头孢他啶与头孢吡肟治疗药物监测的高效液相色谱方法探索及其临床采样流程建立

Determination of cefepime and ceftazidine in human plasma by HPLC method and its establishment of clinical sampling process

目的:建立同时测定头孢他啶和头孢吡肟血药浓度的高效液相色谱(high performance liquid chromatography,HPLC)法及其临床采样流程,并应用于临床治疗药物监测。方法:采用CAPCELL PAK C18(4.6 mm×250 mm,5.0μm)色谱柱进行色谱分离,流动相A为50 mmol·L-1磷酸二氢钾溶液,流动相B为混合有机相(乙腈∶甲醇∶水=7∶2∶1),A∶B(V/V,93∶7),流速1.0 mL·min-1,波长为254 nm,盐酸雷尼替丁为内标,以ACP-1去蛋白剂沉淀蛋白,旋涡离心后进样30μL分析,同时考察全血中两药在不同抗凝管、不同温度下放置不同时间的稳定性。结果:头孢他啶和头孢吡肟的血浆质量浓度线性范围分别是0.57~267.34μg·mL-1、0.54~208.49μg·mL-1,低、中、高质控样品的日内、日间精密度均小于15%,萃取回收率分别为90.9%~95.4%、88.6%~97.7%;全血稳定性试验中,以EDTA-K2管采血的头孢他啶与头孢吡肟血浆在6℃及24℃下均能稳定48 h,37℃下稳定10 h;而以肝素钠管采血的头孢他啶和头孢吡肟血浆在6℃及24℃下能稳定24 h,37℃下能稳定4 h。结论:所建立的方法具有灵敏度高、稳定性好、操作简便等优点,并根据全血稳定性结果建立了一套临床采样流程,为头孢他啶和头孢吡肟的TDM标准化与规范化建设提供参考依据。

OBJECTIVE To establish a high performance liquid chromatography(HPLC)method for the simultaneous determination of ceftazidime and cefepime plasma concentrations and its clinical sampling process,and to apply it to clinical therapeutic drug monitoring.METHODS This method was performed on a CAPCELL PAK C18(4.6 mm×250 mm,5.0μm)column.Mobile phase A was potassium dihydrogen phosphate solution(50 mmol·L-1)and mobile phase B was mixed organic phase(acetonitrile∶methanol∶water=7∶2∶1),A∶B(V/V,93∶7).The flow rate and detection wavelength were 1.0 mL·min-1 and 254 nm.Ranitidine hydrochloride was used as internal standard.ACP-1 deproteinizer was used to precipitate protein.Vortex centrifugation was followed by injection of 30μL for analysis.The stability of the two drugs placed in different anticoagulants at different temperatures for different times was also investigated.RESULTS The linear ranges of plasma concentration of ceftazidine(or cefepime)was 0.57-267.34μg·mL-1(or 0.54-208.49μg·mL-1).The intra-run precision and inter-runprecision of quality control samples of ceftazidine and cefepime for low,medium and high concentrations were less than 15%and the extraction recovery of ceftazidinem(or cefepime)was 90.9%-95.4%(or 88.6%-97.7%).In the whole blood stability results,ceftazidime and cefepime in EDTA-K2 tubes could remain stable for 48 h under 6℃and 24℃,and for stabilized 10 h under 37℃,while they could remain stable for 24 h at 6℃and 24℃in heparin sodium tubes,and remain stable for 4 h at 37℃.CONCLUSION This method had advantages of high sensitivity,high stability and simple operations.According to the whole blood stability results,a set of completed clinical sampling process was established,which provided reference for the TDM standardization of ceftazidime and cefepime.