CN1157475C - Bacterial strain producing phloem fibre degumming enzyme and its application in ramie and hemp degumming - Google Patents
Bacterial strain producing phloem fibre degumming enzyme and its application in ramie and hemp degumming Download PDFInfo
- Publication number
- CN1157475C CN1157475C CNB011274409A CN01127440A CN1157475C CN 1157475 C CN1157475 C CN 1157475C CN B011274409 A CNB011274409 A CN B011274409A CN 01127440 A CN01127440 A CN 01127440A CN 1157475 C CN1157475 C CN 1157475C
- Authority
- CN
- China
- Prior art keywords
- degumming
- enzyme
- phloem
- fibre
- bacterial strain
- 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.)
- Expired - Fee Related
Links
Abstract
The present invention relates to a strain which can generate phloem fiber degummase and an application thereof to the degumming of ramie and hemp, which belongs to the technical field of microbes. The bacterium is identified to be Bacillus subtilis No. 13, and a preservation number is CCTCC, No. M200038. The present invention has the advantages of extensive culture condition, quick growth and propagation and good degumming effect, and various enzymes needed by degumming can be generated under the same culture condition, which is completed for 2 to 6 hours. Degumming rate reaches 95%, and the dispersion degree of fibre is 100%. The fibre is not damaged, environment is not polluted, and the present invention can replace a chemical degumming method and is used for the degumming technique of the ramie and the hemp.
Description
(1) technical field
The present invention relates to the bacterial strain and the application in ramie, hemp degumming thereof of a strain producing phloem fibre degumming enzyme.
(2) technical background
Since over half a century, people attempt to change the employing environmental pollution that chemical degumming technique brought and the part deterioration of flaxen fiber textile performance, therefore many research workers of this area are striving to find a kind of novel method of retting, and biological degumming has been subjected to special concern.Nineteen fifty-three, Shenyang forestry soil Liu of institute of Chinese Academy of Sciences phase pine etc. is studied the China grass degumming micro-flora, Qiao Hege (Chaudhurg.S.D.) once reported some bacterium and fungi during the fermentation can the decompose pectin material and discharge fiber, 1958 Ali (Ali.M.M.) propose to utilize polygalacturonase to carry out coming unstuck of crudefiber crop (as jute, flax etc.) fiber.1972, good fortune Sha carried (Fogarty W.M.) and Ward (Ward D.D.) report and produce the microorganism of polygalacturonase or the possibility that pectase preparation is used in degumming technology.1976, the Japan rattan heavily rises the method (the Japanese Patent spy opens clear 51-149976) that discloses a kind of microorganism forever and carried out China grass degumming, contain cellulase and the suitableeest effect PH in the mycetogenetic polygalacturonase for acid, thereby cause the decline of fibre strength, the degumming effect instability is difficult to use in suitability for industrialized production.Chinese patent CN85104285 and for example, CN-85104284, CN-89104529 disclose and have utilized genus bacillus to carry out the method for biological degumming of ramie.Chinese patent 97109044 discloses a strain Alkaliphilic bacillus and a ramie degumming method thereof, but owing to the relatively lower usually time of pectinase activity long (16～24 hours) of its generation also is difficult in industrial application.Above-mentioned patent does not all relate to the content to hemp degumming.
(3) summary of the invention
The invention provides the bacterial strain and the application in ramie, hemp degumming thereof of a strain producing phloem fibre degumming enzyme.
Technical scheme of the present invention comprises: the separation of bacterial classification, mutagenic and breeding, cultivate, shake bottle and fermentor tank produce enzymic fermentation with and application on ramie, hemp degumming.
By method known to those skilled in the art, gather pedotheque in the suburbs, Jinan, select to separate, carry out just, sieve again through shake flask fermentation, then through ultraviolet ray and processing such as lithium chloride complex mutation and protoplastis fusion, and according to degumming effect and enzymic activity height binomial standard, screen, obtain the bacterial strain of strain laboratory numbering No.13, its biological characteristics is: thalline is shaft-like, Gram-positive (G
+), the statospore near-end is given birth to, and sporocyst is less than or equal to thalline, spore ellipse, aerobic, hydrogen peroxide enzyme positive, to energy fermentation and acids such as glucose, fructose, sucrose, lactose, semi-lactosi are not produced acid, can hydrolyzed starch, well-grown in containing the meat soup of 7%NaCl.Identify that according to uncle's outstanding Bacteria Identification handbook (the 8th edition 1984 Science Presses) and pertinent literature this bacterium is subtilis (Bacillus subtilis) No.13.On November 20th, 2000 this bacterium is submitted to Chinese typical culture collection, guarantor's minus sign is CCTCCNO.M200038.
