CN105505903A - Method for preparing induction sugar and inducing the synthesis of cellulose through yeast cells - Google Patents
Method for preparing induction sugar and inducing the synthesis of cellulose through yeast cells Download PDFInfo
- Publication number
- CN105505903A CN105505903A CN201610003974.4A CN201610003974A CN105505903A CN 105505903 A CN105505903 A CN 105505903A CN 201610003974 A CN201610003974 A CN 201610003974A CN 105505903 A CN105505903 A CN 105505903A
- Authority
- CN
- China
- Prior art keywords
- sugar
- induction
- induction sugar
- synthesis
- zymosan
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Granted
Links
Classifications
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N9/00—Enzymes; Proenzymes; Compositions thereof; Processes for preparing, activating, inhibiting, separating or purifying enzymes
- C12N9/14—Hydrolases (3)
- C12N9/24—Hydrolases (3) acting on glycosyl compounds (3.2)
- C12N9/2402—Hydrolases (3) acting on glycosyl compounds (3.2) hydrolysing O- and S- glycosyl compounds (3.2.1)
- C12N9/2405—Glucanases
- C12N9/2434—Glucanases acting on beta-1,4-glucosidic bonds
- C12N9/2437—Cellulases (3.2.1.4; 3.2.1.74; 3.2.1.91; 3.2.1.150)
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12Y—ENZYMES
- C12Y302/00—Hydrolases acting on glycosyl compounds, i.e. glycosylases (3.2)
- C12Y302/01—Glycosidases, i.e. enzymes hydrolysing O- and S-glycosyl compounds (3.2.1)
- C12Y302/01004—Cellulase (3.2.1.4), i.e. endo-1,4-beta-glucanase
Landscapes
- Chemical & Material Sciences (AREA)
- Life Sciences & Earth Sciences (AREA)
- Organic Chemistry (AREA)
- Health & Medical Sciences (AREA)
- Genetics & Genomics (AREA)
- Engineering & Computer Science (AREA)
- Bioinformatics & Cheminformatics (AREA)
- Zoology (AREA)
- Wood Science & Technology (AREA)
- General Health & Medical Sciences (AREA)
- Biochemistry (AREA)
- General Engineering & Computer Science (AREA)
- Microbiology (AREA)
- Biotechnology (AREA)
- Biomedical Technology (AREA)
- Molecular Biology (AREA)
- Medicinal Chemistry (AREA)
- Preparation Of Compounds By Using Micro-Organisms (AREA)
- Enzymes And Modification Thereof (AREA)
- Micro-Organisms Or Cultivation Processes Thereof (AREA)
Abstract
The invention discloses a method for preparing induction sugar and inducing the synthesis of cellulose through yeast cells. The method includes the following steps of firstly, preparing induction sugar through zymosan, wherein zymosan is hydrolyzed through acid water with concentration of 0.2 mol/L to 3 mol/L at a temperature of 90-130 DEG C for 2-4 hours so that induction sugar can be obtained; secondly, inoculating trichoderma strains to a fermentation culture medium with induction sugar obtained in the first step to be cultured so that the synthesis of cellulose can be achieved. The method of preparing induction sugar through zymosan and the process of adding induction sugar in the synthesis of cellulose are simple, low in cost, abundant in raw material source, capable of achieving multiple purposes through one action and suitable for industrial promotion.
Description
Technical field
The present invention relates to a kind of yeast cell that utilizes and prepare induction sugar and the method for fibrin enzymic synthesis.
Background technology
Ligno-cellulosic materials be abundant on the earth can in production-goods source, it can be broken down into the monose such as glucose, wood sugar and be utilized bio-based chemical such as producing ethanol further.At present, although the method that the agricultural wastes such as stalk produce ethanol through cellulase degradation extensively admitted, the restriction that stover is several key factor in bio-based chemical process is subject to.Cellulase hydrolysis efficiency is low, production cost height is one of restraining factors.
The production method of cellulase mainly adopts ligno-cellulosic materials to do substrate fermentable, and the Nomenclature Composition and Structure of Complexes of ligno-cellulosic materials affects its hydrolysis efficiency, thus affects the production of cellulase.The lignocellulose substrate of the cellulase induction of report has been had to have corn cob, maize straw, straw, wheat bran etc. in research.They are all containing higher xylogen, and compact structure, be difficult to be hydrolyzed utilization.Also have researchist to prepare the inductors such as sophorose, add at fermention medium and carry out the synthesis of cellulase induction, but due to the complicated process of preparation of sophorose, cost is higher, produces be difficult to apply at cellulose fermentation.Therefore screen a kind of abundant raw material source, cheap, preparation method simply induces sugar to have very great help for the fermentative production tool of cellulase.
