摘要
本文对不加中和剂条件下德氏乳杆菌高产菌株的筛分、固定化德氏乳杆菌发酵条件的优化以及吸附固定化德氏乳杆菌发酵耦合离子交换树脂分离提取工艺过程进行了系统的研究。
1．从两株出发菌株LD-1、LD-2中选出不加中和剂条件下可产乳酸9.4g／L的LD-1作为出发菌株。利用德氏乳杆菌LD-1自身的发酵产物乳酸的反馈抑制作用，通过36h不加中和剂耐酸培养，淘汰掉不耐酸的菌株后，添加CaCO_3使发酵液中残留的耐酸菌株复壮、增殖，建立了一种快速、简单的选育不加中和剂时高产乳酸菌的方法。通过耐酸驯化，得到一株不加中和剂时产乳酸14g／L的德氏乳杆菌LD-1-1。
2．利用分批“钙盐法”发酵，对使用木炭等载体物理吸附固定化德氏乳酸杆菌发酵进行了研究。结果表明，利用木炭作为吸附载体效果最好，在分批“钙盐法”发酵过程中，得到了固定化培养的最佳条件：培养温度48℃，木炭的最适添加量为20g／L，底物糖化液的最适浓度为100g／L，麸皮的最适添加量为20g／L，在固定化德氏乳酸杆菌发酵的最佳工艺条件下，72h残糖耗尽，对糖的转化率94％，同样条件下游离细胞发酵72h残糖耗尽，对糖的转化率93.3％，利用木炭吸附固定化德氏乳杆菌发酵达到现有工艺水平，可以用于耦合乳酸发酵与提取技术的进一步研究。
3．研究了树脂的特性。结果表明：改M1_阴离子树脂不易破损、脱色、不吸附葡萄糖等，对LD-1-1无毒性，树脂交换吸附量不受发酵液中的葡萄糖、菌体的影响，适用于耦合发酵与提取技术。
4．研究了耦合固定化发酵与离子树脂提取的工艺过程，确定了耦合发酵与提取的最优工艺：大米糖化液140g／L、乳酸达6g／L时开始循环发酵液提取、提取流速为240ml／h，在最优工艺下，发酵周期缩短到60h，将乳酸从离子交换柱上洗脱，并精制、检验，乳酸收率达到86％，乳酸的生产率达到2.01g．L~(-1)．h~(-1)，各项理化指标合格，色度为1号，优于现有工艺。
In this paper, the breeding of lactic acid-resistant stains, culture condition on the immobilized fermentation and coupling technology of lactic fermentation and Extractive was studied.
The most lactic acid-resistant stain Lactobacillus delbrueckii LD-1 was selected from two original L.delbrueckii LD-1,LD-2.LD-1-1 could resistant 14g/L lactic acid in the ferment medium through Lactic- resistant domestication. A simple and fast method was built for breeding lactic-resistant stain utilizing lactic acid feedback and resistant to stain. In this method not lactic acid-resistant stain was eliminated through 36 hour cultivate without counteract reagent added and lactic acid-resistant stain was rejuvenated and multiplicated with CaCO3 added after 36 hour.
By means of convention batch fermentation, immobilized fermentation was studied. The result showed that carbon was the best material. A maximum yield was obtain under the following immobilized fermentation: temperature 48 C, carbon 20g/L, rice starch saccharification of 140g/L,bran 20g/L.On lactic acid optimum fermentation, the yield of 93.3g/L lactic acid, sucrose conversion of 94% after 72 hour. Under the same condition, free-cell fermentation yields of 93.3g/L after 72 hour. Immobilized fermentation could substitute free-cell fermentation in the coupling technology of lactic acid fermentation and extractive.
The characteristic of anion-exchange resin was studied. It showed that the alter-M] anion-exchange resin few were damaged, discolored, absorbed glucose and pigment, etc. At the same time, alter-Mi didn't affect the cell growth. Glucose and cell in the medium did not effected exchangeable amount of Lactic acid. The alter-Mi anion-exchange could be used to coupling technology of lactic fermentation and extractive.
The coupling technology of carbon immobilized lactic fermentation and alter-Mi anion-exchange extractive were studied. The optimum conditions were ascertained. The maximum yield was obtain under the following conditions: glucose 140g/L, exacting when the concentration of lactic acid was 6g/L, the velocity of extractive flow is 240ml/h, Under the optimum techniques condition, the new technology reduced time to 60 hour. Product lactic acid was pulled off from alter-M1 anion-exchange and refined. The reclaiming rate is 86%, productivity is 2.01g.L-1.h-1, all physics and chemical character of product up to grade. Chromed is below 1 degree and was evidently better than the now technology.
引文
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