CN110241166A - It is a kind of promote riboflavin fermentation composition and its application - Google Patents

Abstract

It is a kind of promote riboflavin fermentation composition and its application, be mainly made of the raw material of following mass fraction: composite precursor 1-3%, compound amino acid 2-10% and tap water 50-70%；The composite precursor is mainly made of sodium gluconate, sorbic acid and malonic acid 1:2:2 in mass ratio；The compound amino acid is mainly made of tyrosine and phenylalanine 2.5:3.5 in mass ratio, promote application of the composition of riboflavin fermentation in riboflavin fermentation, the riboflavin fermentation the following steps are included: level-one, secondary seed medium preparation, the preparation of fermentation medium, first order seed culture, secondary seed culture, fermented and cultured, rush fermentation composition is constantly added during the fermentation, it ensure that the supply of various nutritional ingredients in fermentation process, it ensure that ferment effect, effectively increase the fermentation yield of riboflavin.

Description

It is a kind of promote riboflavin fermentation composition and its application

Technical field

The present invention relates to the preparation fields of riboflavin, and in particular to it is a kind of promote riboflavin fermentation composition and its fermenting In application.

Background technique

Riboflavin (Vitamin B2) is also known as vitamin B2, is a kind of important vitamin.It is promoted in human body cell One of redox important substance, also participates in the metabolism of sugar, protein, fat in vivo, and has and maintain normal vision function Effect just will affect the process of vivo biodistribution oxidation and dysbolism occur, bicker then occur if human body lacks riboflavin Dry, dry skin furfur of inflammation, blepharitis, conjunctivitis, cheilitis, glossitis, ear nose mucous membrane etc..

Riboflavin is mainly fermented by aerobic process and is made at present, and the fermentation process use is with bacillus subtilis (Bacillus subtilis) it is genetic engineering bacterium constructed by receptor, with fermentation period is short, at low cost, riboflavin production is high The advantages that.The biosynthesis and accumulation of riboflavin is a complicated metabolic process, the shadow by many factors such as carbon source, nitrogen sources It rings, continuous feeding zymotechnique is mainly taken in actual industrial production, fermentation level is relatively stable, but due to fermentation medium The limitation of middle material and the control of feed supplement sugar cost, so that horizontal being promoted of riboflavin fermentation has certain limitation.

Summary of the invention

The purpose of the present invention is to provide a kind of alimentation composition for promoting riboflavin strain growth to utilize, the composition energy Effectively improve the fermentation level of riboflavin.

The technical scheme adopted by the invention to solve the technical problem is that:

A kind of composition promoting riboflavin fermentation, is mainly made of the raw material of following mass fraction: composite precursor 1-3%, compound ammonia Base acid 2-10% and tap water 50-70%；The composite precursor is mainly by sodium gluconate, sorbic acid and malonic acid in mass ratio 1: 2:2 composition；The compound amino acid is mainly made of tyrosine and phenylalanine 2.5:3.5 in mass ratio.

Preferably, the component content of the sodium gluconate is 98-99%, the active constituent content of the sorbic acid is >= 99%, the active constituent content of the malonic acid is 98-99%, the active constituent content of the tyrosine and phenylalanine >= 95%。

A kind of application of the composition in riboflavin fermentation promoting riboflavin fermentation, the riboflavin fermentation includes following step It is rapid:

(1) preparation of level-one, secondary seed medium: the prescription quality of the seed culture medium is than composition are as follows: glucose 20- 25g/L, 20 g/L of corn pulp, 12 g/L of yeast extract, epsom salt 0.5 g/L, ammonium chloride 3g/L, potassium dihydrogen phosphate 1.0g/ L, 2.0 g/L of dipotassium hydrogen phosphate adjust pH to 7.2, pH7.0-7.2 before disappearing, in 121 DEG C of sterilizing 20-25min.

(2) preparation of fermentation medium: the prescription quality of the fermentation medium is than composition are as follows: glucose 40g/L, it is beautiful Rice & peanut milk 12g/L, yeast extract 5g/L, beancake powder 8g/L, epsom salt 1g/L, ammonium chloride 1.8g/L, potassium dihydrogen phosphate 0.2g/L, Dipotassium hydrogen phosphate 0.5g/L, precipitated calcium carbonate 0.5g/L adjust PH to 7.2, pH 7.0-7.2 before disappearing, in 121 DEG C of sterilizing 20- 25min to get.