Yeast cells wall is made up of dextran, mannosans etc.Treated yeast cells wall is hydrolyzed to zymosan, and zymosan is hydrolysis also generating portion transglycosylation further under certain condition, and polymerizable is multiple disaccharides.The Induced synthesis of these two sugar mutual-cellulose enzymes has great role, applies its fermentative production inductor making cellulase and has with low cost, is easy to control, and is easy to the advantages such as industrial applications.
Summary of the invention
An object of the present invention is to solve at least the problems referred to above and/or defect, and the advantage will illustrated at least is below provided.
A further object of the invention is to provide a kind of method utilizing the enzymic synthesis of zymosan fibrin, method of the present invention is by being prepared as induction sugar by zymosan, glucose wherein also can be utilized as carbon source simultaneously, solving in cellulase synthesis induces sugared raw material sources few, expensive, the problems such as complicated process of preparation, meanwhile, the output of cellulase is greatly improved.
For this reason, technical scheme provided by the invention is:
Utilize yeast cell to prepare induction sugar and a method for fibrin enzymic synthesis, comprise the following steps:
Step one, utilize zymosan prepare induction sugar: at temperature 90-130 DEG C, utilize the acid hydrolysis zymosan 2 ~ 4h of 0.2mol/L-3mol/L, with obtain induction sugar;
Step 2, Trichoderma is inoculated in be added with the induction sugar obtained in step one fermention medium in cultivate, cellulase synthesis.
Preferably, the described yeast cell that utilizes is prepared in the method for the sugared also fibrin enzymic synthesis of induction, and in described step one, described acid is hydrochloric acid, sulfuric acid, phosphoric acid, acetic acid or nitric acid.
It is further preferred that the described yeast cell that utilizes is prepared in the method for the sugared also fibrin enzymic synthesis of induction, in described step one, described acid is sulfuric acid.
Preferably, the described yeast cell that utilizes is prepared in the method for the sugared also fibrin enzymic synthesis of induction, and in described step one, the concentration of described zymosan is 1%-30% (w/w).
Preferably, the described yeast cell that utilizes is prepared in the method for the sugared also fibrin enzymic synthesis of induction, and in described step 2, the induction sugar in described fermention medium adds in described fermention medium before fermentation culture or in fermentation culture process.
Preferably, the described yeast cell that utilizes is prepared in the method for the sugared also fibrin enzymic synthesis of induction, and described Trichoderma is Trichodermareesei.
Preferably, the described yeast cell that utilizes is prepared in the method for the sugared also fibrin enzymic synthesis of induction, and comprise induction sugar in described fermention medium, in described induction sugar, the concentration of disaccharides is 0.5 ~ 6.5g/L.
Preferably, the described yeast cell that utilizes is prepared in the method for the sugared also fibrin enzymic synthesis of induction, and described zymosan derives from yeast thalline discarded in fermentation industry.
The present invention at least comprises following beneficial effect:
Yeast cell is easy to get, and the making method preparing induction sugared by zymosan is comparatively simple, meet abundant raw material source and cheap, induction sugar in containing in a large number can fibrin synthesis disaccharides, such as cellobiose, gentiobiose, sophorose etc., therefore, the induction sugar that zymosan makes is used for induction fermentation cellulase synthesis, can applies.
The present invention, by adding induction sugar in cellulase fermentations substratum, reduces the cost of fermention medium, solves the problem that cellulase synthesis cost is high.
Method of the present invention is by being prepared as induction sugar by zymosan, glucose wherein also can be utilized simultaneously as carbon source, solve in cellulase synthesis and induce sugared raw material sources few, expensive, the problems such as complicated process of preparation, meanwhile, the output of cellulase is greatly improved.
Present invention process is simple, and cost is low, abundant raw material source, and can achieve many things at one stroke, and is suitable for commercial introduction.
Part is embodied by explanation below by other advantage of the present invention, target and feature, part also will by research and practice of the present invention by those skilled in the art is understood.
Embodiment
Below in conjunction with embodiment, the present invention is described in further detail, can implement according to this with reference to specification sheets word to make those skilled in the art.
