CN109207533A

CN109207533A - A method of improving valine yield

Info

Publication number: CN109207533A
Application number: CN201811153962.5A
Authority: CN
Inventors: 徐庆阳; 徐达; 陈宁; 李燕军; 谢希贤; 马倩; 范晓光; 张成林
Original assignee: Tianjin University of Science and Technology
Current assignee: Tianjin University of Science and Technology
Priority date: 2018-09-30
Filing date: 2018-09-30
Publication date: 2019-01-15

Abstract

本发明提供一种利用蛋氨酸和氯化胆碱复配的方式提高缬氨酸发酵产量的方法，包括以下步骤：在缬氨酸的发酵培养时，葡萄糖初始添加加量为80g/L。当培养基中葡萄糖耗至10g/L左右时，开始流加含有1g/L氯化胆碱(相对于初始发酵液体积而言)的80％葡萄糖溶液，使培养基中残糖浓度保持在10‑15g/L；发酵过程中温度为32℃，通过流加25％氨水维持pH在7.0‑7.2，通过调节转速和通风将溶氧控制在20％左右。本发明将氯化胆碱与蛋氨酸以一定量配比之后加入到发酵培养基底物中，再将氯化胆碱添加到葡萄糖溶液中，使其随糖流加至发酵液中，以提高菌体生产缬氨酸的能力，发酵48h—52h，缬氨酸产量从80—85g/L提高到95—100g/L。The invention provides a method for improving valine fermentation yield by using methionine and choline chloride compound, comprising the following steps: during the fermentation and culture of valine, the initial addition amount of glucose is 80 g/L. When the glucose consumption in the medium reaches about 10g/L, the 80% glucose solution containing 1g/L choline chloride (relative to the initial fermentation broth volume) is added to keep the residual sugar concentration in the medium at 10 ‑15g/L; the temperature during the fermentation process is 32°C, the pH is maintained at 7.0‑7.2 by adding 25% ammonia water, and the dissolved oxygen is controlled at about 20% by adjusting the rotational speed and ventilation. In the present invention, choline chloride and methionine are added to the fermentation medium substrate in a certain proportion, and then choline chloride is added to the glucose solution, so that it is added to the fermentation liquid along with the sugar stream, so as to improve the bacterial cells. The ability to produce valine, fermentation 48h-52h, the valine yield increased from 80-85g/L to 95-100g/L.

Description

A method of improving valine yield

Technical field

The invention belongs to microorganism and fermentation engineering fields, and in particular to a method of improve valine yield.

Background technique

Valine is one of eight kinds of amino acid necessary to humans and animals body, is belonged to leucine, isoleucine Three kinds of branched-chain amino acids physiologically have a variety of regulatory functions, have consequence in human life's metabolism, except use It other than general auxotype dietetic formulation, is also largely used to produce all kinds of mixed amino acids, have extremely in terms of medical treatment It is widely applied.Valine is one of maximum kind of dosage in medicinal amino acid starting material.At present, Valine is main Microbial fermentation production is carried out using corynebacterium glutamicum, this production method has at low cost, mild condition and can advise greatly The advantages that mould produces, is a kind of very economical production method.Therefore, have using Production by Microorganism Fermentation valine important Meaning.

When using Production by Microorganism Fermentation valine, used bacterial strain is Corynebacterium glutamicum mutagenic fungi, the bacterial strain Slow growth, fermentation period is long, and rate of producing acid is slower, therefore, improves thalli growth speed, shortens the fermenting and producing period, accelerates Rate of producing acid is a problem to be solved.

Summary of the invention

Technical problem to be solved by the present invention lies in provide a kind of method for improving valine yield.

