Preparation Of Nisin Nanoparticles Based On Protease Activity Regulation Strategy And Their Antiseptic Application Research

The preservation and antisepsis of meat products have become one of the most concerning issues in the meat production industry.Nisin is a natural antibacterial peptide widely used in the food industry.However,when nisin is used in meat products,it will be affected by protease or other components in meat products,resulting in a decrease in bacteriostatic activity.Isolate soy protein(ISP)and egg white protein(EWP)were widely available,inexpensive and nutritious proteins,which coud not only be used as protease inhibitors,but also ideal raw materials for protein-based nanocarriers.Based on the perspective of protease inhibition in meat products,nisin-isolate soy protein(NISPN)and nisin-egg white protein nanoparticles(NEWPN)were prepared to protect the antibacterial activity and stability of nisin,and the mechanisms of action of two nanoparticles were researched,along with further analysis of the application effect of two nanoparticles in low-salt emulsified sausage,to improve the antibacterial stability of nisin in meat products and solve the problem of limited application of nisin in the meat processing industry.The main research contents were as follows:1.Study on the stability of nisin in meat products(1)To investigate the effect of endogenous proteases in meat products on the antibacterial activity of nisin by measuring the activity of nisin in meat products and proteomic analysis.The results showed that there was still a certain amount of protease in cooked meat products,in which the antibacterial activity of nisin decreased.(2)To protect the bacteriostatic activity of nisin from the perspective of inhibiting the protease activity in cooked meat products,the nisin bacteriostatic experiment was performed to screen natural broad-spectrum protease inhibitors.The results showed that ISP and EWP possessed significant protective effect on the antibacterial activity of nisin in cooked meat products.(3)Micrococcus luteus was used as a model strain to explore the protective effects of ISP and EWP on nisin in cooked meat products.The results showed that compared with nisin alone,the mixture of nisin and ISP or EWP existed better inhibition activity on the growth of M.luteus,indicating that ISP and EWP had the effect of protecting the bacteriostatic activity of nisin in cooked meat products.2.Preparation and properties of nisin nano-formulations(1)Nisin-loaded ISP/EWP nanoparticles were prepared by the acid-heat induction method,both of which represented stable systems with particle sizes of 154 nm and113 nm,respectively,with the encapsulation efficiency of 95.38% and 99.46%,respectively.(2)The results of spectral analysis and scanning electron microscopy indicated that the formations of NISPN and NEWPN were mainly mediated by electrostatic interactions and hydrogen bonding.In the nanoparticle system,the secondary structure of nisin might be affected by ISP and EWP,but the main functional structure of nisin would not be affected.(3)The application experiments of NISPN and NEWPN in emulsified sausage showed that the antibacterial stability of nisin in the form of nanoparticles was higher than that of free nisin during storage of emulsified sausage,and meanwhile they could reduce the degree of spoilage and lipid oxidation of emulsified sausage.3.Application of NISPN and NEWPN in low-salt emulsified sausages(1)Through the measurement of p H,cooking loss,texture,color and sensory evaluation of low-salt emulsified sausages,the results showed that NISPN and NEWPN could maintain or improve the quality of low-salt emulsified sausages during storage to a certain extent and maintained their good organoleptic acceptability.(2)By measuring the total number of colonies(TVC),volatile base nitrogen(TVB-N)and thiobarbituric acid reactive substances(TBARS)of low-salt emulsified sausage,the preservation and antisepsis effects of NISPN and NEWPN were evaluated in low-salt emulsified sausages.The results showed that the reduction of salt content made the emulsified sausages more susceptible to spoilage,but the addition of NISPN or NEWPN could delay this process and further prolong the storage period of the lowsalt emulsified sausages.In conclusion,endogenous proteases in meat products could affect the antibacterial activity of nisin,while ISP and EWP as protease activity regulators could protect the antibacterial activity of nisin in meat products.Both proteins could be used as protein-based nanocarriers to encapsulate nisin to form stable nanoparticles,which could significantly increase the antibacterial stability of nisin in the environment of endogenous proteases and further improve the quality characteristics and sensory evaluation of low-salt emulsified sausages,with the effect of extending the storage period of low-salt emulsified sausages and expanding the application of nisin in meat products.