| As a green and efficient physical field heating technology,microwave has been applied in the processing and production of surimi products.Other studies have confirmed that microwave energy can change the conformation of enzymes and affect the enzymatic reaction process.This study focuses on the degradation effect induced by Cat L(Cat L)on surimi gel,and studied the effect of microwave heating on Cat L(EC 3.4.22.15)-induced myofibriller protein(MPs)degradation and gel formation ability,moreover,its influence mechanism was preliminarily speculated.This study aims to provide a theoretical basis for better use of microwave heating methods to improve the gel properties of surimi products.The main research contents and results are as follows(1)Preliminary isolation and purification of Cat L from the dorsal muscle of silver carpThis study has studied the effects of acidification treatment conditions,ammonium sulfate saturation and molecular weight of the dialysis bag on the Cat L activity in the crude enzyme solution,and explored the effect of different types of ion exchange chromatography columns and equilibrium buffer p H on the protein chromatographic purification results to optimize the preliminary separation and purification scheme of Cat L.The results showed that the optimal condition for acid treatment was p H3.0,and the p H was adjusted to p H6.0 after heat treatment at 40? for 10 min.The salting-out ammonium sulfate saturation range was 55% to 90%,and the optimal molecular weight of the dialysis bag was 7000 Da.The optimized scheme of chromatographic purification was two-step chromatography which was weak anion exchange followed by molecular sieve.In addition,20 mmol/L,p H 6.0 phosphate buffer was selected as the equilibrium buffer of the weak anion column,and 45 k Da electrophoresis pure Cat L is obtained.The purification fold was 487,and the recovery rate was 22 mg/1000 g.This method could remove impurities to the maximum extent and has a high recovery rate.(2)Effect of microwave on degradation of MPs gel induced by Cat LBy simulating the reaction of myofibrillar protein and Cat L in the heat-induced gel formation process in the actual system,the results show that Cat L degraded myofibrillar protein into small molecule peptides,destroyed the protein gel network structure,and made the gel network structure more loose and porous,which in turn led to the apparent collapse of the myofibrillar protein gel and the decrease of the gel strength.At the same heating rate,the protein gel strength of the microwave-enzyme group was 13.5 g·cm higher than that of the water-bath enzyme group,and the TCA soluble peptide content was higher by 0.7 ?mol Tyr/g.At the same time,the SDS-PAGE profile of the water-bath enzyme group showed more low-molecularweight bands,indicating that microwave effectively weakened the degradation of myofibrillar protein by Cat L,thereby making up for and improving gel degradation.(3)Preliminary discussion on the mechanism of microwave weakening Cat L-induced degradation of MPs gelBy exploring the changes in water distribution,protein secondary structure and main chemical forces during the formation of protein gels,the corresponding mechanism for microwaves to effectively weaken the degradation of the gel was preliminarily discussed from the perspective of protein conformation changes.The results showed that microwave interfered with protein conformation and spatial structure,promoted the conversion of myofibrillar protein ?-helix to other secondary structure,induced protein aggregation,enhanced hydrophobic interaction between protein molecules and increased disulfide bonds.Microwave intervention in protein aggregation further reduced the probability of contact between enzymes and active sites of myofibrillar protein,thereby preventing Cat L from degrading myofibrillar protein.In conclusion,microwave can effectively weaken the degradation Cat L-induced degradation of MPs gel,thereby improving the deterioration of surimi gel. |
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