发酵香肠中乳酸菌的分离、筛选及其应用
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摘要
本土的自然发酵香肠中存在着优良的菌种资源,这些菌特别适合用于肉品发酵。尤其是乳杆菌,是最早也是最常用的发酵香肠微生物发酵剂。许多乳杆菌还是人体常见的益生菌。本论文以主要发酵特性、工艺安全性和益生性为主要指标对本地的两种自然发酵香肠中的乳酸菌进行了分离、筛选和应用研究,主要内容包括如下几个部分:
1.对两种自然发酵香肠的微生物和理化指标进行了分析,结果表明这两种发酵香肠有很好的生物安全性,适宜用做菌种分离样品。
2.对两种自然发酵香肠中耐酸、耐胆盐的乳酸杆菌进行富集,从中分离纯化乳酸杆菌184株。通过两级筛选,最终筛选出性能优良的乳酸杆菌3株。初步鉴定T10703、T10709为干酪乳酸杆菌,T20706为植物杆菌。这3株乳酸菌能耐受8%的NaCl和150mg/Kg亚硝酸盐,能产双乙酰,分解乳糖,不产生物胺,不产H2O2,CO2和H2S,不产黏液,它们符合发酵香肠发酵剂的基本要求。另外对金黄色葡萄球菌,大肠杆菌,沙门氏菌、单核细胞增生李斯特菌有很好的抑制作用。对高酸(pH2.5)的存活率都在80%以上,并对0.3%胆盐有很好耐受性,可活体摄入人体肠道,发挥其益生性,可以作为益生型发酵剂用于功能性发酵香肠的生产。3.性能测定和混合法发酵剂筛选实验表明,筛选出的3株菌均生长能力强、产酸快、生长温度适宜、不具备脂肪酶和蛋白酶活性,符合发酵香肠生产益生性乳酸菌筛选标准。T10703与T20706之间没有拮抗作用,可作为混合发酵剂用于发酵香肠的生产。
4.利用优选出的混合发酵剂制作发酵香肠。通过L9(34)正交试验和优化调整,得出了最佳优化发酵工艺条件:接种量107cfu/g,干酪乳杆菌T10703、植物乳杆菌T20706菌种配比2:l,发酵温度22-25℃,相对湿度90-95%,发酵时间1d;随后发酵温度16-17℃,相对湿度85%,发酵2d。
5.采用混合发酵剂进行实验,研究了发酵香肠生产过程中的主要理化和微生物变化。添加混合发酵剂的实验组中乳酸菌在整个生产过程始终处于支配地位;乳酸菌混合发酵剂的加入显著提高了发酵香肠pH、水分含量、Aw的下降速率,并使其最终达到了更低值,提高了产品的质量,保证了其生物安全性,另外,实验组亚硝等残留低。组结合感官评价,认为该混合发酵剂可用于发酵香肠生产,其益生性效果有待进一步研究。
In the local natural fermented sausages, there are good bacteria resources, which are particularly suitable for meat fermentation. Particularly Lactobacillus, it is the first and the most popular microbial start culture used in fermented sausages. Many Lactobacillus are probiotics to human. In this paper, lactic acid bacteria stains were isolated from two local natural fermented sausages by using fermentation characteristics, process safety and Probiotic Properties as the main indicator. Finaly, the applied research were carried out. The main research contents are as follows:
1. Microorganisms and the physical and chemical indicators of two types of natural fermented sausages are studied, and the results show that both fermented sausages have a good bio-safety, and can be used as appropriate sample for strain separation.
2. A total of 184 lactobanillus stains were isolated and purified from two types of natural fermented dry sausages after enrichment by capability of endure low acid, capability of endure Bile Salts. After two steps selection, finally, three strains with excellent characteristies were obtained. By preliminary identification,T10703,T10709 were Lactobacillus casei, T20706 was Lactobacillus plantarum. These three lactic acid bacteria all resistanted to the 8% NaCl and 150 mg / Kg nitrite, producted diacetyl, decomposited lactose, did not produce biogenic amine ,H2O2, CO2 , H2S and slime, and they are consistent with the basic requirements for fermented sausage fermentation agent. In addition, they can inhibit Staphylococcus aureus, Escherichia coli, Salmonella, Listeria monocytogenes bacteria.They have a more than 80% survival rate in the high-acid (pH 2.5) situation, and have a good tolerance to 0.3% bile salt. in vivo uptake of human intestinal, naturally play its benefits ,and can be as beneficial fermentation agent for the Health of functional fermented sausage production.
3. Mixed performance measurement and selection of fermentation experiments show that the three selected bacteria strains all have strong growth ability, quickly acid forming ability , suitable growth temperature, and do not have lipase and protease activity. They all meet the screening criteria of probiotic lactobanillus stains.there are no antagonism between T10703 and T20706, and can be used as mixed start culture for the production of fermented sausages.
4. Use of selected mixed culture products fermented sausages.The best optimum fermentation conditions were gained by orthogonal test L9(34) and optimization.The result showed that, the optimum proportion of Lactobacillus casei T10703 and Lactobacillus plantarum T20706 was 2:1, the viable cell counts was 107cfu/g, temperature of fermentation 22-25℃, and relative humidity was 90-95%, fermented one day; then 16-17℃,relative humidity 85%, fermentation two days.
5. Fermented sausages were produced with selected srtains under the best optimum fermentation conditions, and main microbial and physic-chemical changes were studied. lactic acid bacteria in the test group added mixed start cultures was in a dominant position during the entire production process; the addition of mixed lactic acid bacteria culture significantly speed up the droping rate of pH, moisture content, and Aw, and made them down to a lower value,which improved the quality of the product and ensured its biosafety. Besides,the residues of nitrite in test group was lower.Combined with the sensory evaluation, that the selected mixed culture can be used as fermented sausages start culture, but the action effect of Probiotics needs further research.
