二氧化碳辅助超高压对大肠杆菌和金黄色葡萄球菌的杀菌研究

发布时间：2018-01-09 07:28

  本文关键词：二氧化碳辅助超高压对大肠杆菌和金黄色葡萄球菌的杀菌研究　出处：《中国农业大学》2016年博士论文　论文类型：学位论文

  更多相关文章： 二氧化碳 超高压 大肠杆菌 金黄色葡萄球菌 杀菌机制 自然菌群

【摘要】：本文以脱毒型食源性致病菌革兰氏阴性菌(G-)大肠杆菌(Escherichia coli)和革兰氏阳性菌(G+)金黄色葡萄球菌(Staphylococcus aureus)为目标菌,研究二氧化碳(Carbon dioxide, CO2)辅助超高压(High pressure processing, HPP)对E. coli和S. aureus的杀菌效果,并通过从细胞水平和分子水平分析了可能杀菌机制,最后将其应用于黄瓜汁和苹果汁自然菌群的杀灭。主要研究内容如下：(1)不同因素对C02辅助HPP (CO2+HPP)处理E. coli和S. aureus的影响。真空顶空充入20%CO2对HPP有较强的辅助杀菌作用；两种菌的稳定期比对数期具有更强的抗性；与磷酸盐缓冲液相比,黄瓜汁和苹果汁体系对两种菌有明显的保护作用；HPP处理过程中C02溶解在菌液中最大浓度为0.7 mmol;(2) CO2+HPP对E. coli和S. aureus的杀菌动力学。CO2+HPP对E. coli和S. aureus的杀菌动力学曲线最适拟合模型为Biphasic,20% CO2与200 MPa/3 min、300 MPa/3min和400 MPa/3min辅助杀菌效果显著；(3) CO2+HPP对E. coli和S. aureus细胞形态和结构的影响。经过CO2+HPP处理,细胞褶皱变形,细胞膜透性增强,膜孔洞效应明显,内容物大量流出,细胞质产生空洞现象,呈均匀聚集状态,胞内蛋白变性；显著抑制E. coli胞内亮氨酸芳基酰胺酶和萘酚-AS-BI-磷酸水解酶的活性,以及S. aureus胞内α-葡萄糖苷酶的活性；(4) CO2+HPP对E. coli蛋白组的影响。E. coli经HPP处理后有大量蛋白下调表达,而经CO2+HPP处理后差异下调蛋白明显小于HPP处理样品,可能是由于真空顶空充入C02带来的酸逆境环境使细胞产生逆境相应,导致在大量蛋白上调表达,在一定程度上掩盖了由于CO2+HPP引起的蛋白下调；(5)从细胞水平和分子水平分析CO2+HPP可能的杀菌机制为：1)细胞水平。经过CO2+HPP处理后,由于气体在卸压过程中的爆破效应对细胞膜造成了严重损伤,细胞膜表面孔洞效应明显,细胞内容物大量流失,严重影响了细胞正常的生理代谢；2)分子水平。经过CO2+HPP处理后细胞能量代谢降低,胞内碳氮不平衡,细胞正常的生理活动无法得到维持；转录与翻译水平降低,蛋白合成代谢水平降低,遗传物质RNA和DNA的生成受到干扰；同时无法修复及重新合成受损的细胞膜,使膜主动运输能力减弱,胞内大量内容物流出;(6) 20% CO2+600 MPa/3 min处理黄瓜汁和苹果汁中自然菌群(包括菌落总数、霉菌酵母和乳酸菌群)在22天4℃贮藏过程中呈现“S”型生长趋势,可采用Logistic模型拟合；经过20%CO2+600 MPa/3 min处理的黄瓜汁可实现15天的贮藏,苹果汁在22天贮藏过程中均呈现微生物安全；贮藏过程中,20% CO2+HPP处理苹果汁中果胶甲酯酶和多酚氧化酶残存活性显著降低,苹果汁中的蔗糖转化酶和黄瓜汁中的脂肪氧合酶保持较高水平；20% CO2+HPP处理有利于保持黄瓜汁和苹果汁在贮藏过程中稳定性、颜色和香气物质；经过感官评价分析,经CO2+600 MPa/3 min处理的黄瓜汁在4℃贮藏7天后、苹果汁贮藏15天后的感官评价结果与未处理没有显著差异,依然可以被消费者接受。
[Abstract]:This type of food borne pathogens and detoxification of gram negative bacteria (G-) in Escherichia coli (Escherichia coli) and gram positive bacteria (G+) Staphylococcus aureus (Staphylococcus aureus) as target bacteria, study on carbon dioxide (Carbon dioxide, CO2 (High pressure) auxiliary ultra high pressure processing, HPP) and S. coli E. on bactericidal effect aureus, and from the cellular level and molecular level analyses the sterilization mechanism, finally applied to cucumber juice and Apple Juice natural bacteria killing. The main contents are as follows: (1) different factors on C02 assisted HPP (CO2+HPP) E. coli and S. aureus of the vacuum headspace filled with 20%CO2. Have strong bactericidal effect on HPP; stability of two kinds of bacteria has stronger resistance than the logarithmic phase; compared with phosphate buffer, cucumber juice and Apple Juice system has markedly protective effect on two bacteria; HPP C02 dissolution process in suspension concentration is 0.7 mmol; (2) killing curves on E. CO2+HPP.CO2+HPP bactericidal kinetics of coli and S. aureus of E. coli and S. aureus the best fitting model for the Biphasic, 20% CO2 and 200 MPa/3 min, 300 MPa/3min and 400 MPa/3min (auxiliary bactericidal effect; 3) the effect of CO2+HPP on E. coli and S. aureus cell morphology and