脂质自由基AAPH体外氧化对富硒大豆分离蛋白性质和结构的影响

Effects of Peroxyl Radical Produced by in vitro Oxidation by Lipid Radical AAPH on Properties and Structure of Selenium-enriched SPI

本文采用2,2'-偶氮二(2-脒基丙烷)二盐酸(AAPH)在有氧条件下热分解制备过氧自由基,代表脂肪氧合酶诱导多不饱和脂肪酸脂质过氧化反应产生脂质自由基,研究AAPH体外氧化对富硒大豆分离蛋白性质和结构的影响。随着AAPH添加浓度的增加,FT-IR结果表明大豆分离蛋白(SPI)的α-螺旋、β-转角和无规卷曲结构增加,β-折叠结构相对减少;SDS-PAGE结果表明SPI的亚基在浓缩胶顶端逐步聚集;蛋白溶解性、游离巯基含量下降;表面疏水性、乳化性、乳化稳定性呈现先上升后下降趋势;內源荧光峰位(λmax)表现为逐渐蓝移,且荧光强度先增加后减少。富硒大豆分离蛋白(SSPI)在结构、亚基分布、性质等多项测试指标方面均优于普通大豆分离蛋白(OSPI),可能与SSPI中含有较高的硒含量,因而表现出更强的抗氧化性有关。

Peroxyl radicals were generated by the thermal decomposition of 2,2＇-azobis（2-amidinopropane） dihydrochloride（AAPH） under aerobic conditions in this study and served as a representative free radical derived from lipoxygenase-induced lipid peroxidation of polyunsaturated fatty acids. The effects of the peroxyl radical on the properties and secondary structure of selenium-enriched soy isolate protein（SPI） were investigated. With increasing concentrations of AAPH, the results of Fourier transform infrared spectroscopy showed that the α-helix, β-turn, and random coil content increased, while the β-sheet content decreased. Sodium dodecyl sulfate polyacrylamide gel electrophoresis（SDS-PAGE） patterns indicated that the subunits of SPI accumulated at the top of the stacking gel; protein solubility and the free sulfhydryl content of SPI decreased. Surface hydrophobicity, emulsifying activity, and emulsifying stability of SPI increased initially, but later decreased; the wavelength of the maximum emission（λmax） in the intrinsic fluorescence spectrum was gradually blue-shifted, and fluorescence intensity first increased then decreased. The selenium-enriched soy isolate protein was superior to ordinary soy isolate protein in terms of structure, distribution of subunits, properties, and other test indicators, which may be related to the high selenium content in selenium-enriched soy isolate protein, indicating its stronger antioxidant activity.