(3) first order seed culture: by seed culture medium sterilizing, the cooling of step (1), being inoculated with and produce strain into riboflavin, Condition of culture is that tank presses 0.03-0.05Mpa, vvm0.6-0.8, and tank temperature is 36-36.5 DEG C, revolving speed 150-180rpm, cultivation cycle 20-22h；

(4) secondary seed culture: by seed culture medium sterilizing, the cooling of step (1).It is net to first order seed OD value in step (3) When increasing 10-14, by cultivating in inoculum concentration 6-10% access secondary seed tank, condition of culture is that tank presses 0.03-0.05Mpa, Vvm0.6-1.0, tank temperature are 36-36.5 DEG C, revolving speed 160-200rpm, cultivation cycle 18-20h；

(5) the rush riboflavin fermentation composition fermented and cultured: is added to step (2) according to the additive amount of 2.0-3.0wt% In fermentation medium, PH to 7.0-7.2 is adjusted to have a net increase of 6- to secondary seed OD value in step (4) in 121 DEG C of sterilizing 30min before disappearing It is that 6-10% is accessed in fermentor by inoculum concentration when 8, initial culture conditions 0.03-0.035Mpa, revolving speed 60-70rpm, Vvm0.6-0.65,36-36.5 DEG C of tank temperature, Initial sugar concentration 25-30g/L, phosphorus concentration 0.3-0.4g/L, ammonia nitrogen concentration 160- 200mg/L surveys glucose content, ammonia-nitrogen content, OD, pH and microscopy every 4 hour samplings after inoculation and observes bacterial type, fermenting The fermentation composition is added in the feed supplement liquid of process, and is controlled glucose content in 9-11g/L by adjusting flow feeding amount； By adding liquefied ammonia stability contorting PH；When the early period of OD value≤20, pH controls 6.6-6.8, and 36-36.5 DEG C of temperature；Mid-term OD value is When 20-120, pH control 6.9-7.0,37-38 DEG C of temperature；Later period to fermentation ends control pH7.1-7.2, temperature 38.5-39.5 ℃；To occur gemma in fermentor, and when the every 4 hours increasing degree≤1000mg/L of unit, stop fermentation.

The beneficial effects of the present invention are: compared with prior art, a kind of composition of rush riboflavin fermentation of the invention and Its application has the advantages that (1) present invention promotees the various micronutrient factor additive of fermentation composition, meets hair The appropriate combination of ferment needed nutrient is supplied, and (2) present invention is applied to riboflavin fermentation mistake will promote riboflavin fermentation composition When in journey, rush fermentation composition is constantly added during the fermentation, ensure that the supply of various nutritional ingredients in fermentation process, is protected Ferment effect has been demonstrate,proved, the fermentation yield of riboflavin is effectively increased.

Specific embodiment

A kind of composition for promoting riboflavin fermentation of embodiment 1

A kind of composition promoting riboflavin fermentation, is mainly made of the raw material of following mass fraction: composite precursor 2kg, compound ammonia Base acid 3.5kg and tap water 68.9kg；The composite precursor is mainly by sodium gluconate, sorbic acid and malonic acid in mass ratio 1: 2:2 composition；The compound amino acid is mainly made of tyrosine and phenylalanine 2.5:3.5 in mass ratio；The gluconic acid The component content of sodium is 98.3%, and the active constituent content of the sorbic acid is 99.5%, the active constituent content of the malonic acid It is 98.9%, the active constituent content of the tyrosine and phenylalanine is >=95%.

A kind of composition for promoting riboflavin fermentation of embodiment 2

A kind of composition promoting riboflavin fermentation, is mainly made of the raw material of following mass fraction: composite precursor 2.3kg, compound Amino acid 8.2kg and tap water 56kg；The composite precursor is mainly by sodium gluconate, sorbic acid and malonic acid in mass ratio 1: 2:2 composition；The compound amino acid is mainly made of tyrosine and phenylalanine 2.5:3.5 in mass ratio；The gluconic acid The component content of sodium is 98.9%, and the active constituent content of the sorbic acid is 99.2%, the active constituent content of the malonic acid It is 99.5%, the active constituent content of the tyrosine and phenylalanine is >=95%.

