CN1322681A - Biological flocculant prepared with Nocardia bacteria - Google Patents
Biological flocculant prepared with Nocardia bacteria Download PDFInfo
- Publication number
- CN1322681A CN1322681A CN 01108045 CN01108045A CN1322681A CN 1322681 A CN1322681 A CN 1322681A CN 01108045 CN01108045 CN 01108045 CN 01108045 A CN01108045 A CN 01108045A CN 1322681 A CN1322681 A CN 1322681A
- Authority
- CN
- China
- Prior art keywords
- rea
- biological flocculant
- biological
- obtains
- flocculant
- 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
Images
Landscapes
- Separation Of Suspended Particles By Flocculating Agents (AREA)
- Preparation Of Compounds By Using Micro-Organisms (AREA)
- Micro-Organisms Or Cultivation Processes Thereof (AREA)
Abstract
The present invention relates to microbial preparation technology. In the fermenting culture medium with cane sugar as carbon source and corn milk and urea as nitrogen source, Nocardia bacteria WSH-A11 is used to prepare new tye biological flacculant REA-11 with relatively high flocculation activity. The biological flocculant is one kind of proteoglycan with galacturonic acid as main component. The present invention lays the foundation for directing breeding design and realizing metabolism regulation and promotes the industrial production of biological flocculant.
Description
The present invention relates to a kind of new bio flocculation agent REA-11 of Nocardia bacteria (Nocardia sp.) WSH-A11 preparation, belong to the microbial preparation technical field.
Biological flocculant is a kind of novel, the efficient water treatment agent that microorganism produces during the fermentation.Compare with traditional chemosynthesis flocculation agent, biological flocculant has unique advantages such as application security height, non-secondary pollution, source be wide, thereby in recent years, enhancing gradually along with people's environmental consciousness and health perception, the research of biological flocculant is subjected to home and abroad environment engineering scholar's attention, and the gesture of substituted chemistry synthetic flocculant is arranged greatly.The bacterium for producing flocculant wide material sources of having found at present relate to bacterium, actinomycetes, yeast and mould.Different microorganisms can be synthesized different types of biological flocculant under different fermentation conditions, the biological flocculant of having identified at present mostly is function polysaccharide and function protein matter greatly, the perhaps complex body of several organic constituents such as glycoprotein, proteoglycan, lipoprotein etc. also have the only a few biological flocculant to belong to DNA class material in addition.Nakamura extracts a kind of flocculation agent from the fermented liquid of Aspergillus sojae AJ7002, this flocculation agent is a kind of aminosugar and proteinic complex body (Nakamura J. based on GalN, Miyashiro S., Hirose Y., Agri.Biol.Chem., 1976,40 (3): 619-624); Levy has invented from the mixed polysaccharide biological flocculant F-J1 of a kind of sulphating of Cyanobacterium phrmidium sp.J1 synthetic, its main component is a uronic acid, rhamnosyl, seminose and semi-lactosi (Levy N., Baror Y., Magdassi S., Colloids andSurfaces, 1990,48:337-349); The Al-201 that Alcaligenes cupidus KT201 metabolism produces be a kind of serve as biological flocculant (the Toeda K. of main composition with 42.5% glucose, 36.38% semi-lactosi, 8.52% glucuronic acid and 10.3% acetate, Kurane R., Agric.Biol.Chem.1991,55 (11): 2793-2799); The PF-101. flocculation agent that Paecilomyces sp.I-1 produces be by galn with α-1,4 glycosidic link link to each other the mucopolysaccharide that forms (Levy N., Baror Y., Magdassi S., Colloids and Surfaces, 1990,48:337-349); M.Watanabe has then reported a kind of bacterial strain Rhodovulum sp.PS88 (M.Watanabe that can produce DNA class flocculation agent, K.Sasaki, Y.Nakashimada, T.Kakizono, N.Noparatnaraporn, N.Nishio, Appl.Microbiol.Biotechnol.1998,50 (6): 682-691); The people such as Kurane of Japan have carried out comparatively deep research to the flocculation agent NOC-1 that Rhodococcus erythropolis S-1 (former Nocardia Pseudomonas) produces, include bacterial screening, culture condition is optimized, the separation and purification of agglutinating matter and application thereof, Takeda et al. once utilized propyl carbinol-ammonium sulfate method that the active substance that Rhodococcus erythropolis S-1 produces is separated from fermented liquid, be accredited as proteinaceous flocculation agent NOC-1 (M.Takeda, R.Kurane, J.Kiozumi, I.Nakamura, Agric.Biol.Chem.1991,55 (10): 2663-2664); Thereafter, Kurane but utilizes the acetone extract method to obtain the lipid flocculation agent from the nutrient solution of the similarity condition of same strain bacterium, and its lipid conformation detailed evaluation (R.Kurane.K.Hatamochi, T.Kakuno, K.Klyohara have been carried out, K.Kawaguchi, Y.Mizuno, M.Hirano, Y.Taniguchi, Biosci.Biotech.Biochem.1994,58 (11): 1977-1982).At present the biological flocculant of finding is mostly owing to yield poorly, and a little less than the flocculation activity, production cost is high and limited its applying in practice.