structure. After CO2+HPP treatment, cell fold deformation, cell membrane permeability increased, membrane pore effect, the contents of a large outflow, cytoplasmic voids phenomenon, homogeneous aggregation state, intracellular protein denaturation; E. coli significantly inhibited the intracellular leucine Fang arylamidase and naphthol -AS-BI- phosphate hydrolase activity, and the activity of S. aureus intracellular alpha glucosidase; (4) the influence of CO2+HPP on E. coli protein.E. coli group treated by HPP has a large number of proteins down regulated by CO2+H, and The protein was less than that of HPP treated sample differences down after PP treatment may be due to vacuum headspace filling C02 caused by acid stress causes cells to produce the corresponding stress, resulted in a large amount of protein increased to a certain extent masked due to CO2+HPP protein expression; (5) from cell and molecular level analysis of the sterilization mechanism of CO2+HPP may: 1) the level of cells. After CO2+HPP treatment, because the gas caused serious damage in pressure relief blasting effect in the process of cell membrane, the pores on the surface of the cell membrane effect is obvious, the cell content loss, serious impact on cell metabolism; 2) molecular level after treatment with CO2+HPP cells. The decline of energy metabolism, intracellular carbon and nitrogen balance, the normal physiological activities of cells can not be maintained; reduce the level of transcription and translation, protein synthesis and metabolism level is reduced, and the genetic material RNA The formation of DNA interference; at the same time to repair and re synthesis of the cell membrane is damaged, weakened the membrane active transport ability, the intracellular content of logistics; (6) 20% CO2+600 MPa/3 min natural flora Apple Juice juice and cucumber (including the total number of colonies, mold, yeast and lactic acid bacteria) showed "S" type the growth trend in 22 days at a temperature of 4 DEG C during storage, can be fitted by Logistic model; after 20%CO2+600 MPa/3 min treated cucumber juice can achieve 15 days of storage, Apple Juice showed microbial safety in 22 days during storage; during storage, 20% CO2+HPP Apple Juice in pectin residual activity significantly, Apple Juice in conversion of sucrose fatty enzyme and oxygen synthase in cucumber juice to maintain a higher level; 20% CO2+HPP treatment is conducive to maintaining stability and Apple Juice cucumber juice during storage, color and fragrance Gas substances; after sensory evaluation and analysis, the sensory evaluation results of cucumber juice processed by CO2+600 MPa/3 min after 4 days storage for 7 days and apple juice storage for 15 days had no significant difference from that of untreated, and could still be accepted by consumers.

【学位授予单位】：中国农业大学
【学位级别】：博士
【学位授予年份】：2016
【分类号】：TS201.3
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本文编号：1400518