液相色谱法测定小鼠脑组织中ATP、ADP及AMP含量  

High performance liquid chromatographic determination ofadenosine phosphates in the mouse brain

作　　者：吴燕川[1] 李尧华[2] 谢胜男[2] 李昕[2] 于顺[2] 何士大[1] 

机构地区：[1]首都医科大学宣武医院中心实验室神经变性病教育部重点实验室,北京100053 [2]首都医科大学宣武医院神经生物学研究室,北京100053

出　　处：《首都医科大学学报》2011年第4期534-537,共4页Journal of Capital Medical University

基　　金：北京市自然科学基金重大项目(7050001);北京市属高等学校人才强教计划资助项目(PHR200907113)~~

摘　　要：目的建立小鼠脑组织中腺苷酸含量测定的高效液相色谱(high performance liquid chromatogram,HPLC)-可变波长检测法。方法应用酸沉淀蛋白、Agilent1200高效液相色谱系统、Supelco Discovery C18(150 mm×4.6 mm,5μm)反相色谱柱及相同填充材料的预柱,对小鼠脑组织样品中腺苷酸进行分离分析,波长为254 nm,流动相为30 mmol/L K2HPO4-KH2PO4(含5%甲醇),pH5.85,流速为1.0 mL/min,进样体积为20μL,柱温为30℃。结果腺苷酸ATP、ADP及AMP在10～100 mg/L范围,线性关系良好,线性方程为ATP:A=28.924C+14.144,γ=0.999 90;ADP:A=31.504C-12.608,γ=0.999 92;AMP:A=46.539C-5.255,γ=0.999 95。平均回收率为95.3%～103.2%;日内和日间相对标准差(RSD)分别为1.14%～1.75%、2.45%～3.97%。结论高效液相色谱-可变波长检测法可以简便、准确、有效地测定急性重复低氧小鼠脑组织中ATP、ADP和AMP的含量。Objective To establish a method for the quantitative determination of adenosine phosphates in mouse brain by high performance liquid chromatography-variable wavelength detector（HPLC-VWD）.Methods The protein in mouse brain tissue sample was precipitated with perchloric acid.Separation was performed on a reverse phase Supelco Discovery C18 column（150 mm×4.6 mm,5 μm）,equipped with a pre-column of the same material by Agilent 1200 HPLC-VWD.The wavelength was 254 nm.The mobile phase contained methanol-30 mmol/L potassium dihydrogen phosphate buffer,pH 5.85（5∶ 95,v/v）.The flow rate was set at 1.0 mL/min and the injection volume was 20 μL.The column temperature was 30 ℃.Results The linear relationship between the range（10～100） mg/L was good.The linear equations were ATP： A=28.924C＋14.144,γ=0.999 90;ADP： A=31.504C-12.608,γ=0.999 92;AMP： A=46.539C-5.255,γ=0.999 95.The average recovery rate was 95.3%～103.2%.The RSDs of intra-and interday were 1.14%～1.75% and 2.45%～3.97%,respectively.Conclusion The experimental data proved that the HPLC-VWD method could detect the concentration of ATP,ADP and AMP in the mouse brain on repeated hypoxic exposures accurately,rapidly and simply.

关 键 词：腺苷酸 高效液相色谱法 脑组织 小鼠 

分 类 号：R-3[医药卫生]