The objective of the invention is a kind of biological flocculant REA-11 of seed selection from Nocardia bacteria (Nocardia sp.) WSH-A11 preparation, product of the present invention is the new bio flocculation agent with strong flocculation activity, by to its analysis of the molecular structure, be design breeding and metabolic regulation based theoretical, the actual condition that provides of applying of biological flocculant is provided simultaneously, accelerates the process of industrialization of biological flocculant.
Concrete operations step of the present invention is as follows:
1. bacterial classification
Nocardia bacteria (Nocardia sp.) WSH-A11 screens in soil and obtains (being deposited in Wuhan China typical culture collection center, preserving number CCTCC NO.M201005)
(1) bacterial screening method:
Screening culture medium (gL
-1): sucrose 10, yeast extract paste 0.5, urea 0.5, KH
2PO
40.1, K
2HPO
40.1, NaCl0.1, MgSO
47H
2O0.2, agar 20, pH8.0
Fermention medium (gL
-1): sucrose 10, yeast extract paste 0.5, urea 0.5, KH
2PO
42, K
2HPO
45, NaCl0.1, MgSO
47H
2O0.2, pH8.0
Culture condition is 150 rev/mins, and 28 ℃ of shaking culture detected the flocculation activity of fermented liquid after 2～3 days.
Prescreening method: get the 1mL fermented liquid and add to 40mL kaolin solution and 2.5mL1%CaCl are housed
250mL scale test tube in, be settled to scale with distilled water, behind the mixing, leave standstill a moment, the flocculation situation of range estimation kaolin solution is selected high flocculation activity and is produced bacterial strain as sieving bacterial strain again.
Multiple screen method: from the inclined-plane, respectively get a ring bacterium, be connected in the fermention medium, 3 bottles of every strains, 28 ℃, 150 rev/mins of shaking culture 2～3 days detect the flocculation activity of fermented liquid, select the synthetic bacterial strain of high flocculation agent.
(2) fungus characteristic:
Bacterium colony does not have aerial hyphae, quality softness, white, thickness;
Be rod-short in the liquid nutrient medium, splayed configuration or be arranged in parallel, late stage of culture produces a small amount of yellowish pigment, and thalline becomes near-spherical or extremely short and small shaft-like simultaneously;
Can utilize glucose, sucrose, fructose, sodium acetate and succinate well-grown; Can utilize nitrate, Citrate trianion, lactose and starch growth; Substantially can not utilize the growth of nitrite and formate.
The energy liquefy gelatin, part is antiacid, not hydrolyzed starch.
2, zymotechnique
Liquid fermentation medium is formed (gL
-1): sucrose 10, corn steep liquor 0.5, urea 0.5, KH
2PO
40.1, NaCl0.1, MgSO
4.7H
2O0.2
Can prepare biological flocculant REA-11 when (3) being carbon source with greater activity with 1% sucrose;
(4) corn steep liquor and urea can prepare the biological flocculant REA-11 with greater activity so that 1: 1 (mass ratio) is composite during as culture media nitrogen source;
(5) liquid fermentation condition is 25～30 ℃ (best 28 ℃), the initial pH4.0 of substratum～10.0 (best 6.0), 120～150 rev/mins of (the suitableeest 120 rev/mins) shaking culture of shaking speed 48 hours.
