CN105316370A - Method for producing gamma-polyglutamic acid by using solid-state fermentation of Bacillus natto - Google Patents
Method for producing gamma-polyglutamic acid by using solid-state fermentation of Bacillus natto Download PDFInfo
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- CN105316370A CN105316370A CN201410361850.4A CN201410361850A CN105316370A CN 105316370 A CN105316370 A CN 105316370A CN 201410361850 A CN201410361850 A CN 201410361850A CN 105316370 A CN105316370 A CN 105316370A
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- polyglutamic acid
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Abstract
The invention relates to a method for producing gamma-polyglutamic acid by using solid-state fermentation of bacillus natto. The method of the present invention is not reported in the prior art. The method comprises: preparing a soybean meal solid-state fermentation base material, mixing into a nutrition supplement, preparing a solid state fermentation culture medium, inoculating Bacillus natto, carrying out constant temperature moisturizing for 4-10 days, carrying out stirring extraction of gamma-polyglutamic acid with water, filtering, carrying out rotary evaporation concentration on the filtrate, adding ethanol, precipitating the gamma-polyglutamic acid, and carrying out centrifugation recovery to obtain the gamma-polyglutamic acid. According to the present invention, the method can be used in the fields of agriculture, food, medicine, chemical industry, and the like.
Description
technical field:the present invention relates to the new and high technology in a kind of fermentation engineering of technical field of bioengineering, particularly a kind of method utilizing bacillus natto solid state fermentation to produce gamma-polyglutamic acid-.
background technology: the production method of the gamma-polyglutamic acid-of research mostly at present is liquid fermenting, not yet finds to adopt bean pulp solid-state fermentation in the present invention to produce gamma-polyglutamic acid-method.
summary of the invention:the object of this invention is to provide one utilizes dregs of beans as solid state fermentation base-material, is mixed into a certain proportion of nutritive medium, and inoculation bacillus natto, produces the method for gamma-polyglutamic acid-by solid state fermentation.
Above-mentioned object is realized by following technical scheme:
Bacillus natto solid state fermentation is utilized to produce the method for gamma-polyglutamic acid-, its composition comprises: preparation bean pulp solid-state fermentation base-material, be mixed into nutritious supplementary, make solid-state fermentation culture medium, after inoculation bacillus natto, leave standstill solid state fermentation, add water to stir and extract gamma-polyglutamic acid-, screen filtration, add ethanol after filtrate concentrated by rotary evaporation, precipitation gamma-polyglutamic acid-, centrifugal recovery obtains gamma-polyglutamic acid-.
Described preparation bean pulp solid-state fermentation base-material comprises: after dregs of beans, crushed stalk, according to dregs of beans: the part by weight of stalk=8-10:0-2 mixes.
Described nutritious supplementary comprises: sucrose, Pidolidone, KH
2pO
4, anhydrous CaCl
2, MgSO
4forming with water, their weight thousand points of concentration respectively are 20-30 ‰, 30-40 ‰, 0.5-1 ‰, 0.5-1 ‰, 0.5-1 ‰, and all the other are water, pH7.0-7.5.
Described solid-state fermentation culture medium is made up of solid state fermentation base-material and nutritious supplementary, and both part by weight scopes are 1:1.0-1.5.
Described standing solid state fermentation is: bacillus natto is by 5-10%(V/W) be inoculated in solid-state fermentation culture medium, abundant stirring, bacterial classification is made to be uniformly distributed in media surface, postvaccinal incubator keeps the constant temperature of 30-37oC, incubator humidity strictly remains on more than 50%, fermentation 4-10 days.
Described extraction gamma-polyglutamic acid-is: the water adding solid state fermentation thing 5-10 times of volume stirs and extracts 1-2 hour, after 100-200 eye mesh screen filters, collect filtrate, liquid concentrated by rotary evaporation to concentration 20-60g/L, then adds the ethanol of 1-4 times of volume, leaves standstill 6-48 hour, centrifugal collecting precipitation, then by the washing with alcohol of 1-4 times of volume, vacuum lyophilization, obtains the gamma-polyglutamic acid-refined.
This technical scheme has following beneficial effect:
1, take dregs of beans as fermentation base-material, add a certain proportion of nutritive medium, solid state fermentation γ-polyglutamic acid, provide a kind of efficient a large amount of method of producing gamma-polyglutamic acid-, facilitate comprehensive exploitation and the higher value application of dregs of beans simultaneously.