Described subtilis (Bacillus subtilis) No.13 CCTCC NO.M200038 is containing on the slant medium of following proportioning, the characteristic of maintenance high yield enzyme that can be more stable, slant culture based formulas (being weight proportion) is: 10～20% murphy juices 100, sucrose 1.0～2.0, pectin 0.2～0.4, yeast extract paste 0.03～0.05, agar 1.5～1.8; Or extractum carnis 0.3, peptone 1.0, NaCl 0.5, agar 1.8～2.0, water 100; PH 7.2～7.4, and 100Pa sterilization 20～30 minutes is placed to the inclined-plane, with this bacterial classification of transfering loop streak inoculation, cultivate 24～48 hours for 30 ℃～34 ℃, after cultivation finishes, put 4 ℃ of refrigerators and preserve standby.
Described subtilis (Bacillus subtilis) No.13 CCTCC NO.M200038 is under the culture condition of regulation, can produce has high efficiency degummase (mainly containing polygalacturonase, zytase, mannase etc.) to phloem fiber, but does not produce the cellulase (as the C1 enzyme) of damage fiber.This culture condition is: culture medium prescription (being weight proportion): wheat bran 2.0～5.0, Semen Maydis powder 0.5～2.0, (NH
4)
2SO
40.5～1.5, soybean cake powder 0.5～1.5, K
2HPO
10.05～0.1, MgSO
17H
2O 0.1～0.2, water 100, and before disappearing, pH transfers to 7.5～8.5,100Pa sterilization 20～30 minutes; Fermentating controling condition: 30 ℃～34 ℃ of culture temperature, shake 180～220 rev/mins of bottle rotating speeds, incubation time 48～72 hours; Behind filtration of shake flask fermentation liquid or the low-speed centrifugal, dilute 500～1000 times, measure the enzyme activity of polygalacturonase (is substrate with the poly-half congealed aldehydic acid of gala of Sigma), zytase (is substrate with the Sigma xylan), mannase (is substrate with the Sigma locust bean gum) by the SomogyiShi method.Enzyme activity unit is defined as: under condition determination, discharge the required enzyme amount of 1 μ mol product by the substrate per minute, promptly an enzyme activity unit is represented with U/ml.
The condition that the bacterial strain subtilis of producing phloem fibre degumming enzyme (Bacillus subtilis) No.13 CCTCCNO.M200038 fermentor tank enlarged culturing produces degummase is: 1. technical process: slant strains → shake a bottle bacterial classification → first order seed → secondary seed → enzymatic production, 2. the culture medium prescription of I and II seed and enzymatic production (being weight proportion): wheat bran 3.0～5.0, Semen Maydis powder 1.0～2.0, soybean cake powder 0.5～1.5, (NH
1)
2SO
10.5～1.0, K
2HPO
10.05～0.10, MgSO
17H
2O 0.1～0.2, and before disappearing, pH transfers to 7.5～8.5,3. I and II seed and enzymatic production control condition: tank pressure: 0.03Mpa, and temperature: 32 ℃～34 ℃, air flow (volume of air m
2/ fermentating liquid volume m
1Minute) 1: 0.5～0.8, incubation time I and II seed is 8～14 hours, produce 12～18 hours enzymic fermentation time, 4. fermentation ends puts a jar standard, polygalacturonase 400U/ml～2000U/ml, fermented liquid pH 5.5～6.5, free from extraneous odour, thalli growth are good, and no spore or minority thalline produce spore.
The degummase liquid of above-mentioned production can be used for coming unstuck of ramie or hemp after suitably handling.Suitably the condition of handling is: dilute with water enzyme liquid transfers to the pH that dilutes enzyme liquid between 7.4～9.6; Consumption and the dilution enzyme liquid measure of raw ramie of coming unstuck be 1: 10～20 (weight: volume), the temperature of coming unstuck is controlled at 35 ℃～50 ℃, and it is static or dynamically that degummase liquid keeps, usually time 2～6 hours, the fiber crops of coming unstuck are through washing fiber crops machine washing fiber crops, back operation is carried out routinely.The ramie degummed ramie of making, its residual gum content can reach 2.18% (chemical method 3.0%), breaking tenacity 4.30CN/dtex (chemical method is 4.70CN/drex), the product row yielding is higher than chemical method 2.5%, compare with chemical method in addition, environmentally safe improves workman's labor condition etc.
