【作者】 王秀丽；

【导师】 韩文瑜；

【作者基本信息】 吉林大学 ， 微生物学， 2009， 硕士

【摘要】 乳链菌肽(Nisin)亦称为乳酸链球菌素,是由乳酸乳球菌产生的一种小分子多肽抗菌物质,它可以抑制几乎所有革兰氏阳性菌的生长。由于它对人体无毒,并在酸性条件下极其稳定,因而被公认为天然食品防腐剂。本文探讨了工业废料在乳酸乳球菌发酵中的应用。通过单因素实验对影响乳链菌肽收率的培养基组分玉米浆、糖蜜、K2HPO4和MgSO4的浓度进行了筛选,实验表明,当糖蜜为30.0 g/L、玉米浆为200.0 g/L、K2HPO4为9.0 g/L、MgSO4为1.0 g/L时,乳链菌肽收率较高。应用Box-Behnken设计得到乳链菌肽最佳发酵培养基配方为:玉米浆243.35 g/L,糖蜜26.77 g/L,MgSO40.44 g/L,K2HPO48.56 g/L时。在此培养条件下,乳链菌肽的收率为1316.0IU/mL。本文对乳链菌肽普通发酵培养基及培养条件优化进行了优化。利用单因素实验筛选了乳酸乳球菌发酵培养基的成分。在单因素实验的基础上利用Plackett–Burman实验对影响乳链球菌发酵产生乳酸链菌肽的培养基组分进行关键因素筛选,发现酵母浸粉、葡萄糖和K2HPO4为显著影响因素。利用响应面建立了影响乳酸菌肽产率的二次多项数学模型,探讨因素间的交互作用。由实验结果得最优培养基成分为:葡萄糖24.80 g/L,酵母浸粉22.35 g/L, K2HPO413.65 g/L,采用优化培养基得到最优乳链菌肽得率为2115.0 IU/mL。通过单因素对乳链菌肽发酵过程中的pH值、接种量、转速和温度进行优化,实验结果表明最佳的培养条件为:培养温度36.0℃,摇床转速160r/min,接种量为5%,培养基pH值为7.0。对乳酸乳球菌进行了发酵罐扩大培养,分别进行了好氧分批发酵和厌氧分批发酵的研究。从实验结果知,氧气有助于乳酸乳球菌的生长,在溶氧维持在30 %时,乳酸乳球菌细胞收率是厌氧条件下的4.92倍。在有氧条件下,乳链菌肽收率略高于无氧发酵的收率。更多还原

【Abstract】 Nisin, a polypeptide produced by Lactococcus lactis, restrains most Gram-positive bacteria , especially spores of bacterium. Nisin is cosidered as a safe preservative because of its non-toxic and stability at low pH. The study on Nisin began at 1950s and there are more than 50 countries which permit Nisin as preservative. The standard of Nisin was issued in China these years.The industrial wastes were used as the medium components in the fermentation of Lactococcus lacits subsp.Lactis CGMCC1.2030. It is shown that Nisin productivity was mainly influenced by corn steep liquor, molasses,K2HPO4 and MgSO4 with single factor experiment and the optimal levels were 200 g/L, 30 g/L, 9 g/L and 1 g/L respectively. Box-Behnken design and response surface analysis methods were applied to confirm optimum concentration of mainly effect factor.The results showed that the optimum medium culture was corn steep liquor 243.35 g/L,molasses 26.77 g/L,K2HPO4 8.56 g/L,MgSO4 0.44 g/L.The predictive Nisin productivity is 1334 IU/mL with optimal medium. The validation experiments with optimal medium were conducted 5 times and the average Nisin productivity is 1315.6IU/mL, which confirm the model could fit the fermentation conditions. The cost could be decreased significantly by the industrial waste as medium components to produce Nisin.With the Plackett-Burman method , the factors affected the Nisin titer were studied , and three important components were got , which are glucose , yeast extract and K2HPO4. Box-Behnken experiment were used to optimize the medium , the optimal medium were got as : glucose 24.80 g/L , yeast extract 22.35 g/L,K2HPO4 13.65 g/L. Under the optimal condition , the max predicted Nisin titer is 2157 IU/mL ,while the experimental results 2115 IU/mL .The effects of the medium pH,temperature,rotate speed,inoculum size were studied in this work . The result suggests the optimal culture condition as : medium pH 7 , temperature 36℃, rotate speed 160 r/min , inoculum size 5 %.The pilot-scale fermentation was carried in the Biomate 10 L fermenter . Anaerobic and aerobic batch fermentation was studied. The biomass and Nisin titer were higher in aerobie fermentation. The biomass with aerobic batch fermentation were 4.92 times compared with anaerobic fermentation,while the Nisin titer of aerobic batch fermentation was little higher than that of anaerobic fermentation.更多还原