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目的建立固相萃取-高效液相色谱法(SPE-HPLC)测定人血清中万古霉素和去甲万古霉素浓度,并与荧光偏振免疫法(FPIA)测定结果进行相关性分析。方法血清样品经C18固相萃取小柱净化后进样。色谱柱为Hyper Sil C18柱(250 mm×4.6 mm,5μm),流动相为0.05 mol·L?1磷酸二氢钾-乙腈(91︰9),柱温25℃,流速1 m L·min?1,检测波长236 nm,进样量30μL;28例患者的血浆分别用SPE-HPLC和FPIA测定,考察2种方法的相关程度。结果万古霉素、去甲万古霉素的标准曲线范围分别为0.4~80 mg·L?1和1~50 mg·L?1,回收率分别为87.2%和89.8%,日内、日间精密度均<7%;SPE-HPLC和FPIA 2种方法的测定值差异无统计学意义。结论 SPE-HPLC法快速简便,准确,灵敏度高,适用于万古霉素和去甲万古霉素血药浓度检测;2种测定方法有良好的相关性。
OBJECTIVE To establish a SPE-HPLC method for the determination of vancomycin and norvancomycin in human serum and analyze the correlation with the results of fluorescence polarization immunoassay (FPIA). Methods Serum samples were purified by C18 SPE cartridge. The column was a Hyper Sil C18 column (250 mm × 4.6 mm, 5 μm) with a mobile phase of 0.05 mol·L -1 potassium dihydrogen phosphate-acetonitrile (91︰9). The column temperature was 25 ℃ and the flow rate was 1 m L · min- 1, detection wavelength of 236 nm, injection volume of 30μL; 28 patients were measured by SPE-HPLC and FPIA, respectively, to investigate the correlation between the two methods. Results The standard curves of vancomycin and norvancomycin were 0.4-80 mg · L -1 and 1-50 mg · L -1, respectively. The recoveries were 87.2% and 89.8%, respectively. The intra- and inter-day precision <7% respectively. There was no significant difference between the two methods of SPE-HPLC and FPIA. Conclusion SPE-HPLC method is rapid, simple, accurate and sensitive and suitable for the determination of vancomycin and norvancomycin. The two methods have good correlation.