Study On Sterilizing And Mechanism Of High Pressure Processing Hurdle Technology For Mud Snail(Buttacta Exarata)

High pressure processing?HPP?is a new type of non-thermal sterilization technology,which has attracted much attention in biological and food engineering fields recently.Part of microorganisms can be killed by a certain range of pressure,but there are some limitations in the application for the weak effects of HPP on Gram-positive bacteria and spores.Therefore,how to improve the sterilization effect of HPP is of great significance.In order to improve the reliability and application scope of HPP technology,this paper discussed the feasibility and mechanism of multi-technology synergy.Mud snail of Bullacta exarata is an important shellfish product in China,and it contains abundant protein and human essential amino acids and is loved by consumers.However,the mud snail contained a large number of microbes,and the traditional curing methods and people's eating habits had greater security risks.This was an important problem in the processing of mud snail,and also an ideal experimental material in this study.The purpose of this study was to study the inactivation of microbes in mud snail and the effect of HPP combined with slightly acidic electrolyzed water?SAEW?on Bacillus cereus spore.In this study,single cycle HPP,multi-cycle HPP mode,and combined with different available chlorine content?ACC?of SAEW,were applied.The specific research content was divided into the following six parts:?1?The effects of single-and multi-cycle HPP on the microbes of mud snail during the chilled storage were investigated.The results showed that the increase of pressure could significantly reduce microbes of mud snail.The H₂S-producing bacteria in the sample were not detected by different treatments.The total microbial count of samples treated with the multi-cycle HPP grew slowly in the first 20 d,the psychrophilic microbes was not detected in 0 and 10 d,and the total enterobacteriaceae and H₂S-producing bacteria were not detected in the whole storage.The modified Gompertz model showed that when the single cycle HPP was 500 MPa/20 min,the prediction time of the total bacteria was 25 d,and the growth lag was 9.55d;while the multi-cycle HPP?200 MPa/20 min+500 MPa/20 min?could prolong the prediction time and growth lag time of the total bacteria to 38 d and 14.26 d.?2?Effects of multi-cycle HPP?200 MPa/20 min+500 MPa/5 min?combined with different temperature?30? and 60?,HPP30 and HPP60?on the physical characteristics?pH value and conductivity?and bacterial diversity of B.exarata during refrigerated storage were explored.HPP30and HPP60 groups significantly reduced total microbial counts by about 3log CFU/g,and samples treated with HPP30 had more survivial bacteria than HPP60.Different treatments resulted in a higher pH and conductivity values,and also the conductivities were significantly increased?P<0.05?during refrigerated storage.The diversity of microbes in the three groups increased first,and then decreased,and the genus of Bacillus were dominant bacteria when sample was refrigerated to 20 d.Bacillus cereus was the dominant cultivable bacteria in the mud snail at late stage.The effects of HPP on Gram-positive bacteria and spores were weak,hence,the HPP combined with SAEW were used to treat Bacillus cereus spores.?3?Synergistic effects of HPP and SAEW?24,35,44,and 55 mg/L?on the inactivation of B.cereus spores were examined.The individual effects of HPP or SAEW on spores were less than 1.0 log CFU/mL.Single cycle HPP combined with SAEW treatment was shown to have significantly positive effects on spores'inactivation?P<0.05?,and its optimal conditions was 300 MPa HPP+SAEW with 44 mg/L ACC,producing reductions of3.27 log CFU/mL.The optimal conditions of multi cycle HPP combined with SAEW was 200 MPa HPP+SAEW with 44 mg/L ACC+500 MPa HPP,producing reductions of 3.99 log CFU/mL.?4?The effects of HPP and SAEW on the structure of B.cereus spores were investigated.The morphology of B.cereus spores treated with HPP-SAEW exhibited an incomplete exosporium,and the inner structure of spores had an irregular surface bumps.The DPA release of spores treated with HPP+SAEW was higher than that of treated with HPP.The membrane permeability results indicated that HPP-SAEW methods was not mainly depended on germination,but also the damage of inner membrane.?5?Label free-based proteomic was used to analysis the inactivation mechanism of multi-cycle HPP combined with SAEW against B.cereus spores.Compared with control group,a total of 93 and 83 proteins of HPP-SAEW and HPP were significantly changed?fold-changes?±2 and P<0.05?,also there was 21 proteins was significantly changed?fold-changes?±2 and P<0.05?between HPP-SAEW and HPP.The results indicated the spore cortex-lytic enzyme for spore germinating was upregulated 3.15 fold after multi-cycle HPP treatment.The the metabolic,degradation,signaling,and biosynthesis pathways were important pathways underlying the HPP-SAEW mediated spore inactivation.Amino acid-biosynthetic and quorum sensing were the main KEGG pathway mediated by SAEW under HPP conditions.Specifically,the phosphorylation level was the main reason for quorum sensing pathway.?6?The effects of HPP-SAEW and traditional heat processing on microbe,physical quality,and free amino acids of B.exarata were evaluated.Results showed that the total microbial counts and psychrotrophic bacteria of the mud snail treated with HPP-SAEW were below detection limit.HPP-SAEW had no effect on the water content,also can significantly improve the hardness of the mud snail,and had little influence on the microstructure.The total free amino acid?TFAA?of sample treated with HPP-SAEW was13.115 mg/g dw,and the essential free amino acid?EFAA?had no significant change.The water content,electrical conductivity and drip loss rate of the traditional heat processing group decreased significantly,the tissue was severely contracted,and TFAA was only 6.013 mg/g dw.The research results provided technical support for deep processing of mud snails and similar aquatic products processing.The results indicated that HPP-SAEW could improve HPP in the inactivation of B.cereus spores,also prodive a development in the processing of B.exarata,which promote the application of HPP technology in biological and food engineering fields.