01 mol·L^-1盐酸200μL复溶后进样。色谱柱:Alltech Nucleosil 100(C₁₈)柱(10μm,4.6 mm×250mm);流动相:缓冲液-甲醇-乙腈(66.8:15.1:18.1),缓冲液为三乙胺-磷酸-水(0.37:0.30:99.3);流速1.3 mL·min^-1,荧光检测器的激发波长和发射波长分别为295和418nm。结果在1.0-50.0 μg·mL^-1内线性关系良好,相关系数r=0.999 9(n=7)。高低2个浓度质控样品的日内RSD在0.67％～0.81％之间,日间RSD在0.48％～0.89％之间,方法学回收率为98.43％～103.51％。结论本法灵敏、可靠、耐用、操作简便易行、分析时间短,并成功地应用于莫西沙星体外模型的细菌耐药性发展研究。

A reverse phase high performance liquid chromatographic assay for determination of moxifloxacin in special growth media

OBJECTIVE To develop a HPLC method for the determination of moxifloxacin in special growth media.METHODS With gatifloxacin as the internal standard,the protein in 100μL of samples containing broth media was precipitated by acetonitrile.The supernatant layer was evaporated under nitrogen stream and the residual was reconstituted with 200μL of O.Olmol·L^-1 HC1.A Alltech Nucleosil 100(C₁₈) column was used and the mobile phase consisted of buffer-methanol-acetonitrile (66.8:15.1:18.1), the buffer consisted of triethylamine-phosphoric acid-water(0.37:0.30:99.3).The flow rate of mobile phase was 1.3mL·min^-1.Excitation and emission wavelengths of the fluorometer were at 295 and 418 nm, respectively.RESULTS A linearity was obtained from 1.0 to 50.0μg·mL^-1 of moxifloxacin in broth (5%LHB CAMHB,a cation-adjusted mueller-hinton broth with 5% lysed horse blood) with a good correlation coefficient (r=0.999 9,n=7) .The in tra-run and inter-run validation were less than 0.89% .The mean recoveries were 98.43% and 103.51% for the high and low concentrations of the quality control samples,respectively.CONCLUSION The HPLC assay was sensitive,reliable, reproducible and simple, and the run time was only 10 minutes. The method was successfully applied to unknown samples of moxifloxacin in 5%LHB-CAMHB obtained from an in vitro model BAL study.