尼古丁与BSA相互作用的光谱研究

用紫外-可见光谱和荧光光谱研究了尼古丁与牛血清白蛋白(bovine serum albumin, BSA)的相互作用。荧光研究表明，尼古丁浓度的增加引起BSA 345 nm处荧光有规律地猝灭。Stern-Volmer 方程分析pH 5.0，pH 7.4和pH 11.0体系的荧光猝灭机理发现，pH 5.0体系属动态猝灭，而pH 7.4和pH 11.0体系为静态猝灭。Lineweaver-Burk双倒数方程计算pH 7.4和pH 11.0体系在温度为20和37 ℃条件下尼古丁和BSA的结合常数k分别为：k_{20 ℃}=140.15 L·mol-1 ，k_{37 ℃}=131.83 L·mol-1 (pH 7.4)和k_{20 ℃}=141.76 L·mol-1，k_{37 ℃}=27.79 L·mol-1(pH 11.0)，表明结合常数在pH 7.4条件下受温度的影响要比pH 11.0条件下小，推测是由于不同pH下尼古丁存在的不同形态所致。紫外-可见光谱研究表明，pH 7.4条件下尼古丁浓度的增加引BSA在210 nm处吸收峰吸收强度减小且红移，说明BSA二级结构发生变化，即螺旋结构变松散;紫外二阶导数光谱和同步荧光光谱(Δλ=λ_em-λ_ex=15 nm和Δλ=λ_<i>em-λ_ex=60 nm)分析尼古丁对BSA芳香性氨基酸(Trp, Tyr和Phe)残基微环境的变化，结果表明高浓度的尼古丁使所有这些芳香性氨基酸残基微环境由疏水环境转变为亲水环境。

Spectroscopic Studies on the Interaction of Nicotine and BSA

The interaction of nicotine and bovine serum albumin(BSA) was investigated by fluorescence spectra and UV-vis spectra. The fluorescence spectrum showed that BSA fluorescence quench regularly with the addition of nicotine.The fluorescence quenching mechanisms were also studied in pH 5.0, pH 7.4 and pH 11.0 by Stern-Volmer equation, indicating dynamic quenching(pH 5.0) and static quenching(pH 7.4 and pH 11.0) respectively. Association constants (k) of nicotine and BSA at pH 7.4 and pH 11.0 at the temperatures of 20 and 37 degrees C were given by the Lineweaver-Buck equation, which are: k(20 degrees C) = 140.15 L x mol(-1) and k(37 degrees C) = 131.83 mol x L(-1) (pH 7.4), and k(20 degrees C) = 141.76 mol x L(-1), k(37 degrees C) = 27.79 mol x L(-1) (pH 11.0), suggesting that the association constant is effected by the temperature much more remarkably at pH 7.4 than that at pH 11.0 because of the different states of nicotine at different pHs. The UV-Vis spectra exhibit that the absorbance of BSA(210 nm) shifts to red and decreases gradually with the addition of nicotine, reflecting the transition of secondary structure of BSA, namely, the helix of BSA becomes looser. The UV-Vis second derivative spectra and synchronous spectra (delta wavelength = wavelength(em) - wavelength(ex) = 15 nm and delta wavelength = wavelength(em) - wavelength(ex) = 60 nm) imply the change of the microcircumstance of aromatic amino residues of BSA(Trp, Tyr and Phe) from hydrophobicity to hydrophilicity at high concentration of nicotine.