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曾有报道用 HPLC 同时测定人血清中的卤泛曲林(HAL)和去丁基卤泛曲林(BHAL),但分析时间长,内源性物质与BHAL 几乎同时出峰,对测定有干扰。本文报道的方法迅速、分辨率更高。实验采用 757 可变波长 UV-VIS 检测器。Ultrasphere C_8 柱(5μm,25cm×4.6mm)。内标为2,4-二氯-9-(2-二氯基-1-羟基)乙基-6-三氟甲基菲盐酸盐。流动相为乙腈∶水(75∶25,v/v)与0.2%(w/v)十二烷基硫酸钠和0.2%(v/v)冰醋酸。在含0.5ml 血清的试管中,加入内标50μl(游离碱4μg/ml),乙腈1ml,沉淀出蛋白质,涡流混合20s,1200g 离心10min,将上清液移至 C_8-键合洗脱柱,用真空仪抽入柱体,脂乙腈洗柱后,药物被1.0ml 混合液[1.0mol/L 盐酸∶乙腈,1∶9(v/v)]洗脱下。用硅试管收集洗脱液,洗脱液中加入NH_4OH 1.0ml,CH_2Cl_2 6.0ml,旋转器上旋转25min,1000g 离心5min,分出有机相,在40℃通氮下挥发至干,践留物用150μl 流动相溶解,取25μl 注入 HPLC 柱。
It has been reported that simultaneous determination of halibuterin (HAL) and behenbuterol (BHAL) in human serum by HPLC has been reported for a long time, with almost simultaneous peak of endogenous substances and BHAL, interfering with the assay . The method reported in this paper is rapid and has higher resolution. Experiments using 757 variable wavelength UV-VIS detector. Ultrasphere C_8 column (5 μm, 25 cm × 4.6 mm). The internal standard is 2,4-dichloro-9- (2-dichloro-1-hydroxy) ethyl-6-trifluoromethylphenanthrene hydrochloride. The mobile phase was acetonitrile: water (75:25, v / v) with 0.2% (w / v) sodium lauryl sulfate and 0.2% (v / v) glacial acetic acid. In a test tube containing 0.5 ml of serum, 50 μl of an internal standard (4 μg / ml of free base) and 1 ml of acetonitrile were added to precipitate protein. The mixture was vortexed for 20 seconds and centrifuged at 1200 g for 10 minutes. The supernatant was transferred to a C_8- After the column was vacuumed into the column and the column was washed with lipid acetonitrile, the drug was eluted with 1.0 ml of a mixed solution [1.0 mol / L hydrochloric acid: acetonitrile, 1: 9 (v / v)]. Collect the eluate with a silicon tube. Add 1.0 ml of NH 4 OH and 6.0 ml of CH 2 Cl 2 to the eluate. Spin on a spinner for 25 min and centrifuge at 1000 g for 5 min. Separate the organic phase and evaporate to dryness at 40 ° C. with nitrogen. Dissolve 150μl of mobile phase and inject 25μl into HPLC column.