Should be appreciated that used hereinly such as " to have ", other element one or more do not allotted in " comprising " and " comprising " term or the existence of its combination or interpolation.
The invention provides a kind of yeast cell that utilizes and prepare induction sugar and the method for fibrin enzymic synthesis, comprise the following steps:
Step one, utilize zymosan prepare induction sugar: at temperature 90-130 DEG C, utilize the acid hydrolysis zymosan 2 ~ 4h of 0.2mol/L-3mol/L, with obtain induction sugar, comprise cellobiose, gentiobiose, sophorose;
Step 2, Trichoderma is inoculated in be added with the induction sugar obtained in step one fermention medium in cultivate, cellulase synthesis.
Yeast cell is easy to get, and the making method preparing induction sugared by zymosan is comparatively simple, meet abundant raw material source and cheap, induction sugar in containing in a large number can fibrin synthesis disaccharides, such as cellobiose, gentiobiose, sophorose etc., therefore, the induction sugar that zymosan makes is used for induction fermentation cellulase synthesis, can applies.
The present invention, by adding induction sugar in cellulase fermentations substratum, reduces the cost of fermention medium, solves the problem that cellulase synthesis cost is high.
In one of them embodiment of the present invention, as preferably, in described step one, described acid is hydrochloric acid, sulfuric acid, phosphoric acid, acetic acid or nitric acid.
In such scheme, as preferably, in described step one, described acid is sulfuric acid.
In one of them embodiment of the present invention, as preferably, in described step one, the concentration of described zymosan is 1%-30% (w/w).
In one of them embodiment of the present invention, as preferably, in described step 2, the induction sugar in described fermention medium adds in described fermention medium before fermentation culture or in fermentation culture process.
In one of them embodiment of the present invention, as preferably, described Trichoderma is Trichodermareesei.
In one of them embodiment of the present invention, as preferably, comprise induction sugar in described fermention medium, in described induction sugar, the concentration of disaccharides is 0.5 ~ 6.5g/L.Further, do not need additionally to add glucose in this fermention medium.Because the glucose containing more amount in induction sugar, therefore also saves glucose, reduce further production cost, achieve many things at one stroke.
In one of them embodiment of the present invention, as preferably, described zymosan derives from yeast thalline discarded in fermentation industry.Raw material is easy to get, wide material sources.
Embodiment one
(1) acquisition of liquid glucose is induced: prepare zymosan according to method disclosed in Chinese Patent Application No. 201110043824.3, afterwards with the obtained induction sugar of following method process zymosan: at 90 DEG C, with the H of 0.5mol/L
2sO
4solution is made into the concentration of substrate hydrolysis 3h of 4%, and the hydrolyzed solution of rear generation contains the glucose of 6g/L, 0.8g/L disaccharides.
(2) bacterial classification obtains: Li's Trichoderma strains is inoculated in PDA inclined-plane, in 28 DEG C of constant incubators, cultivates 6d.Then the bacterial strain after activation on PDA slant medium being prepared into concentration is 2 × 10
7individual/mL embraces sub-suspension, and the inoculum size according to 3% is inoculated in fresh seed culture medium, and wherein seed culture medium consists of: Microcrystalline Cellulose 2%, corn steep liquor 1%, glucose 1%, regulates pH to be 4.5.Be placed in 30 DEG C, on the shaking table that rotating speed is 180 revs/min, concussion is cultivated.
(3) seed culture fluid cultivating 24h is seeded in fresh fermention medium according to 5% inoculum size.Consisting of of fermention medium: Microcrystalline Cellulose 3.3%, corn steep liquor 1.7%, (NH
4)
2sO
40.4%, KH
2pO
40.6%, CaCO
30.25%, MgSO
40.1%, hydrolyzed solution (with glucose meter) adds 0.5%, and disaccharides addition is 0.67g/L, regulates initial 5.0.Contrast is Microcrystalline Cellulose 3.3%, corn steep liquor 1.7%, (NH
4)
2sO
40.4%, KH
2pO
40.6%, CaCO
30.25%, MgSO
40.1%, glucose 0.5%, regulates initial 5.0.Carry out shake flask fermentation cultivation, in 26 DEG C, after 120h is cultivated in concussion on the shaking table that rotating speed is 180 revs/min.The International Standards Method adopting IUPAC (IUPAC) to recommend measures fermentation broth enzyme and lives, and obtaining filter paper enzyme activity is 6.95FPU/mL, and control group filter paper enzyme activity is 5.45FPU/mL.