1.对两种自然发酵香肠的微生物和理化指标进行了分析,结果表明这两种发酵香肠有很好的生物安全性,适宜用做菌种分离样品。
2.对两种自然发酵香肠中耐酸、耐胆盐的乳酸杆菌进行富集,从中分离纯化乳酸杆菌184株。通过两级筛选,最终筛选出性能优良的乳酸杆菌3株。初步鉴定T10703、T10709为干酪乳酸杆菌,T20706为植物杆菌。这3株乳酸菌能耐受8%的NaCl和150mg/Kg亚硝酸盐,能产双乙酰,分解乳糖,不产生物胺,不产H2O2,CO2和H2S,不产黏液,它们符合发酵香肠发酵剂的基本要求。另外对金黄色葡萄球菌,大肠杆菌,沙门氏菌、单核细胞增生李斯特菌有很好的抑制作用。对高酸(pH2.5)的存活率都在80%以上,并对0.3%胆盐有很好耐受性,可活体摄入人体肠道,发挥其益生性,可以作为益生型发酵剂用于功能性发酵香肠的生产。3.性能测定和混合法发酵剂筛选实验表明,筛选出的3株菌均生长能力强、产酸快、生长温度适宜、不具备脂肪酶和蛋白酶活性,符合发酵香肠生产益生性乳酸菌筛选标准。T10703与T20706之间没有拮抗作用,可作为混合发酵剂用于发酵香肠的生产。
4.利用优选出的混合发酵剂制作发酵香肠。通过L9(34)正交试验和优化调整,得出了最佳优化发酵工艺条件:接种量107cfu/g,干酪乳杆菌T10703、植物乳杆菌T20706菌种配比2:l,发酵温度22-25℃,相对湿度90-95%,发酵时间1d;随后发酵温度16-17℃,相对湿度85%,发酵2d。
5.采用混合发酵剂进行实验,研究了发酵香肠生产过程中的主要理化和微生物变化。添加混合发酵剂的实验组中乳酸菌在整个生产过程始终处于支配地位;乳酸菌混合发酵剂的加入显著提高了发酵香肠pH、水分含量、Aw的下降速率,并使其最终达到了更低值,提高了产品的质量,保证了其生物安全性,另外,实验组亚硝等残留低。组结合感官评价,认为该混合发酵剂可用于发酵香肠生产,其益生性效果有待进一步研究。
In the local natural fermented sausages, there are good bacteria resources, which are particularly suitable for meat fermentation. Particularly Lactobacillus, it is the first and the most popular microbial start culture used in fermented sausages. Many Lactobacillus are probiotics to human. In this paper, lactic acid bacteria stains were isolated from two local natural fermented sausages by using fermentation characteristics, process safety and Probiotic Properties as the main indicator. Finaly, the applied research were carried out. The main research contents are as follows:
1. Microorganisms and the physical and chemical indicators of two types of natural fermented sausages are studied, and the results show that both fermented sausages have a good bio-safety, and can be used as appropriate sample for strain separation.
2. A total of 184 lactobanillus stains were isolated and purified from two types of natural fermented dry sausages after enrichment by capability of endure low acid, capability of endure Bile Salts. After two steps selection, finally, three strains with excellent characteristies were obtained. By preliminary identification,T10703,T10709 were Lactobacillus casei, T20706 was Lactobacillus plantarum. These three lactic acid bacteria all resistanted to the 8% NaCl and 150 mg / Kg nitrite, producted diacetyl, decomposited lactose, did not produce biogenic amine ,H2O2, CO2 , H2S and slime, and they are consistent with the basic requirements for fermented sausage fermentation agent. In addition, they can inhibit Staphylococcus aureus, Escherichia coli, Salmonella, Listeria monocytogenes bacteria.They have a more than 80% survival rate in the high-acid (pH 2.5) situation, and have a good tolerance to 0.3% bile salt. in vivo uptake of human intestinal, naturally play its benefits ,and can be as beneficial fermentation agent for the Health of functional fermented sausage production.
3. Mixed performance measurement and selection of fermentation experiments show that the three selected bacteria strains all have strong growth ability, quickly acid forming ability , suitable growth temperature, and do not have lipase and protease activity. They all meet the screening criteria of probiotic lactobanillus stains.there are no antagonism between T10703 and T20706, and can be used as mixed start culture for the production of fermented sausages.
4. Use of selected mixed culture products fermented sausages.The best optimum fermentation conditions were gained by orthogonal test L9(34) and optimization.The result showed that, the optimum proportion of Lactobacillus casei T10703 and Lactobacillus plantarum T20706 was 2:1, the viable cell counts was 107cfu/g, temperature of fermentation 22-25℃, and relative humidity was 90-95%, fermented one day; then 16-17℃,relative humidity 85%, fermentation two days.
5. Fermented sausages were produced with selected srtains under the best optimum fermentation conditions, and main microbial and physic-chemical changes were studied. lactic acid bacteria in the test group added mixed start cultures was in a dominant position during the entire production process; the addition of mixed lactic acid bacteria culture significantly speed up the droping rate of pH, moisture content, and Aw, and made them down to a lower value,which improved the quality of the product and ensured its biosafety. Besides,the residues of nitrite in test group was lower.Combined with the sensory evaluation, that the selected mixed culture can be used as fermented sausages start culture, but the action effect of Probiotics needs further research.
引文
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