低温等离子体关键活性粒子对大豆球蛋白结构及致敏性的影响

Effects of the Free Radicals in Cold Plasma Treatment on the Structure and Allergenicity of Glycinin

摘要:

该文通过构建单独活性粒子（•OH、H2O2）氧化体系和基于低温等离子体技术（CP）的活性粒子屏蔽体系（屏蔽•OH：叔丁醇；屏蔽H2O2 ：MnO2）的对比研究，探究CP活性粒子（•OH、H2O2）对大豆球蛋白结构及致敏性影响的特异性。结果表明：单独•OH氧化体系，大豆球蛋白与IgG/IgE 结合能力均随•OH含量增加而逐渐降低至69.00%（20 mmol/L）；CP+叔丁醇的•OH屏蔽氧化体系中，结合能力由59.45%（IgG）、52.50 %（IgE）（40 kV、2 min），分别增加至67.95%（IgG）（100 mmol/L）和73.77%（IgE）（300 mmol/L）。单独H2O2 氧化体系，大豆球蛋白与IgG/IgE结合能力均随H2O2 含量增加而降至65.65%（IgG）、30.84%（IgE）（2.0 mmol/L），基于CP+MnO2 的H2O2 屏蔽体系中，随着MnO2 添加量的增加，结合能力分别增加至91.22%（IgG）、73.90%（IgE）（25 mmol/L）。利用SDS-PAGE 和内源荧光光谱分别表征蛋白一级结构和三级结构变化，结果表明，•OH通过改变大豆球蛋白一级和三级结构，而H2O2 改变蛋白三级结构达到消减大豆球蛋白致敏性目的。该研究证实•OH和H2O2 作为具有氧化作用的活性粒子参与CP处理对蛋白结构的氧化修饰。

Abstract:

A comparative investigation was performed using oxidation systems comprising single active particles, namely •OH and H2O2, and free radical scavenging systems based on cold plasma (CP) (•OH scavenging: CP + tertiary butanol; H2O2 scavenging: CP+MnO2) to explore the specificity of different active particles in CP treatment (•OH and H2O2) on the structure and antigenicity of glycinin. The IgG/IgE binding abilities of glycinin were found to gradually decrease to 69.00% as the •OH concentration increased up to 20 mmol/L in the •OH oxidation system. In the CP + tertiary butanol oxidation system without •OH, the IgG/IgE binding abilities of glycinin increased from 59.45% (IgG) and 52.50% (IgE) with CP treatment (40 kV, 2 min) alone to 67.95% (IgG) (100 mmol/L tertiary butanol) and 73.77% (IgE) (300 mmol/L tertiary butanol), respectively. Similarly, the IgG /IgE binding abilities of glycinin decreased to 65.65% (IgG) and 30.84% (IgE) as the H2O2 concentration increased to 2.0 mmol/L in the H2O2 oxidation system. In the CP + MnO2 oxidation system without H2O2, the binding abilities of glycinin increased to 91.22% (IgG) and 73.90% (IgE) as the MnO2 concentration increased to 25 mmol/L. Structural analysis based on SDS-PAGE and fluorescence spectra indicate that the •OH-induced primary and tertiary structure changes of glycinin are directly related to the reduction of the antigenicity of glycinin. Meanwhile, only the tertiary structure of glycinin was modified by H2O2. Therefore, •OH and H2O2 are key active particles in CP treatment for the oxidative modification of protein structures and its related antigenicity elimination.