固相萃取-高效液相色谱荧光检测法测定人血浆3-硝基酪氨酸含量  

Determination of 3-Nitrotyrosine in Human Plasma by Solid Phase Extraction-High Performance Liquid Chromatography with Fluorescence Detection

作　　者：郭帅[1] 邵丽华[1] 荣琳[1] 王希峰[1] 郗德凤[1] 

机构地区：[1]山东大学公共卫生学院,济南250012

出　　处：《分析化学》2009年第2期242-246,共5页Chinese Journal of Analytical Chemistry

基　　金：山东省自然科学基金(No.Y2007C079)资助项目

摘　　要：建立了人血浆中痕量3-硝基酪氨酸的高效液相色谱-荧光检测法。血浆经乙腈沉淀蛋白后,上清液用氮气吹干,残渣复溶后过C18固相萃取小柱净化浓缩,洗脱液再用氮气吹干,残渣进行荧光衍生化反应。采用高效液相色谱分离,荧光检测器检测,外标法定量。3-硝基酪氨酸在0.50～50.0nmol/L范围内线性关系良好,相关系数为0.9999;样品加标绝对回收率为89.3%～91.9%,相对回收率为99.2%～107.9%;日内相对标准偏差为1.46%～4.79%,日间相对标准偏差为2.75%～9.20%;在血浆中检出限为0.25nmol/L;测定正常人血浆中3-硝基酪氨酸平均浓度为1.18nmol/L(n=21),Ⅱ型糖尿病病人血浆中3-NT平均浓度为3.63nmol/L(n=23)。本方法灵敏度高,重现性好,适用于临床研究中大样本量的测定。A high performance liquid chromatography with fluorescence detection method has been developed for the determination of trace levels of 3-nitrotyrosine in human plasma. After precipitation of plasma proteins by acetonitrile and solid phase extraction, the derivatization reaction was carried out. The reaction mixture was determined by high performance liquid chromatography with fluorescence detection and quantified by external standard method. The coefficient correlation was 0. 9999 in the linear range of 0.50- 50.0 nmoL/L. The average absolute and relative recoveries in spiked sample were 89.3% - 91.9% and 99.2% - 107.9%. The intra-day and inter-day relative standard deviations were 1.5% - 4.8% and 2.8% - 9.2%, respectively. The detection limit was 0.25 nmoL/L. The average concentrations of 3-nitrotyrosine in normal human plasma （n =21 ） and patients with non-insulin dependent diabetes （n = 23） were 1.18 and 3.63 nmol/L, respectively, The current method is sensitive, accurate, reproducible and suitable for analysis of large sample numbers in clinic research.

关 键 词：3-硝基酪氨酸 4-氟-7-硝基-2 1 3-苯并氧杂恶二唑 人血浆 荧光检测 固相萃取 高效液相色谱 

分 类 号：R446.1[医药卫生—诊断学]