Embodiment two
(1) acquisition of liquid glucose is induced: prepare zymosan according to method disclosed in Chinese Patent Application No. 201110043824.3, afterwards with the obtained induction sugar of following method process zymosan: at 130 DEG C, with the H of 1mol/L
2sO
4solution is made into the concentration of substrate hydrolysis 3h of 20%, and the hydrolyzed solution of rear generation contains the glucose of 35g/L, 6g/L disaccharides.
(2) bacterial classification obtains: Li's Trichoderma strains is inoculated in PDA inclined-plane, in 28 DEG C of constant incubators, cultivates 6d.Then the bacterial strain after activation on PDA slant medium being prepared into concentration is 2 × 10
7individual/mL spore suspension, the inoculum size according to 3% is inoculated in fresh seed culture medium, and wherein seed culture medium consists of: Microcrystalline Cellulose 2%, corn steep liquor 1%, glucose 1%, regulates pH to be 4.5, be placed in 30 DEG C, on the shaking table that rotating speed is 180 revs/min, concussion is cultivated.
(3) to be seeded in fresh fermention medium according to 5% inoculum size to carry out shake flask fermentation cultivation, consisting of of fermention medium by cultivating the seed culture fluid of 24h: Microcrystalline Cellulose 3.3%, corn steep liquor 1.7%, (NH
4)
2sO
40.4%, KH
2pO
40.6%, CaCO
30.25%, MgSO
40.1%, hydrolyzed solution (with glucose meter) adds 0.5%, and disaccharides addition is 0.85g/L, regulates initial 5.0.Contrast is Microcrystalline Cellulose 3.3%, corn steep liquor 1.7%, (NH
4)
2sO
40.4%, KH
2pO
40.6%, CaCO
30.25%, MgSO
40.1%, glucose 0.5%, regulates initial 5.0.In 26 DEG C, after on the shaking table that rotating speed is 180 revs/min, 120h is cultivated in concussion.The International Standards Method adopting IUPAC (IUPAC) to recommend measures fermented liquid filter paper enzyme activity 7.24FPU/mL, contrasts as 5.45FPU/mL.
Embodiment three
(1) acquisition of liquid glucose is induced: prepare zymosan according to method disclosed in Chinese Patent Application No. 201110043824.3, sugared at 120 DEG C of temperature with the obtained induction of following method process zymosan afterwards, be made into the concentration of substrate hydrolysis 2.5h of 15% with the hydrochloric acid soln of 1mol/L, the hydrolyzed solution of rear generation contains glucose and the 2.4g/L disaccharides of 24g/L.
(2) bacterial classification obtains: Li's Trichoderma strains is inoculated in PDA inclined-plane, in 28 DEG C of constant incubators, cultivates 6d.Then the bacterial strain after activation on PDA slant medium being prepared into concentration is 2 × 10
7individual/mL embraces sub-suspension, and the inoculum size according to 3% is inoculated in fresh seed culture medium, and wherein seed culture medium consists of: Microcrystalline Cellulose 2%, corn steep liquor 1%, glucose 1%, regulates pH to be 4.5.Be placed in 30 DEG C, on the shaking table that rotating speed is 180 revs/min, concussion is cultivated.
(3) seed culture fluid cultivating 24h is seeded in fresh fermention medium according to 5% inoculum size.Consisting of of fermention medium: Microcrystalline Cellulose 3.3%, corn steep liquor 1.7%, (NH
4)
2sO
40.4%, KH
2pO
40.6%, CaCO
30.25%, MgSO
40.1%, hydrolyzed solution (with glucose meter) adds 0.5%, and disaccharides addition is 0.5g/L, regulates initial 5.0.Contrast is Microcrystalline Cellulose 3.3%, corn steep liquor 1.7%, (NH
4)
2sO
40.4%, KH
2pO
40.6%, CaCO
30.25%, MgSO
40.1%, glucose 0.5%, regulates initial 5.0.Carry out shake flask fermentation cultivation, in 26 DEG C, after 120h is cultivated in concussion on the shaking table that rotating speed is 180 revs/min.The International Standards Method adopting IUPAC (IUPAC) to recommend measures fermentation broth enzyme and lives, and obtaining filter paper enzyme activity is 5.86FPU/mL, and control group filter paper enzyme activity is 5.45FPU/mL.
