Research On High Hydrostatic Pressure Treatment On Quality Of Trichiurus Lepturus During Cold Storage

Trichiurus lepturus is rich in unsaturated fatty acids, protein, lecithin, phosphorus, calcium, vitamins and other nutrients, Trichiurus lepturus from East China Sea has more nutrients in the Trichiurus lepturus species, but it easily cause spoilage due to containing more unsaturated fatty acids. High hydrostatic pressure treatment is a kind of physical technology which put the food with the vacuum packaging into the medium pressure and then process the food by 100 to 1000 MPa. To improve the preservation quality of Trichiurus lepturus during cold storage, three parts of research were done in this paper. High hydrostatic pressure treatment on the preservation of Trichiurus lepturus with cold storage were studied in the first part. High hydrostatic pressure treatment combined with other preservation methods（natural antioxidants and modified atmosphere package） to keep the quality characteristic of Trichiurus lepturus during cold storage were studied in the second part. The last part were studied microbial ecology of Trichiurus lepturus pretreated by high hydrostatic pressure during cold storage identified by phenotypic and 16 S r RNA gene. The mainly research contents and results were listed as follows:1. Try to find the best condition of high hydrostatic pressure treatment. Samples untreated with HHP stored under 4 ℃ were selected as the control. Different high hydrostatic pressure treatment with 3,6,9 min and 220 MPa,290 MPa pressure and stored at 4 ℃ repsectively,the total samples were divided into 7 groups. The samples were tested every two days,the total plate count（TPC）,sensory evaluation, water holding capacity（WHC）, volatile base nitrogen（TVB-N）, trimethylamine nitrogen（TMA-N）, thiobarbituric acid（TBA） were analyzed. It showed that HHP can effectively control spoilage microorganisms, and its ability increased with the pressure and holding time. The shelf life of the control group was 4 days, the shelf life of the samples treated with 220 MPa HHP and 290 MPa HHP were 10 days, 12 days respectively. With the increase of pressure and holding time, the Trichiurus lepturus was becoming less transparent. Although the water holding capacity decreased, TMA-N and oxidation was effectively inhibited. After HHP treatment, the TVB-N was increased with pressure. TVB-N in the control group was growing faster. The longer holding time, the higher the pressure, the better inhibitory effect of TVB-N. Among them, the preservation effect by 290 MPa, 6 min HHP treatment and 290 MPa, 9 min HHP treatment were the recommended processing conditions, the TPC were 6.19 lg（CFU/g）, 6.04 lg（CFU/g） respectively in 12 th, TVB-N were 19.87 mg N/100 g, 20.86 mg N/100 g respectively on the 12 th days, which were still in the second level freshness of the beltfish.2. Study on the high hydrostatic pressure treatment combined with natural antioxidants keeping quality characteristic of Trichiurus lepturus during cold storage. In order to improve the quality characteristics of Trichiurus lepturus during chilled storage, Trichiurus lepturus during 4 ℃ cold storage were selected as controls, HHP（290 MPa, 6 min）,HHP（290 MPa, 6 min） combined with three different compound antioxidants（2.5 % carboxymethyl chitosan+0.05 % Vitamin E TPGS, 2.5 % carboxymethyl chitosan+0.25 % water-solubility rosemary extract, 0.05 % Vitamin E TPGS + 0.25 % water-solubility rosemary extract） to pretreated Trichiurus lepturus. To evaluate the preservation effects of Trichiurus lepturus during 20 days at 4 ℃, sensory evaluation, total plate count, physicochemical（total volatile basic nitrogen, the thiobarbituric acid, acid value and water holding capability） were periodically analyzed. Results showed that the shelf-life of the controls were 6 days, the shelf-life of Trichiurus lepturus pretreated by HHP were 12 days, the shelf-life of Trichiurus lepturus pretreated by HHP conbined with natural antioxidants were about 18 days. The total plate count, TVB-N, TBA, acid value of samples pretreated by natural antioxidants and HHP which were lower than those of HHP groupssignificantly, and lower than those of the controls. Among them, the preservation effects of the samples dipped in 0.25 % water-solubility rosemary extract+0.05 % Vitamin E TPGS had the best preservation effects, which could suppress the growth of spoilage microorganisms effectively, slow down the decomposition of protein, inhibited lipid oxidation and hydrolysis significantly, and maintain the good quality of Trichiurus lepturus effectively.3. Resaerh on high hydrostatic pressure treatment combined with modified atmosphere package keeping quality characteristic of Trichiurus lepturus during cold storage. The vacuum-packaged samples pretreated by HHP（290 MPa, 6 min） and stored at 4 ℃ were used as the control. The HHP（290 MPa, 6 min） and then packed with different proportions of modified atmosphere packages（MAP1: 60 %CO₂+40 %N₂; MAP2: 60 %CO₂+7 %O₂+33 %N₂; MAP3: 