柱前衍生化HPLC法测定注射用氨磷汀的含量

HPLC method using pre-column derivatization for determinating the content of amifostine

目的建立一种测定注射用氨磷汀含量的HPLC法。方法氨磷汀经氯甲酸芴甲酯(9-fluorenylmethylchloroformate,FMOC-Cl)衍生化后用HPLC法分析。采用Agilent C18柱(150 mm×4.6 mm,5μm)色谱柱,流动相A:乙腈,B:水。0～4 min,A∶B=30∶70;4.1～10 min,A∶B=98∶2,每次进样前再平衡7 min;柱温25℃,流速1 mL/min,检测波长265 nm。外标法计算含量。结果在该色谱条件下,氨磷汀在5～100μg/mL浓度范围内有良好的线性关系(r>0.999,n=6),回收率为97%～103%(n=5),RSD<2.0%(n=5)。最低检测质量浓度为100 ng/mL。样品在6 h内稳定。测得样品含量合格,在90%～110%之间。结论经方法学实验验证,该方法简便、快速、结果准确、可靠、重现性好。

Objective To establish a HPLC method for the determination of amifostine. Methods 9-fluorenylmethyl chloroformate （FMOC-C1） was used for derivatization. An agilent C,s column （4.6 mm 150 ram, 5 l,m） was adopted as stationary phase. Elution was carried out with acetonitrile （solvent A） and water （solvent B） at a flow-rate of 1 mI./min. Gradient HPLC was used with the solvent ratio changed from 30：70 （for 4 rain） to 98：2 （from 4. 1 rain to 10 min） for solvent A：B, respectively,and the rebalancing time was 7 rain. The detection wavelength was 265 nm and the temperature of column was 25℃. Results Amifostine had a good linearity （r〉0. 999,n = 6） in the range of 5 - 100 μg/mL. The recovery values were between 97% and 103% ,RSD（2. 0% （n = 3）. The detection limit value was 100 ng/mL. The samples were stable in 6 hour and qualified between 90% and 110%. Conclusions The method was proved to be accurate, sensitive and reliable.