高效液相色谱法测定人血清中氨苄西林、丙磺舒的浓度研究

Determination of ampicillin and probenecid in human serum by high performance liquid chromatography

目的：建立测定人血清中氨苄西林、丙磺舒浓度的高效液相色谱法。方法：（1）氨苄西林,以诺氟沙星为内标,血清样品用0.6%的高氯酸沉淀蛋白后进行测定;色谱柱为HigginsTMC18柱（250 mm×4.6 mm,5μm）;流动相为有机相（甲醇-乙腈-异丙醇,100∶2∶5,v/v）：水相（0.01 mol/L磷酸二氢钾,pH3.5）=23∶77;检测波长210 nm;流速1.0 ml/min;柱温30℃。（2）丙磺舒,以缬沙坦为内标,血清样品用乙腈沉淀蛋白后进行测定;色谱柱为HigginsTMC18柱（250 mm×4.6 mm,5μm）;流动相为有机相（甲醇-乙腈,400∶275,v/v）：水相（乙酸-水,1∶60,v/v）=77∶23;检测波长280 nm;流速1.0 ml/min;柱温30℃。结果：氨苄西林在0.14~11.2μg/ml浓度范围内线性关系良好（r=0.999 5）,最低检测限为0.14μg/ml;丙磺舒在0.20~16.0μg/ml浓度范围内线性关系良好（r=0.999 1）,最低检测限为0.20μg/ml。结论：高效液相色谱法操作简便、灵敏、专一性好,适用于人体药动学研究。

Objective：To establish a method to determine the concentration of ampicillin and probenecid in human serum by high performance liquid chromatography（HPLC）.Methods：（1）Ampicillin was detected by HPLC using norfloxacin as internal standard after sample was sedimented with 0.6% perchloric acid.HigginsTM C18 column（250 mm×4.6 mm,5 μm） was used,the mobile phase consisted of organic phase（methanol-acetonitrile-isopropanol,100∶2∶5,v/v） and aqueous phase（0.01 mol/L potassium dihydrogen phosphate,pH 3.5） with a ratio of 23∶77,the flow rate was 1.0 ml/min,the detection wavelength was set at 210 nm,and column oven at 30 ℃.（2）Probenecid was detected by HPLC using valsartan as internal standard after sample was sedimented with acetonitrile.HigginsTM C18 column（250 mm×4.6 mm,5 μm） was used,the mobile phase consisted of organic phase（methanol-acetonitrile,400∶275,v/v） and aqueous phase（acetic acid-water,1∶60,v/v） with a ratio of 77∶23,the flow rate was 1.0 ml/min,the detection wavelength was set at 280 nm,and column oven at 30 ℃.Results：Good linearity was obtained within the range of 0.14-11.2 μg/ml for ampicillin（r=0.999 5）,and 0.20-16.0 μg/ml for probenecid（r=0.999 1）.The mininum detectable concentrations for ampicillin,and probenecid were 0.14 μg/ml and 0.20 μg/ml,respectively.Conclusions：This method is simple,sensitive and specific,which can be applied to study the pharmacokinetics of ampicillin and probenecid.