CN100532541C - Method for preparing bacterial powder - Google Patents
Method for preparing bacterial powder Download PDFInfo
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- CN100532541C CN100532541C CNB2006101184629A CN200610118462A CN100532541C CN 100532541 C CN100532541 C CN 100532541C CN B2006101184629 A CNB2006101184629 A CN B2006101184629A CN 200610118462 A CN200610118462 A CN 200610118462A CN 100532541 C CN100532541 C CN 100532541C
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Abstract
The invention discloses a process for preparing strain powders, which includes a procedure of cultivating and fermenting strain to be spore, a concentrating procedure, an absorption procedure and a crashing and drying procedure. In the procedure of culturing and fermenting the strain to be spore, the fermentation is stopped at the time when the spore rate reaches more than 80% and 20% of the spore emerges out of a spore sac, in the concentrating procedure, fermentation liquor of the strain is in circle concentrated to be of 10% to 15% of the volume of the original liquor by employing hollow fiber film with diameter of 12 to 18 centimeters, length of 90 to 110 centimeters and molecular weight of 40,000 to 80,000, in the absorption procedure, the concentrated fermentation liquor is absorbed to be particles with diameter less than 5 to 6 millimeters (<5-6millimeters) by respectively utilizing absorbent such as light calcium carbonate and the like, in the absorption and drying procedure, the particles finishing absorption treatment are dried to be bacterial powders by hot air from a pneumatic dryer. The invention employs the hollow fiber film of which filtration pores are slightly smaller than that of bacteria and improves bacteria collecting rate, which is an economical and effective technique for producing the strain powders for microbial fertilizers.
Description
Technical field:
The present invention relates to the manufacture method of microorganism manure strain, specifically, the present invention relates to a kind of preparation method of strain powder.
Background technology:
Tradition microbial fertilizer manufacture method weak point is that all strain fermentating liquids only should be self-produced personal in the our factory, and is big through the strain liquid volume of fermentation culture, is difficult to packed and transported and becomes particle bacterial manure to other bacterial manure factories with local fertilizer raw material hybrid process; And bacterial manure enterprise in various places will produce this kind bacterial manure, certainly will all will set up strain plant fermentative production bacterial classification voluntarily.So, the facility investment and the technical force of the various places similar bio-feritlizer factory that then need be multiplied, and under the not enough situation that the fermentation technique personnel lack at business capital, originating as no bacterial classification certainly will have influence on the development of composite bio-fertilizer and apply.
Also have the high speed centrifugation concentration method of employing and flame filter press dehydration concentration method that bacterial classification is concentrated in the prior art, but the flame filter press filter opening is a lot of greatly than thalline, the collection rate of thalline is lower.
Summary of the invention:
The object of the present invention is to provide a kind of preparation method of strain powder, the described Preparation Method that duplicates has solved in the prior art gemma powder collection rate lower technical problem to bacterial classification.
The invention provides a kind of preparation method of strain powder, comprise a process that spawn culture is fermented into gemma, a spissated process, the process of an absorption, pulverizing and exsiccant process, in the described process that spawn culture is fermented into gemma, reach more than 80% and the gemma that accounts for overall number 20% stops fermentation when deviating from from sporangiocyst in the gemma rate, wherein, in described spissated process, utilizing diameter is 12～18cm, length is that the hollow-fibre membrane of 4 of 90～110cm～80,000 molecular weight is concentrated to 10%～15% of stoste volume with the circulation of the fermented liquid of bacterial classification, in the process of described absorption, utilize the fermented liquid after sorbent material will concentrate to adsorb into the particle of diameter less than 5～6mm, in described pulverizing and exsiccant process, utilize pneumatic drier to become water ratio smaller or equal to 8% with the particle drying of the hot blast under 120～160 ℃ of temperature after with adsorption treatment, fineness is less than 60 purpose bacterium powder.
Further, described sorbent material adopts light calcium carbonate.
Further, described bacterial classification is fixed nitrogen sporeformer CCTCC M95040.
Further, described bacterial classification is bacillus megaterium ACCC 10010.
Perhaps, described bacterial classification is bacillusmusilaginosiengineering ACCC 10013.
Further, in the process of absorption, sorbent material and concentrate after the weight percent of fermented liquid be 3:1.5～2.5.
