Mechanism Of The Aggregation And Gelation Of Soy Protein

Soy protein is one of the most used plant protein.It is contains various amino acid and has good functionality.Therefore,it is widely used in the nutritional formula,meat products,dairy products and so on as an additive.Aggregation and gelation of soy protein greatly influence the nutrition and functionality of the food.Numerous work has been done to investigate the aggregation and gelation of soy protein.However,most of the work focused only on a single protein concentration,pH value,ionic strength,temperature and time.If one the the parameters changed,it will probably lead to completely different aggregation and gelation.The study present here will systematically investigate how different factors influence the aggregation and gelation of soy protein and quantify these influences so that we can well predict,control and use the aggregation and gelation of soy protein.The main studies and the corresponding results are as the following:(1)Aqueous solutions of native soy glycinin was investigated at 20 ?,over a wide range of ionic strength(0-0.5 M),pH(3.0-8.0)and protein concentration(2-40 g/L).The stabilizing effect of ?-conglycinin on native glycinin solutions was investigated at the ratio from 5:1 to 1:5.Results indicated with the change of pH and ionic strength,glycinin has two distinctive macroscopic aggregation,forming large irregular protein flocs or spherical micro domain which can transform into each other when pH or ionic strength changed.Once the irregular protein flocs formed,it stopped evolution,which just slowly sedimented.However,spherical micro domains evolve with time.They coalesced rapidly and sedimented,forming a dense protein layer that accumulate on the bottom.In the native state,?-conglycinin physically binded to the glycinin.As a result,increasing the ratio of ?-conglycinin: glycinin deteriorate the aggregation.Increasing the protein concentration would enhance this stabilization effect.(2)Aqueous solutions of soy globulin were investigated over a wide range of temperatures(5-80 ?),protein concentrations(2-95 g/L),pH(5.3-7.5)and NaCl concentrations(0-0.5 M).The degree and rate of association of proteins was investigated by measuring the turbidity as a function of time.The structure of the protein assemblies in solution was determined by light scattering and confocal laser scanning microscopy.Results indicated that,under certain pH,in the presence of intermediate concentrations of NaCl(0.07 – 0.2 M)a fraction of soy protein phase separate to form spherical microdomains that slowly connect into large clusters and sediment.Microphase separation is strongest at 0.1 M NaCl and favored by decreasing the temperature or the pH and is reverted when the temperature or the pH is increased or when the ionic strength is reduced.Increasing SPI concentration,inhibit phase separation.The microdomains are slightly enriched in glycinin.At temperatures above 30 °C irreversible aggregation of SPI occurs at a rate that increases with increasing temperature.(3)Aqueous solutions of native soy globulin(I?0.003)were characterized over a wide range of protein concentrations(1-100 g/L)and pH(5.8-7.0)at 20 ?.The combined effects of pH and protein concentration on the self-assembly could be best understood by considering charge density(a)of the soy globulins.Soy globulin self-assembled into self-similar aggregates,with a size that increased with increasing concentration and decreasing net charge density.The aggregates slowly dissociated over a period of days when the solutions were diluted.When 6.1 ? pH ? 6.3,the evolution of the aggregates was non-monotonic with increasing protein concentration which was caused by the increasing charge density with increasing protein concentration.When the net charge density is >150,the size of the aggregates didn't change with the increasing protein concentration.A sharp increase of Mw and Rh was observed when the net charge density was reduced below a critical value which might be caused by the conformation change of soy glycinin.Further reducing the charge density,the size of the aggregates gradually increased until precipitated.(4)Aggregation and gelation of soy globulin was studied in salt free aqueous solution at neutral pH over a wide range of concentrations(0.3-90 g/L)and temperatures(50-95 °C).The structure of the aggregates that were formed during heating was characterized with light scattering.In all cases aggregates with the same self-similar structure were observed that were characterized by a fractal dimension df=2.0.Dynamic light scattering showed that the aggregates were flexible.The aggregate size increased with heating time and the rate of growth was characterized by an Arrhenius temperature dependence up to 85 °C with Ea=180 kJ/mol independent of the concentration.For a given temperature the aggregation rate increased very strongly with increasing concentration.Gels were formed at concentrations down to 50 g/L and at temperatures down to 50 °C.(5)Thermal aggregation and gelation of soy protein globulins was studied over a wide range of protein concentrations(1-95 g/L),pH values(5.8-6.8)and heating temperatures(65-95?).The aggregation and gelation rate increased with increasing protein concentration and decreasing pH.At low concentrations or short heating times irregular spherical particles were formed with a hydrodynamic radius that increased with decreasing pH from 30 to 40 nm and a density from 0.1 to 0.2 g/cm3.At longer heating times or at higher protein concentrations,these particles randomly aggregated into self-similar aggregates.With the decreasing pH,fractal dimension increase from 1.7 to 1.9.At a critical heating time,gelation was observed for C340 g/L,which was characterized by oscillatory shear measurements.The gel stiffness increased sharply with increasing protein concentration,but was almost independent of the pH when t>>tg.The temperature dependence(65-85?)of the gelation rate was characterized by an activation energy of 180 kJ/mol independent of the pH.Relationship between the pH and the charge density was not significant influenced by the aggregation determined as determined by potentiometric titration.(6)Thermal aggregation and gelation of soy protein isolate(SPI)was studied over a wide range of protein concentrations(1-95 g/L),NaCl concentrations(0-0.5 M)and temperatures(30-85 ?).The net charge density of the proteins was kept fixed?-175.At all conditions,self-similar aggregates were formed by random association of dense SPI nanoparticles that were formed in a first step.The density of the particles increased with increasing ionic strength.When NaCl?0.1M,increasing salt concentration led to faster gelation,but when NaCl > 0.1 M,gelation rate wasn't influenced by increasing salt concentration.Besides,NaCl concentration did not influence the gel stiffness at steady state but the heterogeneity which increased with increasing salt concentration.When T ?60 ?,the aggregation and gelaiton was mainly caused by the denaturation.When T < 60 ?,aggregation was also observed but the local density of the aggregates is lower than that formed at higher temperature.The effect of varying the ionic strength of thermal aggregation is similar with that of varying the pH.(7)Soy protein aggregates of different size(82->500 nm)were prepared by heating soy protein isolate(SPI)and characterizaed with light scattering.Salt induced gelation of aqueous solutions of the aggregates was investigated as a function of the protein concentration(10-75 g/L),NaCl concentration(0-0.5 M)and temperature(20-80 ?).The gel stiffness did not depend on the size of the aggregates nor the salt concentration,but increased with increasing protein concentration.Gelation of preheated aggregates was compared with that of native SPI,which showed that gelation of aggregates was faster than that of native proteins and occurred at lower protein concentrations and temperatures.The stiffness of gels formed by native SPI and preformed aggregates was not significantly different,but the structure of gels formed by aggregates was more homogeneous.For all systems the gel time had an Arrhenius temperature dependence characterized with an activation energy of 72 kJ/mol.A more limited investigation of gelation induced by adding CaCl2 was done to study the effect of the valence of the cation.