(4) embodiment
Embodiment 1:
CCTCC No.M200038 bacterium preservation slant strains standby or new cultivation is inoculated and is shaken in the bottle product enzyme substratum.Its preparation method is: 300ml shakes bottled product enzyme substratum 50ml and puts 32 ℃ ± 1 ℃, and 200 rev/mins of shaking tables were cultivated 72 hours.Producing the enzyme culture medium prescription counts with weight part: 3.0 parts in wheat bran, 1.0 parts of Semen Maydis powder, (NH
4)
2SO
11.0 part, 0.5 part of soybean cake powder, K
2HPO
40.1 part, MgSO
47H
20.1 part of O, 100 parts in water, before disappearing, pH transfers to 8.0.After cultivation finishes, take off that to shake bottle centrifugal with 4 layers of filtered through gauze or 2000～3000 rev/mins with fermented liquid, get clear liquid, press the SomogyiShi method and measure enzyme activity, polygalacturonase (is substrate with the poly-half congealed aldehydic acid of gala of Sigma) vigor 1075U/ml, zytase (is substrate with the Sigma xylan) vigor 61U/ml, mannase (is substrate with the Sigma locust bean gum) vigor 56U/ml.
Embodiment 2:
The embodiment 1 enzyme liquid that produces is after diluting 5 times, transfer pH 8.5, the bath raio of dilution enzyme liquid and hemp is by 10～16: 1 (volume: weight), putting 40 ℃～50 ℃ water bath heat preservations came unstuck 4 hours, after taking out washing, send Shandong Prov. Fibre Inspection Office to measure, its leading indicator result is: fiber fineness 12.9～18.25dtex, fibre breakage intensity 3.07～3.14CN/dtex, elongation 2.5～1.9CV%.
Embodiment 3:
CCTCC No.M200038 bacterium is at fermentor tank enlarged culturing thalline and produce enzyme and be applied to ramie, hemp degumming.
The technical process that the fermentor tank enlarged culturing produces degummase is: slant strains → shake a bottle bacterial classification → 50L seeding tank → 500L seeding tank → 10M
3Fermentor tank
The condition that the fermentor tank enlarged culturing is produced degummase is: 1. 50L, 500L and 10M
3The fermentor cultivation based formulas is counted with weight part: 3.0 parts in wheat bran, 1.0 parts of Semen Maydis powder, 0.5 part of soybean cake powder, (NH
4)
2SO
41.0 part, K
2HPO
40.10 part, MgSO
17H
20.1 part of O, before disappearing, pH transfers to 7.5～8.5,2. 50L, 500L and 10M3 fermentor tank control condition: tank pressure: 0.03Mpa, temperature: 32 ℃～34 ℃, air flow 1: 0.5～0.8, incubation time 50L jar 12 hours, 500L jar 10 hours, 10M
3Jar, 18 hours, 3. pectinase activity 508U/ml; The whole pH 5.5 of fermented liquid, free from extraneous odour, thalli growth is good, and quantity is more.
About 4.5M
3Degummase liquid is used for former retting 4 cages of ramie, and every cage mounted raw ramie 500kg amounts to 2000kg, adjusts the dilution enzyme liquid pH 7.5～8.5 that comes unstuck when coming unstuck, and is incubated 30 ℃～45 ℃, and ventilation is flowed degummase liquid.First cage comes unstuck and finished in 3 hours, and second and third cage was finished in 3.5 hours, and the 4th cage was finished in 4 hours.The degummed ramie row yielding is 62.5%, is higher than chemical method 2.5%, and residual gum content 2.18%, chemical method are 3.0% (top grade standard≤3.0%), breaking tenacity (CN/drex) 4.30%, and chemical method is 4.70% (top grade product standard 〉=3.53%).
Claims (5)
1. the bacterial strain of a strain producing phloem fibre degumming enzyme, this bacterium is subtilis (Bacillus subtilis) No.13CCTCC No.M200038; Its biological characteristics is: thalline is shaft-like, Gram-positive, statospore near-end are given birth to, sporocyst is less than or equal to thalline, spore ellipse, aerobic, hydrogen peroxide enzyme positive, to energy fermentation and acids such as glucose, fructose, sucrose, to lactose, semi-lactosi do not produce acid, can hydrolyzed starch, well-grown in containing the meat soup of 7%NaCl, this bacterium have been submitted Chinese typical culture collection center preservation on November 20th, 2000.
2. the bacterial strain of a strain producing phloem fibre degumming enzyme as claimed in claim 1, its slant culture and the standby condition of preservation are: the slant culture based formulas, be weight proportion, common beef extract-peptone agar slant: extractum carnis 0.3, peptone 1.0, NaCl 0.5, agar 1.8～2.0, water 100, pH transfers to 7.2～7.4, perhaps murphy juice inclined-plane: 10～20% murphy juices 100, sucrose 1.0～2.0, pectin 0.2～0.4, yeast extract paste 0.03～0.05, agar 1.5～1.8, pH 7.2～7.4,30 ℃～34 ℃ of culture temperature, incubation time 24～48 hours, after cultivation finishes, put 4 ℃ of refrigerators and preserve standby.
