阅读说明：本技术 一种发酵生产氨基葡萄糖的方法 (Method for producing glucosamine by fermentation ) 是由 卢健行 吴祥舟 刘长峰 马善丽 卢建功 卢建智 于 2019-11-12 设计创作，主要内容包括：本发明公开了一种发酵生产氨基葡萄糖的方法,将大肠杆菌、酿酒酵母、葡萄球菌进行活化后制得混合发酵剂；将混合发酵剂接种至含有葡萄糖的发酵培养基中在发酵罐中进行发酵培养,培养温度为35-38℃,培养时间为35-48h,得到含有氨基葡萄糖的发酵液,经分离纯化后得到氨基葡萄糖产品。本发明通过混合发酵剂实现高效发酵生产氨基葡萄糖,半胱氨酸的加入可以降低发酵过程中谷氨酸的产生速度,提高氨基葡萄糖的合成速率。(The invention discloses a method for producing glucosamine by fermentation, which comprises the steps of activating escherichia coli, saccharomyces cerevisiae and staphylococcus to prepare a mixed starter; inoculating the mixed starter to a fermentation medium containing glucose, performing fermentation culture in a fermentation tank at 35-38 deg.C for 35-48h to obtain fermentation liquid containing glucosamine, and separating and purifying to obtain glucosamine product. The invention realizes the high-efficiency fermentation production of the glucosamine by mixing the leaven, and the addition of the cysteine can reduce the production speed of the glutamic acid in the fermentation process and improve the synthesis rate of the glucosamine.)

1. A method for producing glucosamine by fermentation is characterized in that: the method comprises the following steps: activating Escherichia coli, Saccharomyces cerevisiae and Staphylococcus to obtain mixed starter; inoculating the mixed starter to a fermentation medium containing glucose, performing fermentation culture in a fermentation tank at 35-38 deg.C for 35-48h to obtain fermentation liquid containing glucosamine, and separating and purifying to obtain glucosamine product.

2. A method of fermentative production of glucosamine according to claim 1, characterized in that: the mixed starter is prepared from (1-2) Escherichia coli, Saccharomyces cerevisiae and Staphylococcus according to the volume ratio of 3 (0.5-0.8).

3. A method of fermentative production of glucosamine according to claim 2, wherein: the activation of the escherichia coli, the saccharomyces cerevisiae and the staphylococcus is to inoculate strains in a seed culture medium, the culture temperature is controlled to be 36-38 ℃, the shaking table speed is 220-270rpm, and the culture time is 12-14 h.

4. A method of fermentative production of glucosamine according to claim 3, wherein: the seed culture medium comprises 11.5g/L of peptone, 20g/L of yeast extract powder, 10g/L of cysteine, 5g/L of sodium chloride, 2g/L of ammonium sulfate, 5.5g/L of glycerol and pH 6.5-7.0.

5. A method of fermentative production of glucosamine according to claim 1, characterized in that: the fermentation medium comprises: 25g/L of glucose, 20g/L of yeast extract powder, 3g/L of sodium nitrate, 1.05g/L of dipotassium phosphate, 0.1g/L of sodium chloride, 0.5g/L of magnesium sulfate, 0.03g/L of ferrous sulfate, 3.2g/L of lactose, 5.5g/L of glycerol and 6.5-7.0 of pH.

6. A method of fermentative production of glucosamine according to claim 1, characterized in that: the rotation speed of the shaking table in the fermentation tank is 220-300 rpm.

7. A method of fermentative production of glucosamine according to claim 1, characterized in that: the fermentation process adopts constant-speed feeding fermentation, and the feeding speed of glucose is 3-4L/h.

8. A method of fermentative production of glucosamine according to claim 5, wherein: the pH of the fermentation medium is adjusted to 6.7-7.0 by ammonia water.

Technical Field

The invention belongs to the technical field of biological fermentation, and particularly relates to a method for producing glucosamine by fermentation.

Background

Glucosamine (GleN) is an important hexosamine formed by substituting one hydroxyl group of glucose with an amino group, and there are two main types of glucosamine on the market today, one is glucosamine hydrochloride and the other is glucosamine sulfate. D-Glucosamine Hydrochloride (D-Glucosamine Hydrochloride), molecular formula C₆H₁₃NO₅HCl, a white crystal, odorless, slightly sweet, easily soluble in water, slightly soluble in methanol, insoluble in organic solvents such as ethanol, has important physiological functions for human body, participates in liver and kidney detoxification, plays a role in anti-inflammation and liver protection, has good curative effect on rheumatic arthritis and gastric ulcer, and is a main raw material for synthesizing antibiotics and anticancer drugs; can also be used in food, cosmetic and feed additive. Glucosamine hydrochloride is extracted from natural chitin, is a marine biological agent, and is the main component of chondroitin sulfate. It can promote the synthesis of mucopolysaccharide, raise the viscosity of joint synovial fluid, improve the metabolism of joint cartilage, promote the repair of joint cartilage and has obvious antiphlogistic and analgesic effects. It has the effect of promoting the injection efficiency of antibiotics, and can be used as nutritional supplement for diabetic patients.