60 %CO₂+15 %O₂+25 %N₂; MAP4: 50 %CO₂+15%O₂+35%N₂） were used to package fresh Trichiurus lepturus. When all pretreatments were finished, the experimental samples were stored at 4 ℃ during the rest period of processing. During the storage of 21 days, sensory evaluation, total plate count, total volatile basic nitrogen, thiobarbituric acid, actomyosin content and total sulfhydryls content of actomyosin were periodically determined to investigate the preservation effects of Trichiurus lepturus during cold storage at 4 ℃. According to the results of total plate count and total volatile base nitrogen, the shelf life of the control（Trichiurus lepturus pretreated by HHP and vacuum packaging） was only 11 days. The experimental groups of HHP treatment combined with different modified atmosphere packages had longer shelf life of Trichiurus lepturus during cold storage at 4 ℃: the shelf life of Trichiurus lepturus pretreated by HHP and MAP（60 %CO₂+40%N₂） was 16 days, that pretreated by HHP and MAP（60 %CO₂+7 %O₂+33 %N₂） was 19 days, and the shelf lifes of Trichiurus lepturus pretreated by HHP and MAP（60 %CO₂+15 %O₂+25 %N₂ and 50 %CO₂+15 %O₂+35 %N₂） were both 21 days, which was almost 2 times longer than that of the control. This observation suggested that HHP treatment combined with MAP had higher effectiveness on inhibiting total plate count and total volatile base nitrogen. However, modified atmosphere packages could accelerate the decrease of actomyosin content and total sulfhydryls content of Trichiurus lepturus pretreated by HHP, and higher level of O₂ in modified atmosphere packages also had obvious promotion effect for fat oxidation of Trichiurus lepturus pretreated by HHP during cold storage at 4 ℃. In summary, high hydrostatic pressure treatment combined with low proportion O₂ in modified atmosphere packages（60 %CO₂+7 %O₂+33 %N₂） could not only prolong the shelf life of Trichiurus lepturus, but also keep its natural properties at the same time.4. The effect of high hydrostatic pressure（290 MPa, 6 min） on microbial ecology changes of Trichiurus lepturus during vacuum-packaged storage（12 d） was studied by via 16 S r RNA gene sequencing analysis and phenotypic methods. The microbial colonies of aerobic culture and anaerobic culture, which were obtained from 5 microflora’s growth stages（including fresh Trichiurus lepturus, fresh Trichiurus lepturus which were stored under 4 ℃ for 4, 8 and 12 d after high hydrostatic pressure treatment）, were isolated and purified by the tryptic soy agar medium and tryptic soy broth medium, repeatedly classified by the traditional phenotypic methods（morphological, physiological and biochemical characteristics of bacteria strains）, and then the bacteria DNA of pure culture was extracted and amplified by the polymerase chain reaction（PCR）. Finally, 24 bacteria strains of aerobic culture and anaerobic culture in the vacuum-packaged Trichiurus lepturus under the 4 ℃ cold storage were obtained. The result showed that Pseudomonas brenneri, Pseudomonas fulva and Psychrobacter faecalis which appeared in the initial microflora of fresh Trichiurus lepturus were inactivated and non-existent after the high hydrostatic pressure treatment; 5 bacteria strains（including Shewanella baltica, Pseudomonas lundensis, Stenotrophomonas rhizophila, Staphylococcus epidermidis and Microbacterium oxydans） gradually reduced and disappeared after the high hydrostatic pressure treatment. Three bacteria strains gradually grew with the extension of storage time, which were Microbacterium halimionae, Macrococcus caseolyticus and Rhizobium larrymoorei. In the whole process of storage after the high hydrostatic pressure treatment, 4 bacteria strains（Moraxella osloensis, Kocuria rhizophila, Carnobacterium maltaromaticum and Dermacoccus nishinomiyaensis） always existed, and they were not sensitive to the high hydrostatic pressure treatment. Especially Carnobacterium maltaromaticum, which had a high proportion of total bacteria amount under the conditions of aerobic and anaerobic cultivation, could be considered as a dominate strain during the 4 ℃ cold storage. At the end of the shelf life of vacuum-packaged Trichiurus lepturus pretreated by high hydrostatic pressure, 5 bacteria strains（Leucobacter aerolatus, Pantoea agglomerans, Yersinia enterocolitica subsp. Palearctica, Chryseobacterium vrystaatense and Brachybacterium rhamnosum） appeared. The colony evolution of Trichiurus lepturus during the cold storage showed that Gram negative bacteria were sensitive to the high hydrostatic pressure treatment, while Gram positive bacteria had a higher tolerance to the high hydrostatic pressure treatment. Under the influence of high hydrostatic pressure treatment, the microorganisms which have stronger ability of causing corruption are inhibited, and the microorganisms which have weaker ability of causing corruption become dominant bacteria, which may be one of the factors that the high hydrostatic pressure treatment can effectively extend the shelf life of Trichiurus lepturus.