A kind of application of the composition for promoting riboflavin fermentation of embodiment 3 in riboflavin fermentation

A kind of application method of the composition in riboflavin fermentation promoting riboflavin fermentation, the riboflavin fermentation includes following step It is rapid:

(1) preparation of seed culture medium: the prescription quality of the seed culture medium is than composition are as follows: glucose 20g/L, corn pulp 20 G/L, 12 g/L of yeast extract, epsom salt 0.5 g/L, ammonium chloride 3g/L, potassium dihydrogen phosphate 1.0g/L, dipotassium hydrogen phosphate 2.0 G/L, 7.2, pH to 7.12 is adjusted before disappearing, in 121 DEG C of sterilizing 25min.

(2) preparation of fermentation medium: the prescription quality of the fermentation medium is than composition are as follows: the step fermentation medium Prescription quality than composition are as follows: glucose 40g/L, corn pulp 12g/L, yeast extract 5g/L, beancake powder 8g/L, epsom salt 1g/L, ammonium chloride 1.8g/L, potassium dihydrogen phosphate 0.2g/L, dipotassium hydrogen phosphate 0.5g/L, precipitated calcium carbonate 0.5g/L, by the rush Riboflavin fermentation composition is added in the fermentation medium of step (2) according to the additive amount of 2.0wt%, and PH to 7.17 is adjusted before disappearing, In 121 DEG C of sterilizing 30min.

(3) first order seed culture: by seed culture medium sterilizing, the cooling of step (1), being inoculated with and produce strain into riboflavin, Condition of culture is that tank presses 0.04Mpa, vvm0.7, and tank temperature is 36.5 DEG C, revolving speed 170rpm；

(4) secondary seed culture: by seed culture medium sterilizing, the cooling of step (1).It is net to first order seed OD value in step (3) It when increasing 13, accesses in secondary seed tank and cultivates by inoculum concentration 7%, condition of culture is that tank presses 0.04Mpa, vvm0.8, and tank temperature is 36.5 DEG C, revolving speed 160rpm；

(5) fermented and cultured: by seed culture medium sterilizing, the cooling of step (2).When secondary seed OD value in step (4) it is net 6 when, It is initial culture conditions 0.03Mpa, revolving speed 70rpm, vvm0.6 in 8% access fermentor by inoculum concentration, 36.5 DEG C of tank temperature, Initial sugar concentration 30g/L, phosphorus concentration 0.34g/L, ammonia nitrogen concentration 185.2mg/L survey glucose every 4 hour samplings after inoculation Content, ammonia-nitrogen content, OD, pH and microscopy observe bacterial type, and the fementative composition in embodiment 1 is added in the feed supplement liquid of fermentation process Object, and controlled glucose content in 11g/L by adjusting flow feeding amount；By adding liquefied ammonia stability contorting PH；Early period OD When value≤20, pH control 6.6,36.5 DEG C of temperature；When mid-term OD value is 20-120, pH control 6.9,38 DEG C of temperature；Later period is extremely sent out Ferment finishing control pH7.1,39.5 DEG C of temperature；To occur gemma, and the every 4 hours increasing degree≤1000mg/L of unit in fermentor When, stop fermentation.

The present embodiment fermentation period is 52h, and fermentation ends, the fermentation titer of riboflavin is 18936mg/L.

A kind of application of the composition for promoting riboflavin fermentation of embodiment 4 in riboflavin fermentation

A kind of application method of the composition in riboflavin fermentation promoting riboflavin fermentation, the riboflavin fermentation includes following step It is rapid:

(1) preparation of seed culture medium: the prescription quality of the seed culture medium is than composition are as follows: glucose 20-25g/L, corn Starch 20 g/L, 12 g/L of yeast extract, epsom salt 0.5 g/L, ammonium chloride 3g/L, potassium dihydrogen phosphate 1.0g/L, phosphoric acid hydrogen two 2.0 g/L of potassium, pH tune pH to 7.09 before disappearing, in 121 DEG C of sterilizing 20-25min.

