探究了拉曼光谱应用于黑豆蛋白结构变化研究的可行性, 研究了黑豆蛋白溶液在低频超声处理在不同超声强度、 不同处理时间下的结构变化, 并进行了热力学特性分析。 低、 中超声处理强度下TD的降低表明蛋白质分子的内部疏水作用被破坏, 使黑豆蛋白不稳定的聚集体解聚为小分子可溶性聚集物, 而在高超声处理强度下TD的增高表明聚集体重聚。 拉曼光谱分析表明超声处理下除了E样品(300 W, 24 min)所有黑豆蛋白均发生了α-螺旋结构含量降低和β-折叠结构含量增高。 聚集体的聚合/解聚导致黑豆蛋白二级结构的重组, 尤其是β-折叠。 超声处理使拉曼光谱在760 cm-1的色氨酸归属谱线强度降低表明超声处理使黑豆蛋白发生了部分的解折叠。 超声处理下酪氨酸归属谱线强度变化不显著, 表明超声处理并未显著改变黑豆蛋白酪氨酸的微环境。 1 450 cm-1拉曼归属谱带随着超声处理强度和时间的增加而增大, 但随着功率及处理时间的进一步增大此值有所降低。 在超声处理下聚集体的形成使二硫键的g-g-t构型转变为t-g-t构型。 尽管黑豆蛋白聚集体重组的机理仍有待研究, 但拉曼光谱是一种研究超声处理黑豆蛋白结构变化的可行方法, 也可为蛋白质结构研究提供一种新的研究思路。

This article focused on the assessment of the potential of Raman spectroscopy for the determination of structural changes in black-bean protein isolate (BBPI) dispersions with low-frequency (20 kHz) ultrasonication applied at various powers (150, 300 or 450 W) and for different durations (12 or 24 min). It also reported on differential scanning calorimetry analyses. A decrease in TD at low- and medium-power ultrasonication confirmed these ultrasonication treatment disrupted internal hydrophobic interactions of protein molecules and broke up unstable aggregates to smaller soluble protein aggregates, while an increase in TD at high-power was attributed to repolymerization of aggregates. Raman spectroscopy analysis revealed a decrease in the α-helix proportion and an increase in β-sheets after ultrasonic treatment except Sample E (300 W, 24 min). Transformation of aggregation results in a reconstruction in secondary structure of BBPI, especially in β-sheet structure. Ultrasonic-treatment induced a decrease in the normalized intensity of the Raman band near 760 cm-1 which indicated that Tryptophan residues tended to expose and also indicated protein partially unfolding. No significant difference was found in Tyr doublet ratios between unheated and ultrasound-treated BBPI indicated that ultrasound did not change the microenvironment around tyrosyl residues. While the intensity of 1 450 cm-1 band increased with increasing ultrasonic intensity and treatment time, and then decreased with further increase in power and treatment time. In general, the formation of aggregation transferred g-g-t conformation to t-g-t conformation. Though some mechanism of aggregation-repolymerization of BBPI remains to be clearly defined, Raman spectroscopy provide a feasible tool to study the structural changes of BBPI prepared under different ultrasonic conditions, give a new perspective to elucidation of protein structure.