3, the extraction of biological flocculant REA-11
(6) fermented liquid is centrifugal, and condition is: 6000～10000 rev/mins, somatic cells and the solid substance removed in the fermented liquid in centrifugal 20 minutes obtain supernatant liquor;
(7) in the centrifuged supernatant that (6) obtain, add 3 times of volume of ethanol, 4 ℃ leave standstill 6 hours after, 7000 rev/mins centrifugal 10 minutes, obtain throw out;
(8) precipitation that obtains of dissolved in distilled water (7), (PEG-20000) is concentrated into 1/10 of original fermented solution volume with polyoxyethylene glycol, utilizes the DEAE ion exchange column to carry out purifying, obtains active eluant;
(9) collect the active eluant that (8) obtain, after concentrating, utilize gel column SephadexG-100 to be for further processing, obtain gel column and handle active eluant;
(10) collect the gel column processing active eluant that (9) obtain, with ethanol sedimentation and washing;
(11) precipitation that obtains of dissolved in distilled water (10) utilizes preparation high pressure liquid chromatography post Superdex200 HR10/30 to carry out the refinement treatment of active substance, promptly gets product REA-11.
4, the component of biological flocculant is identified
(12) utilize high-pressure liquid phase, gas-chromatography, mass spectrum etc. to analyze means of testing, in conjunction with the feature chemical reaction, the refining REA-11 that obtains is formed evaluation, the result shows, biological flocculant REA-11 by Nocardia sp.WSH-A11 preparation is a kind of acidic polysaccharose based on galacturonic acid, the protein ingredient that also contains denier in its molecule assists to keep the flocculation activity of this biomacromolecule.The GC-MS of REA-11 identifies that collection of illustrative plates is shown in accompanying drawing 1, accompanying drawing 2-1 and accompanying drawing 2-2.
Bacterium for producing flocculant---the Nocardia sp.WSH-A11 that screening obtains from soil can be the biological flocculant REA-11 that the fermented substrate preparation has higher flocculation activity with cheap raw materials such as sucrose, corn steep liquor, urea, has reduced production cost.By refining to the extraction of REA-11, and and then carry out structure and identify, find that REA-11 is a kind of proteoglycans new bio flocculation agent based on galacturonic acid, do not see the report of this type of biological flocculant at present both at home and abroad as yet.More than be found to be and instruct the design breeding and improve output by the metabolic regulation optimization of fermentation conditions and lay a good foundation, also for the production cost that reduces biological flocculant provide may, promoted the practical application process of biological flocculant in industrial production.
Following example has been explained some key point among the present invention in more detail, the invention provides foundation for understanding better.
Choose several different carbon sources respectively, under the identical situation of other zymotechniques (as described in specification sheets), carry out the fermentative preparation of biological flocculant, result such as table 1:
Table 1 carbon source is to the influence of thalli growth and biological flocculant preparation
Carbon source flocculating rate/% dry cell weight/gL
-1
Glucose 85 1.62
Fructose 85.3 1.58
Sucrose 90.5 1.62
Lactose 23.3 0.34
Starch 0 0.31
Sodium acetate 43.3 1.63
Ethanol 9.5 1.19
When result's demonstration is carbon source with sucrose, thalli growth good (dry cell weight), flocculation activity (flocculating rate) is higher, and product cost is lower.
Sucrose with 1% is the carbon source of fermention medium, selects different organic nitrogen sources and urea composite by 1: 1 (mass ratio) respectively, under the identical situation of other zymotechniques (as described in specification sheets), carries out the fermentative preparation of biological flocculant, result such as table 2:
Table 2 compound nitrogen source is to the influence of thalli growth and biological flocculant preparation
Nitrogenous source flocculating rate/% dry cell weight/gL
-1
Urea 53.3 1.08
Yeast extract paste+urea 88.23 1.63
Extractum carnis+urea 65 1.01
Tryptones+urea 87.82 1.45
Peptone+urea 89.93 1.40
Soybean cake powder+urea 69.27 1.66
Corn steep liquor+urea 91.89 1.70
The result shows, corn steep liquor and urea is during with 1: 1 (mass ratio) composite nitrogenous source as fermention medium, thalli growth good (dry cell weight), and flocculation activity (flocculating rate) is also higher, and product cost is lower.