(2) preparation of fermentation medium: the prescription quality of the step fermentation medium is than composition are as follows: the step fermentation training The prescription quality of base is supported than composition are as follows: glucose 40g/L, corn pulp 12g/L, yeast extract 5g/L, beancake powder 8g/L, seven water sulfuric acid Magnesium 1g/L, ammonium chloride 1.8g/L, potassium dihydrogen phosphate 0.2g/L, dipotassium hydrogen phosphate 0.5g/L, precipitated calcium carbonate 0.5g/L will be described Promote riboflavin fermentation composition to be added in the fermentation medium according to the additive amount of 2.8wt%, PH to 7.12 is adjusted before disappearing, in 121 DEG C sterilizing 20-25min to get.

(3) first order seed culture: by seed culture medium sterilizing, the cooling of step (1), being inoculated with and produce strain into riboflavin, Condition of culture is that tank presses 0.05Mpa, vvm0.6, and tank temperature is 36 DEG C, revolving speed 160rpm；

(4) secondary seed culture: by seed culture medium sterilizing, the cooling of step (1).It is net to first order seed OD value in step (3) It when increasing 10, accesses in secondary seed tank and cultivates by inoculum concentration 10%, condition of culture is that tank presses 0.05Mpa, vvm0.8, and tank temperature is 36 DEG C, revolving speed 170rpm；

(5) fermented and cultured: by seed culture medium sterilizing, the cooling of step (2).7 are had a net increase of to secondary seed OD value in step (4) When, it is initial culture conditions 0.03Mpa, revolving speed 70rpm in 10% access fermentor, vvm0.65, tank temperature 36 by inoculum concentration DEG C, Initial sugar concentration 26g/L, phosphorus concentration 0.35g/L, ammonia nitrogen concentration 176.2mg/L survey Portugal every 4 hour samplings after inoculation Grape sugared content, ammonia-nitrogen content, OD, pH and microscopy observe bacterial type, and the fermentation in embodiment 2 is added in the feed supplement liquid of fermentation process Composition, and controlled glucose content in 9g/L by adjusting flow feeding amount；By adding liquefied ammonia stability contorting PH；Early period When OD value≤20, pH control 6.8,36 DEG C of temperature；When mid-term OD value is 20-120, pH control 7.0,37 DEG C of temperature；Later period is extremely sent out Ferment finishing control pH7.2,38.5 DEG C of temperature；To occur gemma, and the every 4 hours increasing degree≤1000mg/L of unit in fermentor When, stop fermentation.

The present embodiment fermentation period is 56h, and fermentation ends, the fermentation titer of riboflavin is 19254mg/L.

Comparative example 1 is compared with embodiment 3, and the difference with embodiment 3 is, in fermented and cultured step, in fermentation process Feed supplement liquid in using the water of same volume replace the fermentation composition in embodiment 1, what is be added in comparative example 1 is in embodiment 3 The water of the fermentation composition of the same volume of addition, in this way formed embodiment 3 comparison, other the step of all and in embodiment 3 Identical, reference examples of the comparative example 1 as embodiment 3, the fermentation period of comparative example 1 is also for 52h, fermentation ends, through detecting core The fermentation titer of flavine is 17602mg/L.

Comparative example 2 is compared with embodiment 4, and the difference with embodiment 4 is, in fermented and cultured step, in fermentation process Feed supplement liquid in using the water of same volume replace the fermentation composition in embodiment 2, what is be added in comparative example 2 is in embodiment 4 The water of the fermentation composition of the same volume of addition, in this way formed embodiment 2 comparison, other the step of all and in embodiment 4 Identical, reference examples of the comparative example 2 as embodiment 4, the fermentation period of comparative example 2 is also for 56h, fermentation ends, through detecting core The fermentation titer of flavine is 17716mg/L.

By the comparison of embodiment 3 and comparative example 1 and embodiment 4 and comparative example 2, rush riboflavin fermentation of the invention Composition can significantly improve the fermentation yield to riboflavin in riboflavin fermentation, ensure that ferment effect.

Above-mentioned specific embodiment is only specific case of the invention, and scope of patent protection of the invention includes but is not limited to The product form and style of above-mentioned specific embodiment, any claims of the present invention and any technical field of meeting The appropriate changes or modifications that those of ordinary skill does it, all shall fall within the protection scope of the present invention.