Further, in the process of absorption, sorbent material and concentrate after the weight percent of fermented liquid be 3:2.
The vinelandii that the present invention adopts are meant bacillus azotobacter (Bacillus azotofixans), because of it contains nitrogenase, can be ammonia-state nitrogen with airborne nitrogen transformation, abbreviate " vinelandii " as.Vinelandii in China typical culture collection center (CCTCC) preservation, are numbered CCTCCM95040, and the public can buy phosphorus bacteria to this preservation center by preserving number.
The phosphorus bacteria that the present invention adopts is meant bacillus megaterium (Bacilllus megatherium) ACCC 10010, because of it can decompose the soil insoluble phosphorus, therefore is commonly called as and is phosphorus bacteria.Phosphorus bacteria in China Committee for Culture Collection of Microorganisms's agricultural microorganism center (ACCC) preservation, is numbered ACCC 10010, and the public can buy phosphorus bacteria to this preservation center by preserving number.
The potassium bacterium that the present invention adopts is bacillusmusilaginosiengineering (Bacilllus mucilaginosus) ACCC 10013, this bacterium can be decomposed the plain ability of potassium in the soil mineral because of it, therefore be commonly called as and be the potassium bacterium, the potassium bacterium is in China Committee for Culture Collection of Microorganisms's agricultural microorganism center (ACCC) preservation, be numbered ACCC 10013, the public can be by number asking for.
The present invention has improved the manufacture method of existing bacterial classification, and the strain fermentating liquid concentrate drying is become the dry strain powder, is condensed into to contain the high density gemma strain powder of (every gram contains 3,000,000,000 of gemma at least).But it is standby to concentrate the gemma strain powder long storage of being processed into: (1) is used as the bacterial classification of manufacturing particle/powdery bacterial manure certainly for the our factory; (2) use to various places bio-feritlizer factory processing and manufacturing bacterial manure through packed and transported, the similar enterprise in various places does not need to invest in addition strain plant.
The cultivation and fermentation of vinelandii can be used known culture condition and method.General incubation time is 20-24 hour, and culture temperature is 25-30 ℃, and cultivating pH is 7.0-7.4, air flow 0.5-1:1 cubic meter/1000 liters, stirring velocity 200-300 rev/min.The preferred incubation time of this bacterial classification is 28 hours formation gemma, and 30 ℃ of culture temperature are cultivated pH7.2, air flow 0.5-1:1 cubic meter/1000 liters, tank pressure 0.5kg/cm
2, 200 rev/mins of stirring velocitys.Its preferred culture medium is as follows:
Screening and preservation bacterial classification nitrogen-free agar:
Starch 0.8%
Peptone 0.15%
Yeast extract paste 0.01%
Ammonium sulfate ((NH
4)
2SO
4) 0.15%
Lime carbonate (CaCO
3) 0.15%
Dipotassium hydrogen phosphate (K
2HPO
4) 0.005%
Manganous sulfate (MnSO
4) 0.005%
Glucose 0.30%
Agar 2.5%
pH＝7.2-7.4
The substratum that is used for the shake-flask culture of CCTCC M95040 vinelandii and fermentative production is to add following composition on the basis of above-mentioned nitrogen-free agar in addition:
Sal epsom (MgSO
4) 0.005%
Potassium primary phosphate (KH
2PO
4) 0.02%
Sodium-chlor (NaCl) 0.01%
The cultivation and fermentation of phosphorus bacteria can be used known culture condition and method.
The substratum of a large amount of cultivation and fermentation of phosphorus bacteria is: starch 0.5%, analysis for soybean powder 0.075%, peptone 0.15%, ammonium sulfate 0.2%, sal epsom 0.005%, manganous sulfate 0.005%, dipotassium hydrogen phosphate 0.2%, yeast powder 0.1%, lime carbonate 0.01%, pH=7.2-7.4.All the other fermentation conditions are identical with the fixed nitrogen sporeformer.
The cultivation and fermentation of potassium bacterium can be used known culture condition and method.
The substratum of a large amount of cultivation and fermentation of potassium bacterium is: starch 1%, analysis for soybean powder 0.08%, dipotassium hydrogen phosphate 0.2%, ammonium sulfate 0.1%, manganous sulfate 0.005%, sal epsom 0.005%, iron trichloride 0.001%, lime carbonate 0.01%, yeast extract paste 0.01%, pH=7.2-7.4.All the other fermentation conditions are identical with fixed nitrogen sporeformer phosphorus bacteria.
