阅读说明：本技术 一种谷氨酸发酵培养基制备方法 (Preparation method of glutamic acid fermentation medium ) 是由 李德衡 赵凤良 翟德亮 赵杰 于 2019-12-13 设计创作，主要内容包括：本发明属于氨基酸生产技术领域,公开了一种谷氨酸发酵培养基的制备方法,其包括如下步骤：取发酵培养基原料：葡萄糖,菌体蛋白浸膏,K<Sub>2</Sub>HPO<Sub>4</Sub>,MgSO<Sub>4</Sub>·7H<Sub>2</Sub>O,MnSO<Sub>4</Sub>·H<Sub>2</Sub>O,FeSO<Sub>4</Sub>·7H<Sub>2</Sub>O,VB<Sub>1</Sub>,生物素；将各原料搅拌均匀后,调节pH为6.5,121℃灭菌15min,自然冷却,制得发酵培养基。本发明制备方法中使用菌体蛋白酶解浸膏来替代酵母浸膏,大幅降低了成本,提高了菌体蛋白的附加值,而且产酸效率更高。(The invention belongs to the technical field of amino acid production, and discloses a preparation method of a glutamic acid fermentation medium, which comprises the following steps: taking fermentation medium raw materials: glucose, mycoprotein extract, K 2 HPO 4 ，MgSO 4 ·7H 2 O，MnSO 4 ·H 2 O，FeSO 4 ·7H 2 O，VB 1 Biotin; stirring the raw materials uniformly, adjusting pH to 6.5, sterilizing at 121 deg.C for 15min, and naturally cooling to obtain fermentation culture medium. The preparation method of the invention uses the mycoprotein enzymolysis extract to replace the yeast extract, thereby greatly reducing the costThe added value of the mycoprotein is improved, and the acid production efficiency is higher.)

1. A preparation method of a glutamic acid fermentation medium comprises the following steps:

1) taking fermentation medium raw materials: glucose, mycoprotein extract, K₂HPO₄，MgSO₄·7H₂O，MnSO₄·H₂O，FeSO₄·7H₂O，VB₁Biotin;

2) adding the raw materials into water, stirring, adjusting pH to 6.5, sterilizing at 121 deg.C for 15min, and naturally cooling to obtain fermentation culture medium.

2. The method according to claim 1, wherein the fermentation medium comprises the following raw materials at the following concentrations: 80g/L glucose, 20g/L mycoprotein extract and K₂HPO₄2g/L，MgSO₄·7H₂O 50mg/L，MnSO₄·H₂O3mg/L，FeSO₄·7H₂O 3 mg/L，VB₁10mg/L, biotin 7. mu.g/L.

3. The preparation method according to claim 1 or 2, characterized in that the mycoprotein extract is prepared by the following processes:

step 1) separation of thalli: separating glutamic acid fermentation liquor by a disc separator, and collecting thallus precipitate;

step 2) high-pressure homogenization: adding a potassium dihydrogen phosphate aqueous solution into the thallus sediment to prepare a thallus cell suspension with the concentration of 100-;

step 3) centrifugation: stopping homogenizing, centrifuging at 1000rpm for 3-5min, and collecting upper layer liquid and precipitate;

step 4), ultrasonic treatment: adding 5-10 times of purified water into the precipitate, stirring, and treating with ultrasonic wave for 20-30 min;

step 5) enzymolysis: adjusting the temperature to 45 ℃, hydrolyzing by adopting neutral protease for 6-9 h;

step 6) evaporation and concentration: heating to 90-100 ℃, inactivating enzyme for 3-5min, mixing with the upper layer liquid obtained in the step 3), and finally evaporating and concentrating to obtain a mycoprotein extract with the dry weight part of 60-70%.

4. The method as claimed in claim 3, wherein the rotation speed of the disk centrifuge is 4000-5000rpm, and the centrifugation time is 3-5 min.

5. The method according to claim 3, wherein the concentration of the aqueous solution of potassium dihydrogen phosphate is 5 to 10 g/L.

6. The method of claim 3, wherein the homogenization parameters are: homogenizing at 25 deg.C under 90Mpa for 3 times.

7. The method according to claim 3, wherein the ultrasonic wave has a frequency of 20 kHz.

8. The method according to claim 3, wherein the neutral protease is added in an amount of 500u enzyme activity/g dry matter.

9. A fermentation medium obtained by the production method according to any one of claims 1 to 8.

Technical Field

The invention belongs to the technical field of amino acid production, and particularly relates to a preparation method of a glutamic acid fermentation medium.