Claims (3)

1. a kind of composition for promoting riboflavin fermentation, it is characterised in that: be mainly made of the raw material of following mass fraction: before compound Body 1-3%, compound amino acid 2-10% and tap water 50-70%；The composite precursor is mainly by sodium gluconate, sorbic acid and third Diacid 1:2:2 in mass ratio composition；The compound amino acid is mainly by tyrosine and phenylalanine 2.5:3.5 group in mass ratio At.

2. a kind of composition for promoting riboflavin fermentation according to claim 1, it is characterised in that: the sodium gluconate Component content is 98-99%, and the active constituent content of the sorbic acid is >=99%, and the active constituent content of the malonic acid is The active constituent content of 98-99%, the tyrosine and phenylalanine is >=95%.

3. a kind of application of composition for promoting riboflavin fermentation in riboflavin fermentation, it is characterised in that: the riboflavin fermentation The following steps are included:

(1) preparation of level-one, secondary seed medium: the prescription quality of the seed culture medium is than composition are as follows: glucose 20- 25g/L, 20 g/L of corn pulp, 12 g/L of yeast extract, epsom salt 0.5 g/L, ammonium chloride 3g/L, potassium dihydrogen phosphate 1.0g/ L, 2.0 g/L of dipotassium hydrogen phosphate adjust pH to 7.2, pH7.0-7.2 before disappearing, in 121 DEG C of sterilizing 20-25min；

(2) preparation of fermentation medium: the prescription quality of the fermentation medium is than composition are as follows: glucose 40g/L, corn pulp 12g/L, yeast extract 5g/L, beancake powder 8g/L, epsom salt 1g/L, ammonium chloride 1.8g/L, potassium dihydrogen phosphate 0.2g/L, phosphoric acid Hydrogen dipotassium 0.5g/L, precipitated calcium carbonate 0.5g/L adjust PH to 7.2, pH 7.0-7.2 before disappearing, in 121 DEG C of sterilizing 20-25min, i.e., ?；

(3) it first order seed culture: by seed culture medium sterilizing, the cooling of step (1), is inoculated with and produces strain, culture into riboflavin Condition is that tank presses 0.03-0.05Mpa, vvm0.6-0.8, and tank temperature is 36-36.5 DEG C, revolving speed 150-180rpm, cultivation cycle 20- 22h；

(4) secondary seed culture: net to first order seed OD value in step (3) by seed culture medium sterilizing, the cooling of step (1) When increasing 10-14, by cultivating in inoculum concentration 6-10% access secondary seed tank, condition of culture is that tank presses 0.03-0.05Mpa, Vvm0.6-1.0, tank temperature are 36-36.5 DEG C, revolving speed 160-200rpm, cultivation cycle 18-20h；

(5) the rush riboflavin fermentation composition fermented and cultured: is added to step (2) according to the additive amount of 2.0-3.0wt% In fermentation medium, PH to 7.0-7.2 is adjusted to have a net increase of 6- to secondary seed OD value in step (4) in 121 DEG C of sterilizing 30min before disappearing It is that 6-10% is accessed in fermentor by inoculum concentration when 8, initial culture conditions 0.03-0.035Mpa, revolving speed 60-70rpm, Vvm0.6-0.65,36-36.5 DEG C of tank temperature, Initial sugar concentration 25-30g/L, phosphorus concentration 0.3-0.4g/L, ammonia nitrogen concentration 160- 200mg/L surveys glucose content, ammonia-nitrogen content, OD, pH and microscopy every 4 hour samplings after inoculation and observes bacterial type, fermenting The fermentation composition is added in the feed supplement liquid of process, and is controlled glucose content in 9-11g/L by adjusting flow feeding amount； By adding liquefied ammonia stability contorting PH；When the early period of OD value≤20, pH controls 6.6-6.8, and 36-36.5 DEG C of temperature；Mid-term OD value is When 20-120, pH control 6.9-7.0,37-38 DEG C of temperature；Later period to fermentation ends control pH7.1-7.2, temperature 38.5-39.5 ℃；To occur gemma in fermentor, and when the every 4 hours increasing degree≤1000mg/L of unit, stop fermentation.