荧光素钠与牛血清蛋白相互作用的荧光光谱研究及其分析应用  

Studies on the Interaction between Sodium Fluorescein and Bovine Serum Albumin by Fluorescence Spectroscopy and Its Analytical Application

作　　者：孙洋[1] 樊君[1] 胡晓云[2] 刘璐莎[1] 胡春梅[1] 尹辰[1] 魏嵩[1] 

机构地区：[1]西北大学化工学院,西安710069 [2]西北大学物理系,西安710069

出　　处：《化学学报》2011年第8期937-944,共8页Acta Chimica Sinica

基　　金：陕西省科技计划(No.2010TG-37);西安市工业应用技术研发(No.CXY09023)资助项目

摘　　要：采用荧光光谱研究了荧光素钠与牛血清蛋白(BSA)间的相互作用,根据荧光淬灭相关方程分别计算了淬灭速率常数、结合常数、结合位点数及热力学参数,确定了BSA对荧光素钠的淬灭机理及作用方式,根据能量转移理论求得荧光素钠与BSA的结合距离及能量转移率,结合三维、同步荧光光谱研究了荧光素钠对BSA构象的影响;在一定范围内荧光素钠的淬灭程度与BSA浓度成正比,据此建立一种荧光素钠测定蛋白的方法,线性范围为0.15×10-7～15×10-7 mol·L–1,方法具有高灵敏度,检测极限为0.146×10-8 mol·L–1,文中还考察了不同pH值和干扰物质对于测定结果的影响,用于人血清中总蛋白含量测定结果与考马斯亮蓝法基本一致.The interactions between sodium fluorescein（SoF） and bovine serum albumin（BSA） were in-vestigated by fluorescence spectroscopy.According to the Stern-Volmer equation,the association constants,quenching constants and the numbers of binding sites were obtained.It is proved that the fluorescence quenching of SoF by BSA was a result of the formation of SoF-BSA complex.The thermodynamic parame-ters for the reaction were calculated according to van＇t Hoff equation.The distance and energy transfer effi-ciency between SoF and BSA were obtained according to the theory of non-radioactive energy transfer.The effect of SoF on the conformation of BSA was also analyzed using 3D fluorescence spectroscopy and syn-chronous fluorescence spectroscopy.The fluorescence intensity of SoF-BSA complex was proportional to the concentration of BSA,based on which,a new quantitative assay of protein was presented.The linear range was 0.15×10-7～15×10-7 mol·L–1,and the sensitivity of the method was high with detection limit of 0.146×10-8 mol·L-1.The effects of pH and interfering substance on the detection were also investigated.The results indicated that the most of the water-soluble amino acids,metal ion and antioxidant do not inter-fer or only interfer slightly under the permission of ±5.0% relative error,whereas SDS,Lecithin,APG,Cu2＋,Fe3＋,Sucrose and Tryptophane produced obvious interference.Determination of proteins in human serum by this method gave results which were very close to those obtained by Coomassie Brilliant Blue col-orimetry.

关 键 词：荧光素钠 牛血清蛋白 荧光光谱 定量分析 

分 类 号：Q51[生物学—生物化学]