2, water-soluble as one, the biodegradable green macromolecular material of gamma-polyglutamic acid-, molecular range is 10kDa-1,000kDa.Application is had in various fields, can be used in food antifreezing agent, thickening material, except astringent etc., moisture saver mask etc. is can be used in cosmetic industry, in pharmaceutical industries, can be used as pharmaceutical carrier, tackiness agent etc., in agricultural, have good water-retentivity to edaphophyte, also can be used as fertilizer synergist etc., industrially can be used for high-molecular biologic flocculation agent, heavy metal absorbent etc.This shows, gamma-polyglutamic acid-all has a good application prospect in a lot of fields.
accompanying drawing illustrates:accompanying drawing 1 is the process schematic representation of present method.
the specific embodiment of the present invention:
Example 1:
Bacillus natto (
bacillussubtilisnatto) activation and cultivation.Primary inclined plane adopts LB inclined-plane, cultivates 24h, be inoculated in secondary LB liquid nutrient medium in 37 DEG C, and 37 DEG C of concussion cultivation 18-24 are little of growth logarithmic phase.For subsequent use as solid fermentation bacterial classification.
Using dregs of beans 1000g as solid fermentation base-material; Another by sucrose 20g, Pidolidone 30g, KH
2pO
40.5g, anhydrous CaCl
20.5g, MgSO
40.5g and water are mixed and made into 1000g nutritious supplementary, are 7.0 with HCl or NaOH aqueous solution adjust pH; Above-mentioned solid fermentation base-material is mixed into nutritious supplementary, and both mix according to the part by weight of 1:1, make solid fermentation substratum, load in polyethylene plastic bag, tighten sack, in 105oC boiling 20 minutes, be cooled to 50 DEG C, the ratio according to 10% is seeded in above-mentioned solid fermentation substratum, after stirring, be laid in pallet by the thickness of 5cm, use polypropylene film capping, in 37oC solid fermentation 4 days, ambient moisture was more than 50%, every day turns, and culture is mixed.
After continuously fermenting 4 days, occurring in fermention medium can the glutinous material of wire drawing, and stir extraction 1 hour with the distilled water of 10 times of volumes, 200 eye mesh screens filter, collection filtrate.Filtrate is revolved and is steamed to gamma-polyglutamic acid-concentration when 40-60g/L, stops revolving steaming.In concentrated solution, add the ethanol of 2 times of volumes, fully stir, 4 DEG C, spend the night, 4800 revs/min centrifugal 10 minutes, the raw product being precipitated as gamma-polyglutamic acid-obtained.
By the washing with alcohol of raw product 2 times of volumes of gamma-polyglutamic acid-obtained, lyophilize, obtains gamma-polyglutamic acid-highly finished product, and the gamma-polyglutamic acid-yield of per kilogram raw material dry-matter is 200g.
Example 2
The activation of bacillus natto and cultivation.Primary inclined plane adopts LB inclined-plane, cultivates 24h, be inoculated in secondary LB liquid nutrient medium in 37 DEG C, and 37 DEG C of concussion cultivation 18-24 are little of growth logarithmic phase.For subsequent use as solid state fermentation bacterial classification.
Dregs of beans 900g is mixed with stalk 100g, as solid state fermentation base-material; Another by sucrose 45g, Pidolidone 60g, KH
2pO
41.5g, anhydrous CaCl
21.5g, MgSO
41.5g and water are mixed and made into 1500g nutritious supplementary, are 7.5 with HCl or NaOH aqueous solution adjust pH; Above-mentioned solid state fermentation base-material is mixed into nutritious supplementary, and both mix according to the part by weight of 1:1.5, make solid-state fermentation culture medium, load in polyethylene plastic bag, tighten sack, in 105oC boiling 20 minutes, be cooled to 50 DEG C, the ratio according to 5% is seeded in above-mentioned solid-state fermentation culture medium, after stirring, be laid in pallet by the thickness of 5cm, use polypropylene film capping, in 37oC solid state fermentation 7 days, ambient moisture was more than 50%, every day turns, and culture is mixed.
After continuously fermenting 7 days, occurring in fermention medium can the glutinous material of wire drawing, and stir extraction 1 hour with the distilled water of 10 times of volumes, 200 eye mesh screens filter, collection filtrate.Filtrate is revolved and is steamed to gamma-polyglutamic acid-concentration when 40-60g/L, stops revolving steaming.In concentrated solution, add the ethanol of 2 times of volumes, fully stir, 4 DEG C, spend the night, 4800 revs/min centrifugal 10 minutes, the raw product being precipitated as gamma-polyglutamic acid-obtained.
By the raw product 2 times of washing with alcohol of gamma-polyglutamic acid-obtained, lyophilize, obtains gamma-polyglutamic acid-highly finished product, and the gamma-polyglutamic acid-yield of per kilogram raw material dry-matter is 184g.
Claims (6)
1. the method utilizing bacillus natto solid state fermentation to produce gamma-polyglutamic acid-, its composition comprises: preparation bean pulp solid-state fermentation base-material, be mixed into nutritious supplementary, make solid-state fermentation culture medium, after inoculation bacillus natto, leave standstill solid state fermentation, add water to stir and extract gamma-polyglutamic acid-, screen filtration, add ethanol after filtrate concentrated by rotary evaporation, precipitation gamma-polyglutamic acid-, centrifugal recovery obtains gamma-polyglutamic acid-.