Embodiment four
(1) acquisition of liquid glucose is induced: prepare zymosan according to method disclosed in Chinese Patent Application No. 201110043824.3, afterwards with the obtained induction sugar of following method process zymosan: at 100 DEG C of temperature, be made into the concentration of substrate hydrolysis 3.5h of 12% with the H2SO4 solution of 0.5mol/L, the hydrolyzed solution of rear generation contains glucose and the 6.5g/L disaccharides content of 23g/L.
(2) bacterial classification obtains: Li's Trichoderma strains is inoculated in PDA inclined-plane, in 28 DEG C of constant incubators, cultivates 6d.Then the bacterial strain after activation on PDA slant medium being prepared into concentration is 2 × 10
7individual/mL embraces sub-suspension, and the inoculum size according to 3% is inoculated in fresh seed culture medium, and wherein seed culture medium consists of: Microcrystalline Cellulose 2%, corn steep liquor 1%, glucose 1%, regulates pH to be 4.5.Be placed in 30 DEG C, on the shaking table that rotating speed is 180 revs/min, concussion is cultivated.
(3) seed culture fluid cultivating 24h is seeded in fresh fermention medium according to 5% inoculum size.Consisting of of fermention medium: Microcrystalline Cellulose 3.3%, corn steep liquor 1.7%, (NH
4)
2sO
40.4%, KH
2pO
40.6%, CaCO
30.25%, MgSO
40.1%, hydrolyzed solution (with glucose meter) adds 0.5%, and disaccharides addition is 1.25g/L, regulates initial 5.0.Contrast is Microcrystalline Cellulose 3.3%, corn steep liquor 1.7%, (NH
4)
2sO
40.4%, KH
2pO
40.6%, CaCO
30.25%, MgSO
40.1%, glucose 0.5%, regulates initial 5.0.Carry out shake flask fermentation cultivation, in 26 DEG C, after 120h is cultivated in concussion on the shaking table that rotating speed is 180 revs/min.The International Standards Method adopting IUPAC (IUPAC) to recommend measures fermentation broth enzyme and lives, and obtaining filter paper enzyme activity is 8.5FPU/mL, and control group filter paper enzyme activity is 5.45FPU/mL.
Embodiment five
(1) acquisition of liquid glucose is induced: prepare zymosan according to method disclosed in Chinese Patent Application No. 201110043824.3, afterwards with the obtained induction sugar of following method process zymosan: at 110 DEG C of temperature, the concentration of substrate hydrolysis 4h of 30% is made into the phosphoric acid solution of 3mol/L, the hydrolyzed solution of rear generation contains glucose and the 3g/L disaccharides content of 27g/L
(2) bacterial classification obtains: Li's Trichoderma strains is inoculated in PDA inclined-plane, in 28 DEG C of constant incubators, cultivates 6d.Then the bacterial strain after activation on PDA slant medium being prepared into concentration is 2 × 10
7individual/mL embraces sub-suspension, and the inoculum size according to 3% is inoculated in fresh seed culture medium, and wherein seed culture medium consists of: Microcrystalline Cellulose 2%, corn steep liquor 1%, glucose 1%, regulates pH to be 4.5.Be placed in 30 DEG C, on the shaking table that rotating speed is 180 revs/min, concussion is cultivated.
(3) seed culture fluid cultivating 24h is seeded in fresh fermention medium according to 5% inoculum size.Consisting of of fermention medium: Microcrystalline Cellulose 3.3%, corn steep liquor 1.7%, (NH
4)
2sO
40.4%, KH
2pO
40.6%, CaCO
30.25%, MgSO
40.1%, hydrolyzed solution (with glucose meter) adds 0.5%, and disaccharides addition is 0.55g/l, regulates initial 5.0.Contrast is Microcrystalline Cellulose 3.3%, corn steep liquor 1.7%, (NH
4)
2sO
40.4%, KH
2pO
40.6%, CaCO
30.25%, MgSO
40.1%, glucose 0.5%, regulates initial 5.0.Carry out shake flask fermentation cultivation, in 26 DEG C, after 120h is cultivated in concussion on the shaking table that rotating speed is 180 revs/min.The International Standards Method adopting IUPAC (IUPAC) to recommend measures fermentation broth enzyme and lives, and obtaining filter paper enzyme activity is 6.16FPU/mL, and control group filter paper enzyme activity is 5.45FPU/mL.
