This research studied the effects of alcalase hydrolysis on the structure, physicochemical properties and aggregation behavior of quinoa protein isolate (QPI), and explored its effects on the gelling properties of QPI.

QPI was extracted by alkaline extraction and acid precipitation method at 4 ℃. Alcalase with different enzyme -substrate ratios was added to the QPI solution to hydrolyze the protein. Thereafter, the changes of the composition, particle size, Zeta potential, solubility, and surface hydrophobicity (S₀-ANS) of QPI were analyzed, and the correlation between these changes and the Th T fluorescence intensity and morphology of protein thermal aggregates were discussed. Furthermore, the concentration of QPI dispersion was elevated to form heat-induced QPI gels. The correlation among alcalase hydrolysis, aggregation behavior and gelling properties of QPI was discussed through the analysis of the microstructure, texture and protein secondary structure of QPI gels. Meanwhile, the reasons for the changes in the gelling properties of QPI induced by alcalase hydrolysis were revealed.

With the increase of enzyme substrate ratio (E/S), the protein particle size of QPI dispersion (2%, w/v) gradually decreased, and its electronegativity increased first and then decreased. Meanwhile, the surface hydrophobicity of QPI gradually increased as the E/S increased. At the same time, the Th T fluorescence intensity of QPI aggregates showed an upward trend as E/S rose from 0 to 0.08%, and then declined when the E/S further increased to 0.14%. Through TEM observation, short fibrils (145-306 nm) and long fibrils (217-406 nm) were formed when E/S was 0.05% and 0.08%, respectively. However, with the further increase of E/S ratio, the length of fibrils became shorter, and more amorphous aggregates appeared. In addition, it was found that the aggregation behavior of QPI had a significant effect on its gelling properties (P<0.05). QPI gels showed enhanced hardness, higher storage modulus and a denser network structure, when fibrillar aggregation was dominant. Compared with short fibrils, long fibrils exerted a more significant effect to improve the gelling properties. Furthermore, alcalase showed a significant effect on the protein secondary structure of QPI gels, and the contents of β-sheet and random coil rose first and then decreased with the increase of E/S.

The limited alcalase hydrolysis promoted QPI to form ordered fibrillar aggregates, and further improved its gelling properties. When the E/S was 0.08%, QPI formed the longest fibrils, and QPI gels exhibited the highest hardness as well as the most favorable viscoelastic properties. Meanwhile, the protein secondary structure of QPI gels was ordered, and their microstructure was dense. However, the higher degree of hydrolysis was unfavorable to form fibrillar aggregates and failed to improve the gelling properties. Therefore, limited alcalase hydrolysis could significantly improve the weak gelling properties of QPI through fibrillation.

Based on the freeze-thaw stability of quinoa stabilized Pickering emulsion, it was incorporated evenly into surimi to generate surimi gels in this study, and its feasibility to improve the freeze-thaw stability of fish protein gel was evaluated. This research aimed to prevent the deterioration of fish protein gel, caused by the temperature fluctuation during storage and transportation.

Quinoa protein Pickering emulsion was prepared and distributed in surimi, followed by heating process to generate surimi gels. The surimi gels with different contents of Pickering emulsion and without emulsion were subjected to three freeze-thaw cycles, and then, the texture, color, water and ice crystal distribution and drip loss of surimi gels were measured.

The quinoa protein Pickering emulsion improved the lightness and whiteness of fish surimi gel, and inhibited the changes of color after freeze-thaw cycles. Meanwhile, the Pickering emulsion addition delayed the changes of hardness and chewiness of surimi gels during freeze-thaw cycles. It was found that quinoa protein Pickering emulsion had no effect on the moisture distribution of surimi gels before freeze-thaw cycles, but significantly increased the proportion of immobile water and decreased free water content in surimi gels after freeze-thaw cycles. Therefore, the drip loss of surimi gel was decreased by emulsion addition. Furthermore, the emulsion addition decreased the diameter of ice crystals formed in surimi gel, reduced the damage to muscle tissues, and decreased the free water content.

The Quinoa protein Pickering emulsion weakened the adverse impact of freeze-thaw cycles on the color and textural properties, maintained the gel structure, and improved the freeze-thaw stability of fish surimi gel, which maintained its quality and nutritional value. Quinoa protein Pickering emulsion was promising to become an innovative antifreeze to be applied in frozen food.

This research studied the effects of different salt ions on the gel properties of quinoa protein, and explored its molecular mechanisms, so as to provide a theoretical basis for the processing of quinoa protein gels.

Quinoa protein was extracted by alkali extraction and acid precipitation. Quinoa protein solution (20%, w/v) was prepared at pH 7.0. NaCl, CaCl₂, CaSO₄ and MgCl₂ was added in quinoa protein solution till the concentration was 50 mmol∙L^-1, and then the solution was heated in a water bath to prepare quinoa protein gels. The effects of salt ions on the texture, water retention, color properties and water distribution of quinoa protein gels were analyzed. Meanwhile, the effects of salt ions on the microstructure and rheological properties of quinoa protein gels were studied by scanning electron microscopy and rheometer. The effects of salt ions on the molecular interactions and secondary structure of protein gels were also analyzed.

The addition of salt ions significantly decreased the hardness and water holding capacity, while increased the springiness of quinoa protein gels under pH 7.0. Quinoa protein gels with MgCl₂ showed the lowest hardness and water holding capacity. NaCl addition had no significant influence on the color properties of protein gels. However, the addition of bivalent salt ions significantly improved the lightness and whiteness of quinoa protein gels, and their whiteness increased from 59.62 to 67.80 with the addition of CaCl₂. Furthermore, the addition of salt ions promoted granular aggregation of quinoa protein, which made the gel network structure become coarse. Coarse and larger gaps were observed in the microstructure of quinoa protein gels when divalent salt ions were added. Meanwhile, compared with blank gels and gels added with NaCl, the addition of divalent salt ions significantly decreased the content of disulfide bond, and weakened the electrostatic interactions within quinoa protein gels. Furthermore, the addition of salt ions decreased the contents of β-sheets and β-turns, increased the contents of α-helix and random coil, which affected the orderliness of protein secondary structure.

Under neutral conditions, the gel properties of quinoa protein and microstructure of gels were affected by the presence of different salt ions to various degrees. Compared with the gel prepared with NaCl, quinoa protein gels with the same concentration of CaCl₂, CaSO₄, and MgCl₂ showed rougher microstructure, lower gel hardness and water holding capacity, as divalent salt ions significantly decreased the disulfide bond content and weakened the electrostatic interactions within quinoa protein gels.