In order to solve the above technical problems, the technical scheme is that

A method of valine yield is improved, valine yield is improved by stream plus choline chloride, the specific steps are as follows:

(1) the methionine additive of 0.5-1.5g/L is added in the routine culture substrate of fermentation, by 48h-52h Fermented and cultured is verified the final acid that produces and is changed；

(2) the choline chloride additive of 0.5-1.5g/L is added in the routine culture substrate of fermentation, by 48h- 52h fermented and cultured is verified the final acid that produces and is changed；

(3) the methionine additive and 0.5-1.5g/L of 0.5-1.5g/L are added in the routine culture substrate of fermentation Choline chloride additive matched, by 48h -52h fermented and cultured, verify and final produce acid variation；

(4) in conventional fermentation process, the choline chloride for being 0.5-1.5g/L with sugar stream plus concentration, by 48h-52h Fermented and cultured, verify and final produce acid variation；

(5) the methionine additive and 0.5-1.5g/L of 0.5-1.5g/L are added in the routine culture substrate of fermentation Choline chloride additive matched, and simultaneously with sugar stream plus concentration be 0.5-1.5g/L choline chloride, by 48h- The fermented and cultured of 52h is verified the final acid that produces and is changed.

Preferably, the method for above-mentioned raising valine yield, in the step (1) in the routine culture substrate of fermentation The methionine additive that 1g/L is added ferments, and the final acid that produces improves 3-5g/L.

Preferably, the method for above-mentioned raising valine yield, in the step (2) in the routine culture substrate of fermentation The choline chloride additive that 1g/L is added ferments, and the final acid that produces improves 7-8g/L.

Preferably, the method for above-mentioned raising valine yield, in the step (3) in the routine culture substrate of fermentation The choline chloride additive of methionine additive and 1g/L that 1g/L is added ferments, and the final acid that produces improves 9-10g/L.

Preferably, the method for above-mentioned raising valine yield flows in the step (4) in conventional fermentation process with sugar Adding concentration is that the choline chloride of 1g/L ferments, and the final acid that produces improves 3-5g/L.

Preferably, the method for above-mentioned raising valine yield, in the step (5) in the routine culture substrate of fermentation The choline chloride additive of methionine additive and 1g/L that 1g/L is added is matched, and is simultaneously 1g/L with sugar stream plus concentration 80% glucose solution of choline chloride ferment, the final acid that produces improves 10-15g/L.

Above-mentioned routine culture substrate refers to using glucose as primary carbon source, be equipped with corn pulp, soybean meal hydrolysate, magnesium salts, The common fermentation medium of the microelements such as microcosmic salt and iron, manganese and other growth factors.

Structure of the invention has the advantages that:

Fermentation bottom is added after methionine and choline chloride are optimized proportion in the method for above-mentioned raising valine yield In object, while making fermentation liquid environment with certain density with sugar stream plus certain density choline chloride during the fermentation It is maintained in the case where choline chloride, provides enough active methyls for thalli growth and production acid, while maintaining thallus vigor, improve The acid producing ability of thallus.

Specific embodiment

To further illustrate the present invention, now by specific implementation example, the present invention will be described in detail.

Embodiment 1

When the fermented and cultured of valine, the initial additive amount of glucose is 80g/L, and the initial additive amount of methionine is 1g/L, when When glucose is consumed to 10g/L or so in culture medium, starts stream plus 80% glucose solution, be maintained at remaining sugar concentration in culture medium 10-15g/L；Temperature is 32 DEG C in fermentation process, maintains pH in 7.0-7.2 by stream plus 25% ammonium hydroxide, by adjust revolving speed and Ventilation controls dissolved oxygen 20% or so.Ferment 52h, and valine ultimate output is increased to 83g/L from 80g/L.

Embodiment 2

When the fermented and cultured of valine, the initial additive amount of glucose is 80g/L, and the initial additive amount of choline chloride is 1g/L. When glucose is consumed to 10g/L or so in culture medium, starts stream plus 80% glucose solution, keep remaining sugar concentration in culture medium In 10-15g/L；Temperature is 32 DEG C in fermentation process, maintains pH in 7.0-7.2 by stream plus 25% ammonium hydroxide, by adjusting revolving speed Dissolved oxygen is controlled 20% or so with ventilation.Ferment 52h, and valine ultimate output is increased to 85g/L from 80g/L.