Embodiment six
(1) acquisition of liquid glucose is induced: prepare zymosan according to method disclosed in Chinese Patent Application No. 201110043824.3, afterwards with the obtained induction sugar of following method process zymosan: at 100 DEG C of temperature, be made into the concentration of substrate hydrolysis 2h of 5% with the salpeter solution of 0.5mol/L, the hydrolyzed solution of rear generation contains glucose and the 0.2g/L disaccharides content of 5g/L.
(2) bacterial classification obtains: Li's Trichoderma strains is inoculated in PDA inclined-plane, in 28 DEG C of constant incubators, cultivates 6d.Then the bacterial strain after activation on PDA slant medium being prepared into concentration is 2 × 10
7individual/mL embraces sub-suspension, and the inoculum size according to 3% is inoculated in fresh seed culture medium, and wherein seed culture medium consists of: Microcrystalline Cellulose 2%, corn steep liquor 1%, glucose 1%, regulates pH to be 4.5.Be placed in 30 DEG C, on the shaking table that rotating speed is 180 revs/min, concussion is cultivated.
(3) seed culture fluid cultivating 24h is seeded in fresh fermention medium according to 5% inoculum size.Consisting of of fermention medium: Microcrystalline Cellulose 3.3%, corn steep liquor 1.7%, (NH
4)
2sO
40.4%, KH
2pO
40.6%, CaCO
30.25%, MgSO
40.1%, hydrolyzed solution (with glucose meter) adds 0.5%, and disaccharides addition is 0.2/L, regulates initial 5.0.Contrast is Microcrystalline Cellulose 3.3%, corn steep liquor 1.7%, (NH
4)
2sO
40.4%, KH
2pO
40.6%, CaCO
30.25%, MgSO
40.1%, glucose 0.5%, regulates initial 5.0.Carry out shake flask fermentation cultivation, in 26 DEG C, after 120h is cultivated in concussion on the shaking table that rotating speed is 180 revs/min.The International Standards Method adopting IUPAC (IUPAC) to recommend measures fermentation broth enzyme and lives, and obtaining filter paper enzyme activity is 5.73FPU/mL, and control group filter paper enzyme activity is 5.45FPU/mL.
Embodiment seven
(1) acquisition of liquid glucose is induced: prepare zymosan according to method disclosed in Chinese Patent Application No. 201110043824.3, afterwards with the obtained induction sugar of following method process zymosan: at 100 DEG C, with the H of 0.5mol/L
2sO
4solution is made into the concentration of substrate hydrolysis 3.5h of 12%, and the hydrolyzed solution of rear generation contains glucose and the 6.5g/L disaccharides of 23g/L.
(2) bacterial classification obtains: Li's Trichoderma strains is inoculated in PDA inclined-plane, in 28 DEG C of constant incubators, cultivates 6d.Then the bacterial strain after activation on PDA slant medium being prepared into concentration is 2 × 10
7individual/mL embraces sub-suspension, and the inoculum size according to 3% is inoculated in fresh seed culture medium, and wherein seed culture medium consists of: Microcrystalline Cellulose 2%, corn steep liquor 1%, glucose 1%, regulates pH to be 4.5.Be placed in 30 DEG C, on the shaking table that rotating speed is 180 revs/min, concussion is cultivated.
(3) seed culture fluid cultivating 24h is seeded in fresh fermention medium according to 5% inoculum size.Consisting of of fermention medium: Microcrystalline Cellulose 3.3%, corn steep liquor 1.7%, (NH
4)
2sO
40.4%, KH
2pO
40.6%, CaCO
30.25%, MgSO
40.1%, hydrolyzed solution (with glucose meter) adds 0.5%, and disaccharides addition is 1.25g/L, regulates initial 5.0.Contrast is Microcrystalline Cellulose 3.3%, corn steep liquor 1.7%, (NH
4)
2sO
40.4%, KH
2pO
40.6%, CaCO
30.25%, MgSO
40.1%, glucose 0.5%, regulates initial 5.0.In fermenting process, sugared mixed solution is induced in gradation interpolation (interval 24h adds once), and final disaccharides concentration is 2g/L.Contrast then adds the glucose of equivalent.Shake flask fermentation is cultivated, in 26 DEG C, after on the shaking table that rotating speed is 180 revs/min, 120h is cultivated in concussion.The International Standards Method adopting IUPAC (IUPAC) to recommend measures fermentation broth enzyme and lives, and obtaining filter paper enzyme activity is 9.68FPU/mL, and control group filter paper enzyme activity is 6.34FPU/mL.