Background

The waste thallus from glutamic acid fermentation is a by-product in the production process of monosodium glutamate, is a single-cell protein separated in the process of extracting and refining monosodium glutamate, and has an annual output of up to millions of tons. In order to save resources, most of monosodium glutamate production enterprises sell glutamic acid mycoprotein as feed, and although certain economic benefit is obtained, the application value of the glutamic acid mycoprotein is not completely excavated. The analysis of the nutrient components of the mycoprotein shows that the mycoprotein has rich protein content, complete amino acid components, rich vitamins, nucleic acid, polysaccharide and the like. Therefore, it is important to search for glutamic acid mycoprotein as a raw material and develop a byproduct with higher value. Currently, the research in this area mainly focuses on developing mycoprotein into animal feed, biological fertilizer, seasoning, fermentation medium preparation and the like.

The cost of a fermentation culture medium in a process for preparing glutamic acid by fermentation is a core problem restricting the development of enterprises, and how to reduce the cost is the research direction of people. Yeast powder is used as a nitrogen source of a glutamic acid fermentation medium, the price is high, and in the aspect of byproducts, the fermentation residual thalli are the most main byproducts. In a general thallus treatment method, thallus is simply treated and then used as an organic fertilizer, so that the added value of the product is reduced, and the resource is greatly wasted. Therefore, in order to reduce the production cost and realize green production, the waste mycoprotein from fermentation is treated and used as the fermentation raw material, so that the recycling of resources is realized.

The optimization of a fermentation culture medium in a process for preparing glutamic acid by fermentation is an important factor for improving the fermentation efficiency, and the prior patent technology of the applicant 'a method for preparing the fermentation culture medium by fermenting waste thalli with glutamic acid' improves and optimizes the culture medium in the prior art, and comprises 15% of sorghum straw hydrolysate, 12% of thalli hydrolysate, 5% of glucose, 1.2% of rice bran extract, 1% of corn steep liquor, 0.02% of shell powder, 0.02% of ferrous sulfate heptahydrate, 0.02% of magnesium sulfate heptahydrate, 0.01% of potassium dihydrogen phosphate and the balance of water; the culture medium reduces the cost, but has fermentation nitrogen sources and carbon source substances with more pigments, high viscosity and more impurities, so that the fermentation process is difficult to control, and the instability of fermentation and the difficulty of separation and extraction are easily caused; in addition, hydrochloric acid is adopted in the method for treating mycoprotein in the patent document, which is easy to damage amino acid, and the nutritive value of hydrolysate is low. Further research on fermentation media is continued, and the aim is to develop a fermentation medium with high fermentation efficiency and low cost.

The compound amino acid obtained by enzymolysis of the thalli has complete components, also contains rich vitamins and the like, is very suitable for being used as the components of an amino acid fermentation culture medium, can replace expensive yeast extract, vitamins and other components, and reduces the fermentation cost; however, how to obtain an enzymolysis thallus product with high nutritive value, and the quality is controllable, and the stability of fermentation efficiency is maintained is a technical problem that needs to be solved.

Disclosure of Invention

In order to overcome the defects of the prior art, the invention provides a preparation method of a glutamic acid fermentation culture medium, which uses a mycoprotein enzymolysis extract to replace a yeast extract, greatly reduces the cost, improves the additional value of mycoprotein and has higher acid production efficiency.

The invention is realized by the following scheme:

a preparation method of a glutamic acid fermentation medium comprises the following steps: taking fermentation medium raw materials: glucose, mycoprotein extract, K₂HPO₄，MgSO₄·7H₂O，MnSO₄·H₂O，FeSO₄·7H₂O，VB₁Biotin; stirring the above materials uniformly, adjusting pH to 6.5 and 121 deg.CSterilizing for 15min, and naturally cooling to obtain fermentation culture medium.

Further, the fermentation medium is prepared from the following raw materials in concentration: 80g/L glucose, 20g/L mycoprotein extract and K₂HPO₄2g/L，MgSO₄·7H₂O 50mg/L，MnSO₄·H₂O 3mg/L，FeSO₄·7H₂O 3mg/L，VB₁10mg/L, biotin 7. mu.g/L.