Claims (9)
1, a kind of biological flocculant for preparing from Nocardia bacteria, it is bacterium for producing flocculant that the present invention is characterized as with Nocardia bacteria (Nocardia sp.) WSH-A11, with sucrose is carbon source, and corn steep liquor and urea are composite to prepare a kind of new bio flocculation agent REA-11 during for nitrogenous source
(1) bacterial classification:
Nocardia bacteria (Nocardia sp.) WSH-A11 (being deposited in Wuhan China typical culture collection center, numbering CCTCC NO.M201005)
(2) zymotechnique:
Liquid fermentation medium is formed (gL
-1): sucrose 10, corn steep liquor 0.5, urea 0.5, KH
2PO
40.1 NaCl 0.1, MgSO
47H
2O 0.2
A. can prepare biological flocculant REA-11 with sucrose during for carbon source with greater activity;
B. corn steep liquor and urea are composite can prepare the biological flocculant REA-11 with greater activity during for nitrogenous source;
C. liquid fermentation condition is 25～30 ℃ (best 28 ℃), the initial pH4.0 of substratum～10.0 (best 6.0), 120～150 rev/mins of (the suitableeest 120 rev/mins) shaking culture of shaking speed 48 hours;
(3) extraction of biological flocculant REA-11:
D. fermented liquid is centrifugal, obtains centrifuged supernatant;
E. in the centrifuged supernatant that (d) obtains, add the long-pending ethanol of triploid, centrifugal, obtain throw out;
F. the precipitation that obtains of dissolved in distilled water (e) is utilized treatment on ion exchange columns, obtains active eluant;
G. collect the active eluant that (f) obtains, after concentrating, utilize gel column to handle, obtain gel column and handle active eluant;
H. collect the gel column processing active eluant that (g) obtains, with ethanol sedimentation and washing;
I. the precipitation that obtains of dissolved in distilled water (h) utilizes preparation high pressure liquid chromatography post to make further refinement treatment, promptly gets product REA-11;
(4) component of biological flocculant is identified:
J. utilize high-pressure liquid phase, gas-chromatography, mass spectrum etc. to analyze means of testing, in conjunction with the feature chemical reaction, REA-11 forms evaluation to product.
2, by the biological flocculant of the described preparation of claim 1, it is characterized by Nocardia sp.WSH-A11, in the substratum that with 1% sucrose is carbon source, can prepare biological flocculant with higher flocculation activity.
3, by the biological flocculant of the described preparation of claim 1, it is characterized by Nocardia sp.WSH-A11, can prepare biological flocculant during with 1: 1 (mass ratio) composite nitrogenous source as fermention medium with higher flocculation activity at corn steep liquor and urea.
4, by the biological flocculant of the described preparation of claim 1, be purifying biological flocculation agent REA-11, it is characterized by described centrifugal condition is 6000～10000 rev/mins, obtains supernatant liquor in centrifugal 20 minutes.
5, by the biological flocculant of claim 1 and 4 described preparations, be purifying biological flocculation agent REA-11, it is characterized by and in centrifuged supernatant, add 3 times of volume of ethanol, 4 ℃ leave standstill 6h after, 7000 rev/mins centrifugal 10 minutes, obtain throw out.
6, press the biological flocculant of claim 1 and 5 described preparations, be purifying biological flocculation agent REA-11, after it is characterized by dissolved in distilled water ethanol sedimentation thing, (PEG-20000) is concentrated into 1/10 of original fermented solution volume with polyoxyethylene glycol, utilize the DEAE ion exchange column to carry out purifying, obtain active eluant.
7, by the biological flocculant of claim 1 and 6 described preparations, for purifying biological flocculation agent REA-11, it is characterized by the active eluant that obtains from the DEAE post, the gel column that passes through is Sephadex G-100.
8, press the biological flocculant of claim 1 and 7 described preparations, be purifying biological flocculation agent REA-11, it is characterized by the active eluant that obtains from Sephadex G-100, after ethanol sedimentation and washing, the preparation high pressure liquid chromatography post that passes through is Superdex 200 HR10/30.
