适宜物理-酶联合改性提高酸性条件下大豆分离蛋白乳化性

Physical-enzymatic modification methods improving emulsifying properties of soybean protein isolate under acidic conditions

为提高酸性条件下大豆分离蛋白(soy protein isolates,SPI)的乳化性能,该文研究了物理-酶联合改性对SPI(p H值为4)的乳化性能影响,通过对比确定了物理-酶联合改性,即超声波-酶复合改性和挤压膨化-酶复合改性两种改性方法在酸性条件下的乳化性能效果最好;并通过对改性后SPI(p H值为4)进行溶解性、游离巯基、二硫键、粒径、扫描电镜(scanning electron microscope,SEM)和激光共焦扫描显微镜(confocal laser scanning microscopy,CLSM)分析,从蛋白结构变化上进一步揭示了乳化性能提高现象的原因。结果表明:超声波联合植酸酶-酸性蛋白酶改性的SPI(Uphy-aci-SPI)的乳化活性(emulsifying activity index,EAI)为0.53 m2/g,比未改性SPI(0.18 m2/g)显著提高了196%(P<0.05),乳化稳定性(emulsifying stability index,ESI)为17 min,比未改性SPI(13.5 min)显著提高了25.9%(P<0.05);挤压膨化联合菠萝蛋白酶改性的SPI(Ebro-SPI)的EAI为0.46 m2/g,比未改性SPI显著增加了155%(P<0.05),ESI为17 min,比未改性SPI显著增加了25.9%(P<0.05)。在p H值为4的条件下对物理-酶联合改性的SPI的性质分析发现,物理-酶联合改性的SPI与未改性SPI相比,物理-酶联合改性的SPI的溶解性显著增加(P<0.05);物理-酶联合改性的SPI的乳状液平均粒径减小,CLSM观察乳状液中油与蛋白溶液稳定共融,改善了油滴之间的空间排斥力。物理-酶联合改性的SPI游离巯基的含量显著增加(P<0.05),二硫键含量显著降低(P<0.05)。SEM观察物理-酶联合改性的SPI为结构松散、破碎均一的微观结构。由此可见,乳化性能的提高是通过深层改变蛋白的结构来实现的。该研究可为探索提高酸性条件下SPI的乳化性能的方法提供理论依据。

Soybean protein isolated（SPI） is a kind of important raw material and widely used in food processing industry due to the excellent nutrition and functionality. Among all the properties towards SPI, emulsifying property is one of the most important functions. If adding SPI to foods, it can maintain emulsified state of system and improve the taste. However, because of the compact globular structure stabilized mainly by hydrogen bonds and disulfide bonds, native soy proteins have lower molecular flexibility and a rather poor emulsifying capability compared to other protein emulsifiers such as milk protein. Emulsifying performance of ordinary SPI is degraded obviously in acidic conditions, which limits its application in a few kinds of food（such as acidic beverages, salad dressings）. Therefore, an effective approach to improve emulsifying properties of SPI in acidic conditions has became a needed technology. In order to improve the emulsifying properties of SPI under acidic conditions, this paper mainly studied the effects of physical-enzymatic modification methods on the emulsifying properties of SPI. The comparison between different modification methods showed that the physical-enzymatic modification, also called ultrasound-enzymatic modification and extrusion-enzymatic modification, had the best emulsifying property under the acid condition of pH value of 4. The study used extrusion＇s characteristics, which produced the effect of shearing, extensional deformations, inertial effects and cavitations to destroy cell tissue and structure, and also used the ultrasonic＇s characteristics of rapidness and energy saving by producing the effect of cavitation, shearing and intense mixing in liquid with the dispersion effect to damage the cell wall. Micro-structure features of modified SPI at p H value of 4 were analyzed based on particle size distribution, circular dichroism spectroscopy, scanning electron microscope（SEM） and confocal laser scanning microscopy（CLSM）, and the influence of different modification methods on protein structure was also analyzed. Results were as follows： the emulsifying activity index（EAI） and emulsifying stability index（ESI） of the SPI modified by ultrasound combined with phytase-acidic protease separately reached 0.53 m^2/g and 17 min, which significantly increased by 196% and 25.9% compared to those of the original SPI condition（P〈0.05）. Likewise, the EAI and ESI of the SPI modified by extrusion combined with bromelain separately reached 0.46 m^2/g and 17 min, which increased by 155% and 25.9%（P〈0.05）. Compared with the original SPI at p H value of 4, the physical-enzymatic treatment significantly improved the solubility of SPI（P〈0.05） under the ultrasound combined with phytase-acidic protease modification and the extrusion combined with bromelain modification. The analysis of structure illuminated that compared with ordinary SPI, the SPI under physical-enzymatic modification methods could decrease average particle size. The CLSM showed that in emulsion, oil and protein solution were melting stably, which improved the spatial repulsion force between oil droplets. The number of free hydrosulfide thiols in tertiary structure of the SPI under physical-enzymatic modification was significantly increased（P〈0.05）, and the amount of disulfide bonds was significantly reduced（P〈0.05）. Under the observation of SEM, the microstructure of the SPI under physical-enzymatic modification was loose and crushing uniformly. Thus, the improvement of emulsifying property of the SPI was achieved by the deep change of protein structure. This research can provide references for the study on improving emulsifying properties of SPI under acidic conditions.