Preparation And Interaction Mechanism Analysis Of Antimicrobial Fermented Broth And The Application Of Preservation Of Rosa Roxburghii Tratt

Roxburgh rose is rich in a variety of bioactive compounds,but has an extremely high decay and loss rate because of mechanical damage and microbial infection during postharvest storage.Edible films provide a protective barrier to prevent material transfer between the fruit and environment,improving the shelf life of fruit,but the existing natural antimicrobial agents were expensive and had unclear target action.According to the previous research,it was found that the enzymatic hydrolysate of walnut meal had antibacterial activity.In this study,the fermentation of walnut meal was used to promote the antimicrobial activity against the dominant microorganism that causing rot to roxburgh rose.The interaction mechanism between the antimicrobial polyphenol and peptide in the fermented walnut meal was analyzed.The ultrasonic treatment was assisted to prepare carboxymethyl chitosan/gelatin/fermented walnut meal film,and the antimicrobial film was applied to extend the shelf life of roxburgh rose.The results are as follows:1、Penicillium.victoriae was the dominant microorganism that causing rot to roxburgh rose,which was isolated and identified by ITS amplified sequencing technology.According to the results of the antimicrobial activity against P.victoriae,Escherichia.Coli,and Staphylococcus.aureus,Lactiplantibacillus plantarum B7 and Bacillus subtilis were selected from 6 strain combinations to ferment walnut meal.Following the optimization by random-centroid,the optimal fermentation process was 35℃,6.50%of inoculation amount,and 26 h.In the whole fermentation,the total polyphenol and peptide contents showed a trend of decreasing firstly and increasing until 20 h,then decreasing.The p H value decreased continuously until 16 h with the extension of fermentation time.Fifteen phenolic compounds were quantified in the fermented broth,among which the concentration of gallic acid,caffeic acid,salicylic acid(SA),and chlorogenic acid increased by 125-fold,119-fold,117-fold,and112-fold,respectively,but naringin-7-O-glucoside disappeared.SA showed the highest antimicrobial activity among seven antimicrobial phenolic compounds.Ultrafiltration was used to separate fermented walnut meal into four molecular weight components,and 0～3 k Da fermented walnut meal presented the strongest antimicrobial activity against P.victoriae and had the highest hydrophobic amino acid content.2、143 peptides were obtained by LC-MS/MS from 0～3 k Da fermented walnut meal.FGGDSTHP,ALGGGY,YVVPW and PLLRW were screened by Py Rx virtual,and the antimicrobial capacities of 4peptides were different,among which the YVVPW had the best antimicrobial activity.The computer simulation results demonstrated that the YVVPW was closely bonded with Rho1 GTPase,and hydrogen bonding,hydrophobic interaction,andπ-πstacking were the main interaction forces between the YVVPW and Rho1 GTPase.According to the minimum inhibitory concentration(MIC)against P.victoriae,the YVVPW and SA were mixed according to the checkerboard method,and the MIC values of complex were 93.75%～93.86%lower than the MIC values of the two compounds alone.The results of fluorescence spectra,FTIR spectra,and molecular docking indicated that hydrogen bond,hydrophobic interaction,andπ-πstacking were the main interaction forces between YVVPW and SA.3、According to the results of contact angle,mechanical properties,and antimicrobial activities,the optimal parameters of the ultrasonic assisted preparation of carboxymethyl chitosan(CMCS)/gelatin(GL)/fermented walnut meal(FW)film were as follows:ultrasonic time 5 min,ultrasonic power 560 W,and 1.0 wt%of fermented walnut meal.Compared to CMCS/GL film,the oxygen,carbon dioxide,and water vapor permeability of U-CMCS/GL film were reduced by 72.47%,25.62%,and 0.25%,respectively.Compared to U-CMCS/GL film,the oxygen,carbon dioxide,and water vapor permeability of U-CMCS/GL/FW film were decreased by 0.33%,4.63%,and 0.62%,respectively.The FTIR,XRD,TG,and SEM results showed that the ultrasonic treatment enhanced the hydrogen bond force between CMCS and GL,weakened the diffraction peak intensity,improved the thermal stability,and made the film become smooth and uniform.The intermolecular interaction between CMCS and GL was destroyed by added FW,and there were a few particles on the surface,but the film still had high thermal stability.The TEM results demonstrated that the cell wall of P.victoriae was thinned by U-CMCS/GL film,and the plasma membrane of P.victoriae was separated and the internal structure was blurred by U-CMCS/GL/FW film.4、The roxburgh rose was treated by U-CMCS/GL film and U-CMCS/GL/FW film.Compared to the control group,the U-CMCS/GL/FW film could significantly inhibit roxburgh rose rot,and the maximum respiration rate was decreased by 18.29%.The hardness,soluble solid,titratable acid,and total polyphenol contents were increased by 12.20%,5.23%,11.71%,and 8.11%,respectively,and the ascorbic acid content was still as high as 15.80±0.12 mg/g.Furthermore,the activities of SOD,POD,and CAT in the three groups were all increased first and then decreased,and the antioxidant enzyme activities were improved by the U-CMCS/GL/FW film.At the end of storage,the aerobic bacterial count of all groups exceeded 10~5CFU/g,but the U-CMCS/GL/FW film decreased the number of yeasts and molds.Comparing the results of the diversity of fungal before and after storage,it was found that the relative abundance of P.victoriae decreased to 0.007%by U-CMCS/GL/FW film,indicating that antimicrobial film not only reduced the number of colonies,but also decreased the diversity of fungal.In summary,this study explored the synergistic mechanism between the polyphenol and antimicrobial peptide in the fermented walnut meal,and provides theoretical basis for extending the shelf life of roxburgh rose.