超声对豌豆分离蛋白结构及乳化性能的调控效应

doi:10.3864/j.issn.0578-1752.2021.22.015

摘要/Abstract

摘要：

【目的】考察超声波处理对豌豆分离蛋白（pea protein isolate,PPI）结构和理化性质的影响,揭示超声处理对PPI乳化特性的调控机制,为豌豆蛋白作为天然乳剂及其相关产品在食品领域中的应用提供理论依据。【方法】选用频率为20 kHz、功率为600 W的超声波经不同时长（0、20、30、40和60 min）的预处理后制备改性豌豆蛋白（ultrasonic-pea protein isolate,U-PPI）,再经高压均质制备U-PPI乳液。通过自由氨基、总巯基、粒径、溶解度及SDS-PAGE探究超声波处理对豌豆蛋白理化性质的影响;借助圆二色谱仪分析U-PPI二级结构的变化;通过内源性色氨酸荧光测定分析U-PPI三级结构的变化;通过乳化活性指数、粒径、乳液界面蛋白分布、Zeta电位和表观黏度表征U-PPI的乳化能力和乳液稳定性;借助激光共聚焦荧光显微镜观察乳状液的微观结构。【结果】超声波处理对PPI结构具有显著修饰作用,30—40 min的短时间超声处理能够显著降低α-螺旋并提高β-折叠含量,使PPI的结构更加舒展柔韧,更多的疏水基团暴露在界面上,同时超声波的解聚效应还引起PPI的平均粒径减小、溶解度显著增大;因而在此条件下超声处理对PPI结构的修饰有利于其在油/水界面形成致密而稳定的蛋白膜,有效地提高了PPI的乳化活性和乳状液的稳定性,微观结构也显示其乳液粒径更小、分布更加均匀。然而,60 min的长时间超声处理会导致PPI的疏水重聚,溶解度降低,不利于其在油/水界面的吸附重排,降低了其乳化活性和乳液稳定性。【结论】30和40 min超声处理产生的空化效应、机械效应等对PPI具有显著的解聚作用,促使蛋白分子结构舒展,有利于其在油/水界面的吸附重排,从而显著改善了豌豆蛋白的乳化性能。

关键词: 超声波改性, 豌豆分离蛋白, 结构修饰, 乳液稳定性

Abstract:

【Objective】 This study was designed to investigate the effects of ultrasonic treatment on the structure and physicochemical properties of pea protein isolate (PPI), and to explore the regulation mechanism of ultrasonic treatment on the emulsifying characteristics of PPI, so as to provide the theoretical basis for the application of PPI as natural emulsions or related products in the food field. 【Method】 The ultrasonic-pea protein isolate (U-PPI) was prepared by ultrasonic wave with frequency of 20 kHz and power of 600 W under different times (0, 20, 30, 40 and 60 min), and then U-PPI emulsions were prepared by high pressure homogenization. The effects of ultrasonic treatment on the physicochemical properties of PPI were investigated by the tests of free amino group, total sulfhydryl group, particle size, solubility and SDS-PAGE. The changes in secondary and tertiary structure of U-PPI were analyzed by circular dichroism and intrinsic tryptophan fluorescence, respectively. The emulsifying ability and emulsion stability of U-PPI were characterized by emulsion activity index, particle size, protein distribution at the interface of emulsion, Zeta potential and apparent viscosity. The microstructure of the emulsions was observed by using laser confocal fluorescence microscopy. 【Result】 Ultrasonic treatment significantly modified the structure of PPI, i.e., a short time (30 to 40 min) ultrasonic treatment significantly reduced the α-helix content and improved the β-sheet content of PPI, which made the structure of the PPI more flexible and more hydrophobic groups exposed to the interface. Simultaneously, the depolymerization effect of ultrasonic also caused the decrease of the average particle size and the significant increase of the solubility of PPI. Therefore, the modification of PPI by a short time ultrasonic treatment was conducive to the formation of a dense and stable protein film at the oil/water interface, which effectively improved the emulsifying activity of PPI and the stability of the emulsions, and the microstructure of the emulsions also showed that the particle sizes of emulsions stabilized with U-PPI were smaller, and the droplet distribution was more uniform. However, a long time (60 min) ultrasonic treatment caused the hydrophobic repolymerization and insolubility of PPI, which was not conducive to the adsorption and rearrangement of PPI in the oil/water interface and thus reduced the emulsifying activity and emulsion stability. 【Conclusion】 The cavitation effect and mechanical effect produced by a short time (30, 40 min) ultrasonic treatment had significant depolymerization effect on PPI, which promoted the protein structure more flexible and was beneficial to the adsorption and rearrangement of PPI at the oil/water interface, and thus significantly improved the emulsifying properties of PPI.

Key words: ultrasonic modification, pea protein isolate, structural modification, emulsion stability

李朝蕊,韩馨蕊,范鑫,黄峻榕,曹云刚,熊幼翎. 超声对豌豆分离蛋白结构及乳化性能的调控效应[J]. 中国农业科学, 2021, 54(22): 4894-4905.

LI ZhaoRui,HAN XinRui,FAN Xin,HUANG JunRong,CAO YunGang,XIONG YouLing. Regulation Effects of Ultrasound on the Structure and Emulsification Properties of Pea Protein Isolate[J]. Scientia Agricultura Sinica, 2021, 54(22): 4894-4905.

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时间 Time (min)	水相蛋白 Aqueous protein (%)	界面蛋白 Interface protein (%)	分配系数 Partition coefficient
Con	38.0±0.0e	62.0±0.0e	14.75
20	29.7±1.2b	70.3±2.1b	21.63
30	27.3±1.2a	72.7±1.5a	23.95
40	31.7±0.6c	68.3±2.1c	19.51
60	33.0±0.0d	67.0±0.0d	18.19