As can be seen from above-described embodiment, the interpolation of glucose can not affect the output of cellulase, therefore, the induction that utilizes method of the present invention to prepare sugar can the synthesis of cellulase induction, makes it synthesize increase (enzyme amount in unit volume fermented liquid increases).
Here the module number illustrated and treatment scale are used to simplify explanation of the present invention.Application, the modifications and variations of being prepared by yeast cell of the present invention to the method and fermention medium etc. of zymosan will be readily apparent to persons skilled in the art.
The present invention prepares the method for induction sugar by yeast cell, and the technique of being added in cellulase synthesis by induction sugar is simple, and cost is low, abundant raw material source, and can achieve many things at one stroke, and is suitable for commercial introduction.
Although embodiment of the present invention are open as above, but it is not restricted to listed in specification sheets and embodiment utilization, it can be applied to various applicable the field of the invention completely, for those skilled in the art, can easily realize other amendment, therefore do not deviating under the universal that claim and equivalency range limit, the present invention is not limited to specific details and illustrates here and the embodiment described.
Claims (8)
1. utilize yeast cell to prepare induction sugar and a method for fibrin enzymic synthesis, it is characterized in that, comprise the following steps:
Step one, utilize zymosan prepare induction sugar: at temperature 90-130 DEG C, utilize the acid hydrolysis zymosan 2 ~ 4h of 0.2mol/L-3mol/L, with obtain induction sugar;
Step 2, Trichoderma is inoculated in be added with the induction sugar obtained in step one fermention medium in cultivate, cellulase synthesis.
2. utilize yeast cell to prepare induction sugar and the method for fibrin enzymic synthesis as claimed in claim 1, it is characterized in that, in described step one, described acid is hydrochloric acid, sulfuric acid, phosphoric acid, acetic acid or nitric acid.
3. utilize yeast cell to prepare induction sugar and the method for fibrin enzymic synthesis as claimed in claim 2, it is characterized in that, in described step one, described acid is sulfuric acid.
4. utilize yeast cell to prepare induction sugar and the method for fibrin enzymic synthesis as claimed in claim 1, it is characterized in that, in described step one, the concentration of described zymosan is 1%-30% (w/w).
5. utilize yeast cell to prepare induction sugar and the method for fibrin enzymic synthesis as claimed in claim 1, it is characterized in that, in described step 2, the induction sugar in described fermention medium adds in described fermention medium before fermentation culture or in fermentation culture process.
6. utilize yeast cell to prepare induction sugar and the method for fibrin enzymic synthesis as claimed in claim 1, it is characterized in that, described Trichoderma is Trichodermareesei.
7. utilize yeast cell to prepare induction sugar and the method for fibrin enzymic synthesis as claimed in claim 1, it is characterized in that, comprise induction sugar in described fermention medium, in described induction sugar, the concentration of disaccharides is 0.5 ~ 6.5g/L.
8. utilize yeast cell to prepare induction sugar and the method for fibrin enzymic synthesis as claimed in claim 1, it is characterized in that, described zymosan derives from yeast thalline discarded in fermentation industry.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201610003974.4A CN105505903B (en) | 2016-01-05 | 2016-01-05 | Induction sugar and cellulase induction synthetic method are prepared using yeast cells |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201610003974.4A CN105505903B (en) | 2016-01-05 | 2016-01-05 | Induction sugar and cellulase induction synthetic method are prepared using yeast cells |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| CN105505903A true CN105505903A (en) | 2016-04-20 |
| CN105505903B CN105505903B (en) | 2019-03-22 |
Family
ID=55714198
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN201610003974.4A Active CN105505903B (en) | 2016-01-05 | 2016-01-05 | Induction sugar and cellulase induction synthetic method are prepared using yeast cells |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN105505903B (en) |
Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN107418943A (en) * | 2017-04-12 | 2017-12-01 | 上海大学 | The method of cellulase production derivant and its application in straw saccharification are extracted from stalk |
Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102604916A (en) * | 2002-09-10 | 2012-07-25 | 金克克国际有限公司 | Induction of gene expression using a high concentration sugar mixture |
| CN105131146A (en) * | 2015-08-18 | 2015-12-09 | 陈莉 | Combined extraction of beta-glucan and beta-mannan in yeast cell walls |
-
2016
- 2016-01-05 CN CN201610003974.4A patent/CN105505903B/en active Active
Patent Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102604916A (en) * | 2002-09-10 | 2012-07-25 | 金克克国际有限公司 | Induction of gene expression using a high concentration sugar mixture |
| CN105131146A (en) * | 2015-08-18 | 2015-12-09 | 陈莉 | Combined extraction of beta-glucan and beta-mannan in yeast cell walls |
Non-Patent Citations (3)
| Title |
|---|
| ANNA COLL´EN等: "Protein production and induction of the unfolded protein response in Trichoderma reesei strain Rut‐C30 and its transformant expressing endoglucanase I with a hydrophobic tag", 《ADVANCED BIO SYSTEMS》 * |
| 李娜等: "离子液体[Bmim]Cl预处理对两种高结晶多糖酸水解的影响", 《食品工业科技》 * |
| 陈俊等: "高效液相色谱法测定酵母葡聚糖的前处理条件的研究及优化", 《轻工科技》 * |
Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN107418943A (en) * | 2017-04-12 | 2017-12-01 | 上海大学 | The method of cellulase production derivant and its application in straw saccharification are extracted from stalk |
Also Published As
| Publication number | Publication date |
|---|---|
| CN105505903B (en) | 2019-03-22 |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| CN102199554B (en) | Saccharomyces cerevisiae strain with multiple-stress resistance, and application thereof in cellulose alcohol fermentation | |
| CN102174433B (en) | Clostridium beijerinckii with high stress resistance and application thereof | |
| CN102229920B (en) | Method for improving submerged fermentation level of trichoderma reesei cellulase liquid | |
| CN103993042A (en) | Method for combined production of bioethanol and pullulan from lignocellulose substances | |
| CN106834368A (en) | A kind of method that utilization lignocellulose for fermentation produces L lactic acid | |
| CN101933439A (en) | Method for improving phellinus igniarius hypha amount of submerged culture by utilizing plant oil | |
| CN104911169A (en) | Method for culturing cordyceps militaris mycelium and plasmin | |
| CN105969702B (en) | Serratia marcescens RZ 21-C6 and its application | |
| CN103409382A (en) | Method used for accelerating lignin degradation in phanerochaete chrysosporium solid state fermentation | |
| CN102174600B (en) | Method for producing L-lactic acid through continuous fermentation | |
| CN103343156B (en) | A kind of cellulose raw material solid state fermentation produces the method and apparatus of alcohol fuel | |
| CN107760753A (en) | Method for producing butanol by co-culture fermentation of high-temperature anaerobe for pyrolyzing sugar and clostridium acetobutylicum | |
| CN102234670B (en) | Method for producing bacterial cellulose through solid state fermentation by using inert adsorption carrier | |
| CN103952447A (en) | Method for producing succinic acid by fermentation under anaerobic condition | |
| CN103740675A (en) | Method for producing cellulase | |
| CN103461654A (en) | Method for producing vinasse protein feed from corn steep liquor | |
| CN105505903A (en) | Method for preparing induction sugar and inducing the synthesis of cellulose through yeast cells | |
| CN103343151B (en) | Preparation method of liquid medium for bacterial cellulose film | |
| CN109536565A (en) | Method for producing succinic acid by utilizing mixed fermentation of high-temperature anaerobic bacteria for pyrolyzing sugar and actinobacillus succinogenes | |
| CN109536558B (en) | Process for preparing beta-carotene | |
| CN102010856B (en) | Solid fermentation method for producing salt-tolerant cellulase by using oceanic Aspergillus niger | |
| CN103614418A (en) | Method for producing fuel ethanol through synchronous saccharification and fermentation | |
| CN104178438B (en) | One strain is suitable for the moral formula lactobacillus of molasses fermented production high-purity L-lactic acid and fermentation process and application | |
| CN103497941A (en) | Method for preparing cellulase through trichoderma viride high-efficiency fermentation | |
| CN102433321B (en) | Fused lactobacillus capable of utilizing xylose for fermenting lactic acid and resisting high temperature and breeding method of fused lactobacillus |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| C06 | Publication | ||
| PB01 | Publication | ||
| C10 | Entry into substantive examination | ||
| SE01 | Entry into force of request for substantive examination | ||
| GR01 | Patent grant | ||
| GR01 | Patent grant |