| Soybean protein which has already been widely used in food processing area is an important product in soy industry. Among all the properties of soy protein, gelation property is important and unique. During the recent years, high intensity ultrasound (HIU) technology has attracted a lot of attentions. Moreover, some recent researches have pointed out that HIU can change the physicochemical properties of soy protein. However, to the best of our knowledge, few systematic researches on using HIU to improve the gelation property of soy protein have been reported.In this study, the first step is to use HIU to change the physicochemical properties of soy protein. Then three kinds of gelation models, namely,"glucono-deta-lactone (GDL) tofu" heated gel,"traditional" tofu heated gel and TGase induced cold gel, were chosen. It was observed that HIU increased the gelation property of the above three kinds of gels. After that, in order to develop the soy protein macro-hydrogel as drug or nutritional compound carrier, TGase induced cold gel was chosen to encapsulate riboflavin. The reason for choosing TGase gel is because this gel can be formed at mild temperature thus can protect a lot of heat sensitive materials. In vitro experiments showed that40min HIU reduced the release speed of riboflavin obviously. Finally, soy7S and11S were treated by HIU and we found that the effects of HIU on7S were more profound than those of11S. Our specific findings were listed below:(1) The effects of low-frequency (20kHz) HIU at varying power (200,400or600W) and time (15or30min) on functional and structural properties of reconstituted soy protein isolate (SPI) dispersions were examined. HIU treatments reduced both the storage modulus and loss modulus of SPI dispersions and formed more viscous SPI dispersions (fluid character). Moreover, HIU treatment significantly decreased the consistency coefficients and increased the flow behaviour index of SPI dispersions. Scanning electron microscopy of lyophilized HIU SPI showed different microstructure with larger aggregates compared to non-treated SPI. No significant change was observed in the protein electrophoretic patterns by SDS-PAGE. However, free sulfhydryl content (SH), surface hydrophobicity and protein solubility of SPI dispersions were all increased with HIU treatment. Differences in solubility profiles in the presence versus absence of denaturing (0.5%sodium dodecyl sulfate and6M urea) and reducing (mercaptoethanol) agents suggested a decrease in non-covalent interactions of SPI in dispersion after HIU. Secondary structure analysis by circular dichroism indicated lower a-helix and random coil in SPI treated at lower power, in contrast to higher a-helix and lower β-sheet in SPI treated with higher power (600W). HIU resulted in partial unfolding and reduction of intermolecular interactions as demonstrated by increases in free sulfhydryl groups and surface hydrophobicity, leading to improved solubility and fluid character of SPI dispersions, while larger aggregates of HIU SPI in the dry state were formed after lyophilization.(2)HIU (20k Hz,400W) pre-treatments of SPI improved the water holding capacity (WHC), gel strength and gel firmness (final elastic moduli) of glucono-δ-lactone induced SPI gels (GISG). Sonication time (0,5,20, and40min) had a significant effect on the above three properties.20min HIU-GISG had the highest WHC (95.53±0.25%), gel strength (60.90±2.87g) and gel firmness (96340Pa), compared with other samples. Moreover, SH groups and non-covalent interactions of GISG also changed after HIU pre-treatments. The HIU GISG had denser and more uniform microstructures than the untreated GISG. Rheological investments showed that the cooling step (reduce the temperature from95℃to25℃at a speed of2℃/min) was more important for the HIU GISG network formation while the heat preservation step (keep temperature at95℃for20min) was more important for the untreated GISG. HIU reduced the particle size of SPI and Pearson correlation test showed that the particle size of SPI dispersions was negatively correlated with WHC, gel strength and gel firmness.(3) HIU (20kHz at400W for5,20or40min) pre-treatments of SPI changed the particle distribution and reduced particle size of SPI dispersions. Surface hydrophobicity and free SH content of SPI increased with HIU time. Free SH content of CaSO4-induced SPI gels (CISG) and protein solubility in the presence of SDS and urea decreased after HIU pretreatments, suggesting HIU facilitated disulfide bond formation during CISG formation. HIU resulted in more uniform and denser gel network, WHC and gel strength of CISG. Furthermore, WHC and gel strength were positively correlated with free SH content of heated SPI and surface hydrophobicity of unheated SPI, and negatively correlated with particle size of heated SPI and free SH content of CISG. In conclusion, HUS induced structural changes in SPI molecules, leading to different microstructure and improved WHC and gel strength of CISG.(4)The mechanism of HIU improvement of gelation properties of GISG and CISG could be summarized as follow: HIU reduced the particle size of soy protein, meanwhile, the hydrophobic and SH groups were exposed from the interior of SPI aggregate or molecular to the surface. The following heat step further reduced the particle size and exposed active groups. Moreover, HIU facilitated the formation of soluble protein aggregates which might be formed as non-soluble aggregates during the heating process. After that, coagulates were added and aggregates were formed. HIU changes the structures and conformation of SPI, which may accelerate the formation of intermolecular hydrophobic interactions and S-S bonds, finally resulting in dense and uniform3D structure.(5) HIU increased the gelation property of TGase induced cold SPI gel (TISG). TISG was used as control release model to encapsulate riboflavin because TISG was a cold gel which can protect heat sensitive materials.40min HIU (20kHz,400W) increased the gel yield from6.02to11.27, increased the gel strength from11.4g to37.5g and increased the encapsulate efficiency from88.8%to100%. Moreover,40min HIU reduced the riboflavin release speed in simulated gastric or intestinal fluid. Further investigations showed that the swell property of TISG reduced while the anti-erosion property increased. SDS-PAGE indicated that HIU increased the cross-link degree of SPI when treated by TGase. Raman spectroscopy revealed that HIU pretreatment of TISG changed the microenvironment of polypeptide and the chemistry of amino acid side chain, indicating the modification of tertiary structure.(6) The effects of HIU (20kHz at400W for5,20or40min) on soybean P-conglycinin (7S) and glycinin (11S) fractions were investigated in this study. HIU decreased turbidity and particle size of7S in0.05M Tris-HCl buffer at pH7.0, while it increased surface hydrophobicity, solubility, emulsifying activity (EAI) and emulsion stability (ESI). Similarly, HIU of soybean glycinin (11S) decreased turbidity while increasing EAI but it had minimal effects on particle size and ESI. Furthermore, surface hydrophobicity and solubility of11S decreased during the first20min of HIU but then increased upon longer treatment. The SH groups of both7S and11S fractions decreased after HIU. HIU did not change7S or11S secondary structure, but it slightly increased the percentage of high molecular-weight aggregates under non-reducing SDS-PAGE, and changed the microenvironment of aromatic and aliphatic side chains as observed by Raman spectroscopy of freeze-dried samples. The physicochemical changes of11S and especially of7S proteins induced by HIU treatment may contribute to improved applications of soy proteins in food products. |
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