用于传统书画修复的不同加热温度豆浆水中大豆蛋白的疏水性分析

Hydrophobicity Analysis of Soy Protein in Soybean Water with Different Heating Temperatures during Painting and Calligraphy Restoration

豆浆水是中国传统书画修复中的施胶材料,加热可导致大豆蛋白产生热变性,使纸张产生不同程度的疏水性,从而影响到书画修复的效果。本研究利用8-苯胺基-1-萘磺酸(ANS)荧光探针法分析大豆蛋白在加热过程中疏水性的变化;利用十二烷基硫酸钠-聚丙烯酰胺凝胶电泳(SDS-PAGE)分析加热前后的蛋白质差异条带,并以液相色谱-串联质谱(LC-MS/MS)进行蛋白鉴别;衰减全反射傅里叶变换红外光谱法(ATRFTIR)去卷积、二阶导数曲线拟合分析不同加热温度豆浆水大豆蛋白二级结构的变化。ANS荧光探针实验结果表明,80℃的豆浆水的疏水性最大。SDS-PAGE与LC-MS/MS分析表明,加热消失的两个条带的蛋白分别为球蛋白Glycinin G3亚基及糖蛋白凝集素Lectin亚基; ATR-FTIR分析表明,随着温度从25℃上升到80℃,大豆蛋白β-片层结构含量由41.14%增加到48.87%,α-螺旋、β-转角和无规卷曲结构向β-片层结构转变,而β-片层结构含量与表面疏水性呈正相关,加热温度为80℃的豆浆水最适于书画修复。

Soybean water is a common sizing agent that is used in the restoration of Chinese paintings and calligraphies.Heating soybean water leads to denaturation of soy protein and thus alters the hydrophobicity of paper after sizing.In this work,the change of surface hydrophobicity of soybean protein in the heating process was analyzed by 8-aniline-1-naphthlene sulfonic acid(ANS)fluorescence probe.Sodium dodecyl sulfate polyacrylamide gel electrophoresis(SDS-PAGE)was adopted to sort out different protein contents which later were identified by LC-MS/MS.The self-deconvolution and second derivative curve fitting techniques of attenuated total reflectance-Fourier transform infrared spectroscopy(ATR-FTIR)were used to analyze the change of secondary structures of soybean protein at different heating temperatures.ANS fluorescence probe experiment showed that the soybean water achieved its largest hydrophobicity when the temperature was 80℃.The results of SDS-PAGE and LC-MS/MS revealed the disappearance of globulin glycinin G3 and glycoprotein lectin at high temperature.ATR-FTIR analysis showed that the increase of heating temperature of soybean water raisedβ-sheet content from 41.14%to 48.87%and the originalα-helix,β-turn andγ-random structures shifted toβ-sheet structure.There was a positive correlation between the structural content ofβ-sheet and the surface hydrophobicity.The research showed that the heating temperature of soybean milk at 80℃was suitable for calligraphy and painting restoration.