2. the method utilizing bacillus natto solid state fermentation to produce gamma-polyglutamic acid-according to claim 1, it is characterized in that described preparation bean pulp solid-state fermentation base-material comprises: after dregs of beans, crushed stalk, according to dregs of beans: the part by weight of stalk=8-10:0-2 mixes.
3. the method utilizing bacillus natto solid state fermentation to produce gamma-polyglutamic acid-according to claim 1, is characterized in that described nutritious supplementary comprises: sucrose, Pidolidone, KH
2pO
4, anhydrous CaCl
2, MgSO
4forming with water, their weight thousand points of concentration respectively are 20-30 ‰, 30-40 ‰, 0.5-1 ‰, 0.5-1 ‰, 0.5-1 ‰, and all the other are water, pH7.0-7.5.
4. the method utilizing bacillus natto solid state fermentation to produce gamma-polyglutamic acid-according to claim 1, it is characterized in that described solid-state fermentation culture medium is made up of solid state fermentation base-material and nutritious supplementary, both part by weight scopes are 1:1.0-1.5.
5. the method utilizing bacillus natto solid state fermentation to produce gamma-polyglutamic acid-according to claim 1, it is characterized in that described standing solid state fermentation is: bacillus natto is by 5-10%(V/W) be inoculated in solid-state fermentation culture medium, abundant stirring, bacterial classification is made to be uniformly distributed in media surface, postvaccinal incubator keeps the constant temperature of 30-37oC, incubator humidity strictly remains on more than 50%, fermentation 4-10 days.
6. the method utilizing bacillus natto solid state fermentation to produce gamma-polyglutamic acid-according to claim 1, it is characterized in that described extraction gamma-polyglutamic acid-is: the water adding solid fermentation thing 5-10 times of volume stirs 1-2 hour, extract bacillus natto secretion gamma-polyglutamic acid-in the fermentation medium, after 100-200 eye mesh screen filters, collect filtrate, filtrate concentrated by rotary evaporation is to concentration 20-60g/L, add the ethanol of 1-4 times of volume again, leave standstill 6-48 hour, centrifugal collecting precipitation, then by the washing with alcohol of 1-4 times of volume, vacuum lyophilization, obtain the gamma-polyglutamic acid-refined.
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Cited By (6)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN106834190A (en) * | 2017-03-11 | 2017-06-13 | 鲁东大学 | Culture medium, bacterial strain and the method for polyglutamic acid are produced with pea protein wastewater fermentation |
| CN107198219A (en) * | 2017-05-02 | 2017-09-26 | 上海纳德生物科技有限公司 | A kind of preparation method of the food additives of the polyglutamic acid containing γ |
| CN108048499A (en) * | 2018-02-09 | 2018-05-18 | 烟台市佳益有机肥料有限公司 | A kind of method of solid fermentation production gamma-polyglutamic acid |
| CN108484266A (en) * | 2018-04-21 | 2018-09-04 | 重庆尚奕辉生物科技有限公司 | A kind of production method of liquid compound fertilizer and its use and methods for using them |
| CN108793424A (en) * | 2018-06-15 | 2018-11-13 | 江苏远山生物技术有限公司 | One seed shrimp crab breeding water body antidote and preparation method thereof |
| CN110951798A (en) * | 2020-01-07 | 2020-04-03 | 广东丸美生物技术股份有限公司 | Biological fermentation method of gamma-polyglutamic acid and application thereof |
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Cited By (6)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN106834190A (en) * | 2017-03-11 | 2017-06-13 | 鲁东大学 | Culture medium, bacterial strain and the method for polyglutamic acid are produced with pea protein wastewater fermentation |
| CN107198219A (en) * | 2017-05-02 | 2017-09-26 | 上海纳德生物科技有限公司 | A kind of preparation method of the food additives of the polyglutamic acid containing γ |
| CN108048499A (en) * | 2018-02-09 | 2018-05-18 | 烟台市佳益有机肥料有限公司 | A kind of method of solid fermentation production gamma-polyglutamic acid |
| CN108484266A (en) * | 2018-04-21 | 2018-09-04 | 重庆尚奕辉生物科技有限公司 | A kind of production method of liquid compound fertilizer and its use and methods for using them |
| CN108793424A (en) * | 2018-06-15 | 2018-11-13 | 江苏远山生物技术有限公司 | One seed shrimp crab breeding water body antidote and preparation method thereof |
| CN110951798A (en) * | 2020-01-07 | 2020-04-03 | 广东丸美生物技术股份有限公司 | Biological fermentation method of gamma-polyglutamic acid and application thereof |
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Application publication date: 20160210 |