HPLC法检测人红细胞膜总胆固醇水平及其临床应用  

Determination of cholesterol in erythrocyte membranes by HPLC method and its clinical application

作　　者：李克[1] 吴龙琴[1] 曹璐颖[1] 蔡加炉 牛冬梅[1] 汪俊军[1] 

机构地区：[1]南京军区南京总医院检验科,210002

出　　处：《中华检验医学杂志》2014年第3期179-183,共5页Chinese Journal of Laboratory Medicine

摘　　要：目的 建立人红细胞膜胆固醇高效液相色谱测定方法.方法 实验方法学研究.选择2012年9月至2013年2月间因胸闷、胸痛在南京军区南京总医院住院并行选择性冠状动脉造影患者167例分为3组：稳定性心绞痛（SAP）组76例、急性冠状动脉综合征（ACS）组46例和对照（Control）组45例.采集红细胞样品经溶血和洗涤后,在70℃氢氧化钾溶液中皂化反应.样品经正己烷-异丙醇混合溶剂提取后,以Lichrospher色谱柱为固定相,乙腈～异丙醇为流动相进行色谱等度分离,208 nm波长下定量检测.检测3组研究对象的红细胞膜胆固醇水平.多组间均数比较用Oneway ANOVA检验;相关性比较采用Spearman相关性分析.结果 在所建立的分析条件下,胆固醇色谱保留时间约为6.1 min,峰形清晰对称,与样品中其余内源性物质分离完全,定量准确.胆固醇进样量在（0.05 ～2.00）μg范围与峰面积呈良好线性关系,对红细胞膜标本检测结果与胆固醇酶法检测试剂盒测定值相关良好.Control组、SAP组以及ACS组红细胞膜胆固醇水平分别为：（87.0μg/mg,75.4 ～98.9 μg/mg）、（92.9 μg/mg,83.8 ～ 109.0 μg/mg）和（173.9 μg/mg,140.0～188.8 μg/mg）.统计学分析显示,ACS组与Control组和SAP组相比,CEM水平明显增高,差异均有统计学意义（t值分别为-14.44和-14.68,P均＜0.01）;SAP组与Control组相比,CEM水平有增高,但差异无统计学意义（t=-1.88,P＞0.05）.结论 成功建立了一种检测红细胞膜总胆固醇含量的方法.灵敏度高、特异、重复性较好.Objective To develop a high performance liquid chromatographic method （HPLC） for the analysis of of cholesterol in erythrocyte membranes.Methods The study included 167 consecutive chest pain patients who underwent coronary artery angiography in the Department of Cardiology,Nanjing General Hospital of Nanjing Command between September 2012 and February 2013.According to the clinical symptoms and t angiographic results,patients were divided into three groups：acute coronary syndrome （ACS） group （n =46）,stable angina pectoris （SAP） group （n =76） and the control group （n =45）.After the erythrocyte sample was hypotonically lysed and washed,saponification was carried out in a polassium hydroxide solution at 70 ℃.After extraction by Hexane/isopropanol mixture,the sample was separated on a Lichrospher column and detected by ultraviolet absorbance at 208 nm.A mobile phase composed of acetonitrile-isopropyl alcohol was found to be the most suitable for this separation.Concentrations of cholesterol in erythrocyte membranes were tested.Analysis of variance with covariates （ANOVA） was used to evaluate differences in CEM levels among groups.The relationship between continuous variables was evaluated by Spearman＇s correlation coefficient.Results Under the chromatographic conditions described,retention time of the cholesterol was approximately 6.1 min.Good separation and detectability of cholesterol in erythrocyte membranes were obtained.The method proved to be linear in the injection range of cholesterol from 0.05 g to 2.00 g.Cholesterol content in erythrocyte membranes were （87.0 μg/mg,75.4-98.9 μg/mg）,（92.9 μg/mg,83.8-109.0 μg/mg） and （173.9 μg/mg,140.0-188.8 μ g/mg） in the control,SAP and ACS groups,respectively.Cholesterol content in erythrocyte membranes was significantly higher in ACS group than that in SAP and control groups （P 〈 0.01）.Conclusion We have successfully developed a method for the determination of cholesterol in erythrocyte membranes with good sensitiv

关 键 词：红细胞膜 胆固醇 色谱法 高压液相 急性冠状动脉综合征 动脉粥样硬化 

分 类 号：R446.1[医药卫生—诊断学] O657.72[医药卫生—临床医学]