3. the bacterial strain of a strain producing phloem fibre degumming enzyme as claimed in claim 1 or 2, the condition that its shake flask fermentation produces enzyme is: culture medium prescription is weight proportion: wheat bran 2.0～5.0, Semen Maydis powder 0.5～2.0, (NH
4)
2SO
40.5～1.5, soybean cake powder 0.5～1.5, K
2HPO
10.05～0.1, MgSO
47H
2O 0.1～0.2, water 100, and before disappearing, pH transfers to 7.5～8.5,100Pa sterilization 20～30 minutes; Fermentating controling condition: 30 ℃～34 ℃ of culture temperature, shake 180～220 rev/mins of bottle rotating speeds, incubation time 48～72 hours; Behind filtration of shake flask fermentation liquid or the low-speed centrifugal, dilute 500～1000 times, press the vigor that the SomogyiShi method is surveyed polygalacturonase, zytase, mannase.
4. the application of the bacterial strain of a strain producing phloem fibre degumming enzyme as claimed in claim 3 in hemp, China grass degumming.
5. the application of the bacterial strain of a strain producing phloem fibre degumming enzyme as claimed in claim 4 in hemp, China grass degumming, its bacterial classification in the processing condition that 10 cubic metres of fermentor tank enlarged culturing produce degummases is: 1. technical process: slant strains → shake a bottle bacterial classification → first order seed → secondary seed → enzymatic production, 2. the culture medium prescription of I and II seed and enzymatic production, be weight proportion: wheat bran 3.0～5.0, Semen Maydis powder 1.0～2.0, soybean cake powder 0.5～1.5, (NH
4)
2SO
10.5～1.0, K
2HPO
10.05～0.10, MgSO
17H
2O 0.1～0.2, and before disappearing, pH 7.5～8.5,3. I and II seed and enzymatic production control condition: tank pressure: 0.03Mpa, temperature: 32 ℃～34 ℃, air flow 1: 0.5～0.8, incubation time I and II seed is 8～14 hours, produce 12～18 hours enzymic fermentation time, 4. fermentation ends puts a jar standard, polygalacturonase 400U/ml～2000U/ml, fermented liquid pH 5.5～6.5, free from extraneous odour, thalli growth are good, and no spore or minority thalline produce spore.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CNB011274409A CN1157475C (en) | 2001-09-26 | 2001-09-26 | Bacterial strain producing phloem fibre degumming enzyme and its application in ramie and hemp degumming |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CNB011274409A CN1157475C (en) | 2001-09-26 | 2001-09-26 | Bacterial strain producing phloem fibre degumming enzyme and its application in ramie and hemp degumming |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| CN1371990A CN1371990A (en) | 2002-10-02 |
| CN1157475C true CN1157475C (en) | 2004-07-14 |
Family
ID=4667395
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CNB011274409A Expired - Fee Related CN1157475C (en) | 2001-09-26 | 2001-09-26 | Bacterial strain producing phloem fibre degumming enzyme and its application in ramie and hemp degumming |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN1157475C (en) |
Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN100422313C (en) * | 2006-07-31 | 2008-10-01 | 江南大学 | Basic catalase high-yield strain and basic catalase prepared by fermentation method using same |
Families Citing this family (12)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101575817B (en) * | 2009-06-15 | 2011-05-04 | 大连工业大学 | Method for degumming mulberry bast fiber microbe in solid state |
| CN101654660B (en) * | 2009-07-14 | 2012-05-09 | 东华大学 | Bacillus subtillis strain containing ramie degumming activity, preparation and application thereof |
| CN102206874B (en) * | 2010-03-30 | 2014-07-30 | 上海康地恩生物科技有限公司 | Degumming method of flax and enzyme preparation for using in degumming |
| CN101948788B (en) * | 2010-09-03 | 2011-11-09 | 湖南润久科技有限公司 | Screening method for degumming strain and degumming strain screened by same |
| CN102337235B (en) * | 2011-08-25 | 2013-04-17 | 西华大学 | Complex bacterial agent for efficiently ungluing bast fiber and preparation method for Complex bacterial agent |
| WO2016061827A1 (en) * | 2014-10-24 | 2016-04-28 | 华中科技大学 | Biological degumming method for ramie |
| CN105400729A (en) * | 2015-07-16 | 2016-03-16 | 山东省食品发酵工业研究设计院 | Antibacterial bacillus subtilis strain producing xylanase |
| CN105567592A (en) * | 2016-01-05 | 2016-05-11 | 天津科技大学 | Bacillus subtilis capable of simultaneously producing pectase and hemicellulase, screening method and applications thereof |
