改良2,4-二硝基苯肼法测定脑内源甲醛  

Assay of Brain Endogenous Formaldehyde With 2,4-Dinitrophenylhydrazine Through UV-HPLC

作　　者：苏涛[1,2] 魏艳[1] 赫荣乔[1] 

机构地区：[1]中国科学院生物物理研究所,脑与认知科学国家重点实验室,北京100101 [2]中国科学院研究生院,北京100049

出　　处：《生物化学与生物物理进展》2011年第12期1171-1177,共7页Progress In Biochemistry and Biophysics

基　　金：国家重点基础研究发展计划资助项目(2010CB912303);国家重点基础研究发展计划(973)(2012CB911004);中国科学院创新方向项目(CAS-KSCX2-YW-R-119)及创新预研项目(KSCX2-YW-R-256);国家自然科学基金面上项目(30970695);强生集团科技公司基金(GJHZ1131)资助项目~~

摘　　要：内源甲醛代谢失衡所造成的脑慢性损伤被认为是衰老过程中记忆丢失的危险因素之一,因此,有必要准确测定脑内不同区域甲醛的含量为相关研究提供参考.作者通过2,4-二硝基苯肼(2,4-DNPH)偶联高效液相色谱(UV-HPLC),测定了家猪脑额叶、顶叶、颞叶、枕叶、海马、小脑、脑干内源甲醛含量(75.5～83.4μmol/kg).采用10%三氯乙酸处理脑组织匀浆(pH=1.0),不但可以避免蛋白质及其他分子的干扰,还可以省略现有方法中的萃取步骤,且提高了测试的灵敏度(P<0.05).该方法的回收率约95.96%～102.04%,相对标准偏差(n=5)小于10%.结果表明,改良2,4-DNPH法用于脑内源甲醛的测定,其操作简便、可信度高.Chronic impairments of brain caused by accumulation of endogenous formaldehyde as ageing is thought as one of the risk factors for sporadic age-related dementia.Thus,it is necessary to determine the endogenous concentration of formaldehyde in brain.Using 2,4-dinitrophenylhydrazine（2,4-DNPH） in the assay, the concentrations of formaldehyde in parietal lobe,frontal lobe,temporal lobe,occipital lobe,hippocampus, cerebellum and brainstem of porcine brain（n=5） were measured through UV-HPLC.Homogenizing brain tissues with 10% trichloroacetic acid（pH =1.0） not only avoided the disturbance from contaminated protein and other compounds,but also eliminated the step extraction of FA-DNPH derivative（2,4-dinitrophenylhydrazone） and improve the methodological sensitivity（P〈 0.05）.The porcine brain formaldehyde was 75.5～83.4 μmol/kg fresh brain tissues measured by this improved method.Recoveries of spiked formaldehyde at low and high level were 95.96%～102.04% with relative standard deviations less than 10%.This suggests that the modified 2,4-DNPH method is effective enough to measure the endogenous formaldehyde in brain tissues.

关 键 词：定量分析 内源甲醛 2 4-二硝基苯肼 高效液相色谱 猪脑 认知功能损伤 

分 类 号：Q42[生物学—神经生物学] O65[生物学—生理学]