The current methods for producing glucosamine mainly comprise an acid hydrolysis method, an enzymolysis method and a microbial fermentation method. The production raw materials of the acid hydrolysis method and the enzymolysis method are from exoskeletons of fishes, shrimps, crabs and the like, and glucosamine is obtained by extracting chitin and chitosan and then carrying out acidolysis or enzymolysis; however, a large amount of concentrated hydrochloric acid is needed in the acid hydrolysis process, which can cause serious industrial pollution; the enzymolysis method is to degrade the exoskeleton of fishes, shrimps and crabs by using chitosan, and has low process efficiency and higher production cost, so that the search for a proper microbial fermentation method to realize the industrial production of glucosamine is the current environmental and market demand.

Disclosure of Invention

In order to make up the defects of the prior art, the invention provides the method for producing the glucosamine by fermentation, which has the advantages of high production efficiency, low cost and simple operation.

The invention is realized by the following technical scheme:

a method for producing glucosamine by fermentation is characterized in that: the method comprises the following steps: activating Escherichia coli, Saccharomyces cerevisiae and Staphylococcus to obtain mixed starter; inoculating the mixed starter to a fermentation medium containing glucose, performing fermentation culture in a fermentation tank at 35-38 deg.C for 35-48h to obtain fermentation liquid containing glucosamine, and separating and purifying to obtain glucosamine product.

Preferably, the mixed starter is prepared from the Escherichia coli, the saccharomyces cerevisiae and the staphylococcus according to the volume ratio of 3 (0.5-0.8) to 1-2, and the mixed starter can enhance the fermentation activity of the Escherichia coli.

Further, the activation of the escherichia coli, the saccharomyces cerevisiae and the staphylococcus is to inoculate strains in a seed culture medium, the culture temperature is controlled to be 36-38 ℃, the shaking table speed is 220-270rpm, and the culture time is 12-14 h.

Preferably, the seed culture medium comprises 11.5g/L of peptone, 20g/L of yeast extract powder, 10g/L of cysteine, 5g/L of sodium chloride, 2g/L of ammonium sulfate, 5.5g/L of glycerol and pH 6.5-7.0.

Preferably, the fermentation medium comprises: 25g/L of glucose, 20g/L of yeast extract powder, 3g/L of sodium nitrate, 1.05g/L of dipotassium phosphate, 0.1g/L of sodium chloride, 0.5g/L of magnesium sulfate, 0.03g/L of ferrous sulfate, 3.2g/L of lactose, 5.5g/L of glycerol and 6.5-7.0 of pH; the mixed starter is inoculated on a fermentation medium according to the inoculation amount of 10%.

Preferably, the rotation speed of the shaking table in the fermentation tank is 220-300 rpm.

Preferably, the fermentation process adopts constant-speed feeding fermentation, and the feeding speed of glucose is 3-4L/h.

Preferably, the pH of the fermentation medium is adjusted to 6.7-7.0 with ammonia water.

The invention has the beneficial effects that: the invention realizes the high-efficiency fermentation production of the glucosamine by the mixed leaven, the addition of the cysteine can reduce the production speed of the glutamic acid in the fermentation process, improve the synthesis rate of the glucosamine, not only reduce the reaction cost, but also improve the yield of the glucosamine, and can realize the industrial production.

Detailed Description

The present invention will be described in further detail with reference to specific embodiments thereof to assist those skilled in the art in providing a more complete, accurate and thorough understanding of the inventive concept and aspects thereof, and the scope of the present invention includes, but is not limited to, the following examples, and any modifications in the details and form of the technical aspects thereof that fall within the spirit and scope of the present application are intended to be included therein.

The following examples all follow the following medium composition:

the seed culture medium comprises: 11.5g/L of peptone, 20g/L of yeast extract powder, 10g/L of cysteine, 5g/L of sodium chloride, 2g/L of ammonium sulfate, 5.5g/L of glycerol and 6.5-7.0 of pH;

the fermentation medium comprises: 25g/L of glucose, 20g/L of yeast extract powder, 3g/L of sodium nitrate, 1.05g/L of dipotassium phosphate, 0.1g/L of sodium chloride, 0.5g/L of magnesium sulfate, 0.03g/L of ferrous sulfate, 3.2g/L of lactose and 5.5g/L of glycerol.