| CN105567604A (en) * | 2016-02-16 | 2016-05-11 | 江苏中通生物科技有限公司 | Compound microorganism bacterium agent and preparation method thereof |
| CN105567605A (en) * | 2016-02-19 | 2016-05-11 | 黑龙江省科学院大庆分院 | Lysinibacillus sp. and application thereof, degumming auxiliary containing lysinibacillus sp. and preparation method of degumming auxiliary |
| CN107828757B (en) * | 2017-10-11 | 2018-12-25 | 中国农业科学院麻类研究所 | It can be used for the enzyme preparation and its crudefiber crop bast degumming tech of crudefiber crop degumming |
| CN110331109B (en) * | 2019-07-24 | 2020-11-03 | 山东省农业科学院农业资源与环境研究所 | Bacillus subtilis and culture method and application thereof |
-
2001
- 2001-09-26 CN CNB011274409A patent/CN1157475C/en not_active Expired - Fee Related
Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN100422313C (en) * | 2006-07-31 | 2008-10-01 | 江南大学 | Basic catalase high-yield strain and basic catalase prepared by fermentation method using same |
Also Published As
| Publication number | Publication date |
|---|---|
| CN1371990A (en) | 2002-10-02 |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| CN102559506B (en) | Penicillium oxalicum and application thereof | |
| CN1157475C (en) | Bacterial strain producing phloem fibre degumming enzyme and its application in ramie and hemp degumming | |
| Chinedu et al. | Assessment of growth and cellulase production of wild-type microfungi isolated from Ota, Nigeria | |
| CN101864366B (en) | Penicillium citrinum bacterial strain and application thereof | |
| CN103540581A (en) | Production method and applications of compound enzyme liquid used in papermaking and pulping process of wheat straws and rice straws | |
| KR20110119961A (en) | Cellulase producing nectria cinnabarina and its use for saccharification | |
| CN111518731B (en) | Bacillus subtilis with antagonistic effect for degrading cellulose at low temperature and application thereof | |
| CN101157903A (en) | Producing Strain for beta- mannose and preparation method thereof | |
| CN101463327B (en) | Use of Eupenicillium javanicum strain in producing cellulase | |
| CN1135263C (en) | Process for preparing pectinase for degumming ramie and its application in degumming ramie | |
| CN1055118C (en) | Bacillus subtilis and producing tech. for solid alkaline degumming enzyme | |
| CN101684571A (en) | Method for biologically degumming bast fibers | |
| CN1188513C (en) | Prepn and use of enzyme prepn for use in hemp degluing industrial process | |
| JPH047191B2 (en) | ||
| CN1177003A (en) | Novel basophilic Bacillus strain and application in degumming of ramie | |
| CN103131639B (en) | Trichoderma longibrachiatum strain and application thereof | |
| CN102337235B (en) | Complex bacterial agent for efficiently ungluing bast fiber and preparation method for Complex bacterial agent | |
| KR100449170B1 (en) | Cellulases and xylanase-producing Aspergillus niger KK2, produced enzymes and solid materials thereof | |
| CN112226380B (en) | Bacillus subtilis capable of degrading cellulose and application and preparation thereof | |
| CN110835610B (en) | Composite microbial inoculum suitable for degrading straw and preparation method thereof | |
| CN113005058A (en) | Apocynum degumming biological preparation and preparation method thereof | |
| RU2195490C2 (en) | Strain of mycelioid fungus trichoderma longibrachiatum as producer of carbohydrases complex containing cellulase, beta-glucanase, xylanase, pectinase and mannanase | |
| CN1130216A (en) | Technology and equipment for ramie biological degumming process | |
| CN102787076B (en) | Cold-resistant pseudogymnoascus roseus and application in preparing cold water cellulase | |
| CN1267547C (en) | High yield fungus for alkali pectinase and screen method as well as ferentation method for producing alkali pectinase by using the strain |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| C10 | Entry into substantive examination | ||
| SE01 | Entry into force of request for substantive examination | ||
| C06 | Publication | ||
| PB01 | Publication | ||
| C10 | Entry into substantive examination | ||
| SE01 | Entry into force of request for substantive examination | ||
| C14 | Grant of patent or utility model | ||
| GR01 | Patent grant | ||
| C19 | Lapse of patent right due to non-payment of the annual fee | ||
| CF01 | Termination of patent right due to non-payment of annual fee |