9, press the biological flocculant of the described preparation of claim 1, for identifying the molecular composition of REA-11, the biological flocculant REA-11 that it is characterized by Nocardia sp.WSH-A11 preparation is a kind of acidic polysaccharose based on galacturonic acid, also contain the trace of albumin component in its molecule, assist to keep the flocculation activity of biomacromolecule.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CNB011080450A CN1139546C (en) | 2001-01-11 | 2001-01-11 | Biological flocculant prepared with Nocardia bacteria |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CNB011080450A CN1139546C (en) | 2001-01-11 | 2001-01-11 | Biological flocculant prepared with Nocardia bacteria |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| CN1322681A true CN1322681A (en) | 2001-11-21 |
| CN1139546C CN1139546C (en) | 2004-02-25 |
Family
ID=4656934
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CNB011080450A Expired - Fee Related CN1139546C (en) | 2001-01-11 | 2001-01-11 | Biological flocculant prepared with Nocardia bacteria |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN1139546C (en) |
Cited By (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN100334012C (en) * | 2003-11-13 | 2007-08-29 | 山东齐东方投资有限公司 | Method for preparing biological flocculant from arthrobacter |
| CN100420754C (en) * | 2006-10-26 | 2008-09-24 | 浙江海正药业股份有限公司 | Method for preparing anthracene Ensamu bacterial |
| CN102952761A (en) * | 2011-08-25 | 2013-03-06 | 天津工业生物技术研究所 | Nocardia sp. capable of converting quininone into (R)-3-quinuclidinol and conversion method |
| CN102952760A (en) * | 2011-08-25 | 2013-03-06 | 天津工业生物技术研究所 | Rhodococcus erythropolis capable of converting quininone into (S)-3-quinuclidinol and conversion method |
| CN109111041B (en) * | 2018-09-13 | 2021-05-07 | 福建海峡环保集团股份有限公司 | Method for inhibiting biological foams caused by Nocardia in aeration tank |
-
2001
- 2001-01-11 CN CNB011080450A patent/CN1139546C/en not_active Expired - Fee Related
Cited By (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN100334012C (en) * | 2003-11-13 | 2007-08-29 | 山东齐东方投资有限公司 | Method for preparing biological flocculant from arthrobacter |
| CN100420754C (en) * | 2006-10-26 | 2008-09-24 | 浙江海正药业股份有限公司 | Method for preparing anthracene Ensamu bacterial |
| CN102952761A (en) * | 2011-08-25 | 2013-03-06 | 天津工业生物技术研究所 | Nocardia sp. capable of converting quininone into (R)-3-quinuclidinol and conversion method |
| CN102952760A (en) * | 2011-08-25 | 2013-03-06 | 天津工业生物技术研究所 | Rhodococcus erythropolis capable of converting quininone into (S)-3-quinuclidinol and conversion method |
| CN109111041B (en) * | 2018-09-13 | 2021-05-07 | 福建海峡环保集团股份有限公司 | Method for inhibiting biological foams caused by Nocardia in aeration tank |
Also Published As
| Publication number | Publication date |
|---|---|
| CN1139546C (en) | 2004-02-25 |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| CN106635869B (en) | A method of producing surfactin using bacillus amyloliquefaciens | |
| CN102978134B (en) | Lactobacillus and method for producing D-lactic acid by fermenting using lactobacillus | |
| CN104342390B (en) | A kind of Sinorhizobium meliloti strain and combinations thereof and application | |
| Son et al. | Isolation and cultivation characteristics of Acetobacter xylinum KJ-1 producing bacterial cellulose in shaking cultures | |
| CN110373359B (en) | Streptomyces albus X-18 and method for producing epsilon-polylysine by using same | |
| CN101037661A (en) | Pseudoalteromonas and its usage | |
| CN102796673A (en) | Feruloyl esterase production strain and method for producing feruloyl esterase by using same | |
| US6255085B1 (en) | Production of chitosan and chitin | |
| CN117778479B (en) | Method for preparing 1, 2-propylene glycol by microbial fermentation method | |
| CN1101855C (en) | Liquid fermentation process for preparing both ganoderic polyose and ganoderic acid | |
| CN101113423A (en) | Microbacterium and method for producing chirality pharmaceutical intermediate compound by using the bacterial conversion | |
| CN1944626A (en) | Microbial bacteria for preparing ginkalide by microbial fermenting and method for preparing ginkalide | |
| Akponah et al. | Bio-ethanol production from cassava effluent using Zymomonas mobilis and Saccharomyces cerevisiae isolated from rafia palm (Elaesis guineesi) SAP | |
| CN1139546C (en) | Biological flocculant prepared with Nocardia bacteria | |
| CN107446868B (en) | One plant of Methylotrophic bacillus and its degradation of feather produce the application of oligopeptides | |
| CN101457250B (en) | Method for synthesizing betulic acid from betulin through microbial cell bioconversion | |
| CN1141392C (en) | Process for preparing ganoderic polyose and ganoderic acid by fermentation during which raw materials are supplemented | |
| CN1269421A (en) | Process for producing xylitol or D-xyluose | |
| CN116218690A (en) | Curvularia robusta producing cloth Lei Feide bacteria A and fermentation method thereof | |
| CN1355318A (en) | Process for preparing natural active dormin | |
| US6485946B1 (en) | Production of chitosan and chitin | |
| CN1027084C (en) | Process for preparing L-sorbose | |
| CN1033394A (en) | The method for making of amino deoxy mannitol | |
| KR20010083600A (en) | Process for Preparing Pullulan Employing Waste Materials Containing High Concentration of Sugars from Food-processing Industry | |
| KR900003740B1 (en) | New saccharomyces cere visiae strain and method for fermentation by its 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 | ||
| 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 |