摘要
目的 建立离子对反相高效液相色谱（RP-HPLC）同时检测小鼠海马组织中ATP、ADP和AMP含量的方法。方法 高氯酸沉淀蛋白后,碳酸钾-甲醇溶液中和,-20℃冷冻保存样本;流动相：50 mmol/L磷酸盐缓冲液（缓冲液为K2HPO4-KH2PO4,pH=6.60,含22%甲醇和4 mmol/L四丁基硫酸氢铵）。C_（18）色谱柱（4.6 mm×250 mm,5μm）及相同填充材料的预柱,测定小鼠海马组织中ATP、ADP和AMP的含量;检测波长254 nm,流速0.6 ml/min;柱温为室温;进样量20μl。结果 本法的日内和日间精密度分别为1.27%~3.42%和0.88%~3.52%,回收率为95.67%~104.05%。结论 本研究建立的RP-HPLC法可同时测定小鼠海马ATP、ADP和AMP的含量,方法稳定、准确、可靠。
Objective To establish an ion-pair reverse-phased high-performance liquid chromatography（RP-HPLC）method for simultaneous determination of ATP,ADP and AMP in the hippocampus of mice. Methods The protein of mouse hippocampus was precipitated with perchloric acid,and neutralized with potassium carbonate-methanol mixture. Mobile phase was as follows：50mmol/L phosphate buffer（buffer for K_2HPO_4-KH_2PO_4,pH 6.60,containing 22% methanol,and 4 mmol/L tetrabutylammonium bisulfate）. Shimadzu HPLC system and Agilent C18column（4.6 mm×250 mm,5 μm）filled with the same material pre-column（12.5 mm×4.6 mm,5 μm）were used. The contents of ATP,ADP and AMP in mouse hippocampus were analyzed at a wavelength of 254 nm,the flow rate of 0.6 ml/min and room column temperature. Results Stability tests showed that intra-day and inter-day precision of the method were 1.27%-3.42% and 0.88%-3.52%,respectively,and recovery rates were 95.67%-104.05%. Conclusion The HPLC method established in this study is simple,accurate and efficient in detecting the levels of ATP,ADP,and AMP in mice hippocampus.
出处
《国际药学研究杂志》
CAS
CSCD
北大核心
2016年第6期1151-1155,共5页
Journal of International Pharmaceutical Research
基金
国家重点研发计划重点专项资助项目(2016YFC1306301)