Embodiment 3

When the fermented and cultured of valine, the initial additive amount of glucose is 80g/L, choline chloride and the initial additive amount of methionine It is all 1g/L.When glucose is consumed to 10g/L or so in culture medium, starts stream plus 80% glucose solution, make residual in culture medium Sugared concentration is maintained at 10-15g/L；Temperature is 32 DEG C in fermentation process, maintains pH in 7.0-7.2 by stream plus 25% ammonium hydroxide, leads to It overregulates revolving speed and ventilation controls dissolved oxygen 20% or so.Ferment 52h, and valine ultimate output is increased to 90g/ from 80g/L L。

Embodiment 4

When the fermented and cultured of valine, the initial additive amount of glucose is 80g/L.When glucose is consumed to 10g/L in culture medium When left and right, starts stream plus 80% glucose solution containing 1g/L choline chloride (for Preliminary fermentation liquid product), make Remaining sugar concentration is maintained at 10-15g/L in culture medium；Temperature is 32 DEG C in fermentation process, maintains pH to exist by stream plus 25% ammonium hydroxide 7.0-7.2 is controlled dissolved oxygen 20% or so by adjusting revolving speed and ventilation.Ferment 52h, and valine ultimate output is from 80g/L It is increased to 83g/L.

Embodiment 5

When the fermented and cultured of valine, the initial additive amount of glucose is 80g/L, choline chloride and the initial additive amount of methionine It is all 1g/L.When glucose is consumed to 10g/L or so in culture medium, start stream plus containing 1g/L choline chloride (relative to initial For fermentating liquid volume) 80% glucose solution, so that remaining sugar concentration in culture medium is maintained at 10-15g/L；Fermentation process medium temperature Degree is 32 DEG C, maintains pH in 7.0-7.2 by stream plus 25% ammonium hydroxide, by adjusting revolving speed and ventilation for dissolved oxygen control on 20% left side It is right.Ferment 52h, and valine ultimate output is increased to 98g/L from 80g/L.

The above is only a preferred embodiment of the present invention, it is noted that for the ordinary skill people of the art For member, various improvements and modifications may be made without departing from the principle of the present invention, these improvements and modifications are also answered It is considered as protection scope of the present invention.

Claims

1. a method for improving valine output, is characterized in that: improve valine output by stream adding choline chloride, concrete steps are as follows:

(1) Add 0.5-1.5g/L methionine supplement to the fermented conventional medium substrate, and verify the final acid production change after 48h-52h fermentation and culture;

(2) Add 0.5-1.5g/L of choline chloride additive to the fermented conventional medium substrate, and verify the final acid production change after 48h-52h fermentation culture;

(3) Add 0.5-1.5g/L methionine supplement and 0.5-1.5g/L choline chloride supplement to the fermented conventional medium substrate for proportioning, and after 48h-52h fermentation and culture, verify the final product acid changes;

(4) In the conventional fermentation process, choline chloride with a concentration of 0.5-1.5g/L was added with the sugar stream, and the final acid production change was verified after 48h-52h of fermentation culture;

(5) Add the methionine additive of 0.5-1.5g/L and the choline chloride additive of 0.5-1.5g/L in the fermented conventional culture medium substrate for proportioning, and at the same time, the concentration is 0.5- 1.5g/L choline chloride, after 48h-52h fermentation and culture, to verify the final acid production change.

2. The method for improving valine yield according to claim 1, characterized in that: in the step (1), a methionine additive of 1 g/L is added to the fermented conventional medium substrate to ferment.

3. the method for improving valine yield according to claim 1 is characterized in that: in described step (2), in the conventional medium substrate of fermentation, add the choline chloride additive of 1g/L to carry out fermentation.

4. the method for improving valine output according to claim 1, is characterized in that: in described step (3), in the conventional medium substrate of fermentation, add the methionine additive of 1g/L and the chlorine of 1g/L Fermentation with choline supplementation.

5. the method for improving valine output according to claim 1, is characterized in that: in the described step (4), in the conventional fermentation process, add the choline chloride that concentration is 1g/L with sugar stream and carry out fermentation .

6. the method for improving valine output according to claim 1 is characterized in that: in the described step (5), in the conventional medium substrate of fermentation, add the methionine additive of 1g/L and the chlorine of 1g/L The addition of choline chloride is proportioned, and at the same time, 80% glucose solution of choline chloride with a concentration of 1 g/L is added to the sugar stream for fermentation.