Further, the mycoprotein extract is prepared according to the process:

step 1) separation of thalli: separating glutamic acid fermentation liquor by a disc separator, and collecting thallus precipitate;

step 2) high-pressure homogenization: adding a potassium dihydrogen phosphate aqueous solution into the thallus sediment to prepare a thallus cell suspension with the concentration of 100-;

step 3) centrifugation: stopping homogenizing, centrifuging at 1000rpm for 3-5min, and collecting upper layer liquid and precipitate;

step 4), ultrasonic treatment: adding 5-10 times of purified water into the precipitate, stirring, and treating with ultrasonic wave for 20-30 min;

step 5) enzymolysis: adjusting the temperature to 45 ℃, hydrolyzing by adopting neutral protease for 6-9 h;

step 6) evaporation and concentration: heating to 90-100 ℃, inactivating enzyme for 3-5min, mixing with the upper layer liquid obtained in the step 3), and finally evaporating and concentrating to obtain a mycoprotein extract with the dry weight part of 60-70%.

Preferably, the rotation speed of the disc centrifuge is 4000-.

Preferably, the concentration of the potassium dihydrogen phosphate aqueous solution is 5 to 10 g/L.

Preferably, the homogenization parameters are: homogenizing at 25 deg.C under 90Mpa for 3 times.

Preferably, the frequency of the ultrasonic wave is 20 kHz.

Preferably, the addition amount of the neutral protease is 500u enzyme activity/g dry matter.

The invention also claims a fermentation medium obtained by the preparation method.

Compared with the prior art, the invention has the advantages that the following aspects are mainly included but not limited:

the invention does not adopt an acid or alkali hydrolysis mode, reduces pollution, avoids corrosion to equipment, only adopts an enzymolysis mode, has mild conditions, does not damage nutrient substances, has stable components of enzymolysis products and controllable quality, and is beneficial to the quality control of a large-scale fermentation system.

The invention adopts high-pressure homogenate treatment, can fully break cell walls and release intracellular protein, adopts monopotassium phosphate aqueous solution as buffer solution, can improve the breaking rate, and monopotassium phosphate can also be used as a component of a fermentation medium, thereby killing two birds with one stone;

the operation pressure of the homogenization treatment is crucial to cell disruption and protein release, the disruption dynamics and the disruption process effect homogenization pressure are integrated, and multiple tests are carried out, preferably 90 MPa; the cell breakage rate is rapidly increased 2 times before homogenization, the cell breakage rate is not obviously improved after the homogenization again, but after the homogenization for the 3 rd time, macromolecular protein is also cracked into micromolecular short peptide or free amino acid under the action of the homogenization, the content of amino acid nitrogen is increased by a certain amount, the degree of proteolysis is increased, the number of times of homogenization is continuously increased, the degree of proteolysis is not obviously increased, and certain damage is caused to equipment;

the supernatant liquid is centrifuged after homogenization, the level of amino acid nitrogen contained in the supernatant liquid is higher, and the partial product is not required to be subjected to enzymolysis treatment, so that the use amount of enzyme is reduced, and the cost is reduced; the precipitate is cell fragment and macromolecular protein, still contains a certain amount of protein in the cell wall fragment moreover, and some protein can adhere on the cell wall fragment in addition, through carrying out ultrasonic treatment to the precipitate for protein and polysaccharide on the cell fragment release, and the polysaccharide is utilized as nutrient by the bacterial strain, and after ultrasonic treatment, and then adopt neutral protease to carry out enzymolysis, the use amount and the action time of enzyme reduce by a wide margin, have practiced thrift the cost.

Compared with the conventional yeast extract nitrogen source, the fermentation culture medium disclosed by the invention is slightly improved in terms of fermentation thallus concentration and glutamic acid yield, so that the thallus protein extract prepared by the invention is richer and more comprehensive in nutrient content and is beneficial to utilization of microorganisms and acid production compared with the conventionally used nitrogen source yeast extract (the dry matter content is 65-70%).

Drawings

FIG. 1: influence of high-pressure homogenization times on the cell breakage rate of thalli;

FIG. 2: the effect of the number of high pressure homogenizations on the degree of hydrolysis.

Detailed Description

Those skilled in the art can modify the process parameters appropriately to achieve the desired results with reference to the disclosure herein. It is expressly intended that all such similar substitutes and modifications which would be obvious to one skilled in the art are deemed to be included in the invention. While the products and methods of this invention have been described in terms of preferred embodiments, it will be apparent to those of skill in the art that variations and modifications, or appropriate alterations and combinations, of the products and methods described herein may be made and utilized without departing from the spirit, scope, and spirit of the invention. For a further understanding of the present invention, reference will now be made in detail to the following examples.