HPLC法测定注射用环磷腺苷的有关物质

Determination of the Related Substances in Adenosine Cyclophosphate for Injection

目的:采用高效液相色谱法测定注射用环磷腺苷的有关物质,并对已知杂质腺苷、5’-腺苷酸进行控制。方法:采用Waters XBridge BPC_(18)(250 mm×4.6 mm,5μm)色谱柱,以磷酸二氢钾溶液与四丁基溴化铵的混合溶液(取磷酸二氢钾6.8 g与四丁基溴化铵3.2 g,用水溶解并稀释至1 000 ml,摇匀,用磷酸调节pH至4.3)-乙腈(90∶10)为流动相,流速为1.0 ml·min^(-1),检测波长为258 nm,柱温为30℃,进样量为20μl。结果:HPLC法测定的腺苷的线性范围为6.545 3~5.365 0×10~3ng·ml^(-1),r=0.999 9,最低检测限为4.491 4 ng·ml^(-1),回收率为100.4%(RSD=1.47%,n=9);5’-腺苷酸的线性范围为8.685 6~5.170 0×10~3ng·ml^(-1),r=1.000 0,最低检测限为3.42 ng·ml^(-1),回收率为98.6%(RSD=2.76%,n=9)。结论:采用HPLC法测定注射用环磷腺苷的有关物质腺苷、5’-腺苷酸、方法简便,结果可靠。

Objective： To determine the related substances in adenosine cyclophosphate for injection, and determine the contents of adenosine and 5＇- adenosine by HPLC as well. Methods： The column was a Waters XBridge BP C18 column （250 mm×4.6 mm, 5 μm）. The mobile phase was monopotassium phosphate and tetrabutylammonium bromide buffer （monopotassium phosphate 6.8 g, tet- rabutylammonium bromide 3.2 g, both were dissolved in 1 000 ml water, and pH value was adjusted to 4.3 by phosphoric acid）- ace- tonitrile（90 ： 10）. The flow rate was 1.0 ml·min^-1, the determination wavelength was 258 nm, the column temperature was 30℃ and the injection volume was 20μl. Results： The linear range of adenosine was 6. 545 3-5. 365 0×10^3 ng·ml^-1 （r =0. 999 9）, the limit of detection was 4. 491 4 ng·ml^-1, and the recovery was 100.4% （ RSD = 1.47%, n = 9）. The linear range of 5＇- adenosine was 8. 685 6-5. 170 0 × 10^3 ng·ml^-1 （ r = 1. 000 0）, the limit of detection was 3.42 ng·ml^-1, and the recovery was 98.6% （ RSD = 2.76%, n = 9）. Conclusion： The determination method for the related substances in adenosine cyclophosphate for Injection and the known impurities of adenosine and 5＇- adenosine by HPLC is easy to operate and the determination results are accurate.