Gemma bacterial classification concentrate drying becomes the powder technology
1, fermented liquid concentration
Vinelandii, phosphorus bacteria and the fermentation of potassium microbial culture 25-30 hour, through the gemma rate of formation of three kinds of thalline of microscopy, when the gemma rate reaches 80% or more and has 20% gemma can stop to ferment in deviating from from sporangiocyst.Then the fermented liquid of three kinds of bacterium is used respectively by diameter 12-18cm, the circulation of the 4-8 ten thousand molecular weight hollow-fibre membranes of long 90-110cm is concentrated to the 10%-15% of stoste volume.
2, concentrated solution adsorption treatment
Fermented liquid after concentrating is become the particle of diameter＜5-6mm respectively with adsorbents adsorb such as light calcium carbonates.
Join sorbent quantity in the fermented liquid according to following ratio proportioning:
Sorbent material: concentrated solution=3:1.5-2.5 (mass ratio)
3, hot blast pulverizing, drying treatment
Particle after the adsorption treatment is become water ratio≤8%, fineness＜60 purpose bacterium powder through pneumatic drier 120-160 ℃ warm air drying.
4, hybrid packed
Can package storage after dried sporeformer powder is mixed in proportion standby.
The gemma dry bacterial powder contains the bacterium number and detects
According to the living spores bacterium number in the dilution plate number scale detection dry bacterial powder of routine.Take by weighing dry bacterial powder 10 grams, join in 90 ml sterile waters and be diluted to proper concn, (every milli contains bacterium approximately counts 20-200), pipetting 0.1 milliliter of dilution bacterium liquid then joins in the aseptic cultivation first ware, add vinelandii or phosphorus bacteria or potassium bacteria culture medium after dissolving in right amount respectively, form flat board after the thorough mixing condensation, in 30 ℃ of insulation cans, cultivated two days, calculate the colony number on the culture dish flat board.The calculating of every kind of bacterium is dull and stereotyped to be repeated 3 times, gets the average colony number that grows on the numeration flat board, multiply by the viable count that the diluted sample multiple is to be had in every gram dry bacterial powder (being the living spores number), and its calculation formula is as follows:
Dry bacterial powder living spores number
(hundred million/gram)=on average each flat-plate bacterial colony number (individual) * diluted sample multiple (individual/gram)
The dry bacterial powder final product quality requires:
3 kinds of ripe total amounts of bacterium living spores: 〉=30 * 10
8(hundred million/gram) (wherein vinelandii, phosphorus bacteria, potassium bacterium respectively account for 1/3) dry bacterial powder water content:＜5%
Dry bacterial powder fineness: cross 80 mesh sieve holes more than 95%
The present invention compares with prior art, adopt tubular fibre membrane concentration after heat wind to be dried to powder method concentrated strain gemma, more save the energy and cost and rate of recovery height than high speed centrifugation concentration method, (the flame filter press filter opening is a lot of greatly than thalline than flame filter press dehydration concentration method bacterial classification enrichment factor height, and the hollow-fibre membrane filter opening that the present invention uses is smaller than thalline, improved the microorganism collection rate), be that a kind of most economical effective microorganism manure strain powder is made new technology through facts have proved.
Embodiment:
Embodiment 1 (vinelandii gemma powder)
1, fixed nitrogen bacterial classification cultivation and fermentation
Vinelandii sporeformer CCTCC M95040 is inoculated into the triangular flask shake-flask culture 18 hours that fill 1000ml foregoing is used for vinelandii substratum of the present invention, be inoculated in 300 liters of seed fermentation jars by 1% inoculum size then, ventilation (1:0.5), stir, 30 ℃ of heat insulating culture 15 hours, be inoculated in 5000 liters of big fermentor tanks by 3% inoculum size again, similarity condition cultivate be warmed up to after 20 hours 35 ℃ and strengthen air flow (1:1) cultivate again treated in 10 hours thalline gemma rate of formation reach 80% and have 20% gemma from sporangiocyst, to deviate from after can go out jar and become and produce bud fixed nitrogen strain fermentating liquid.
2, the preparation of gemma bacterial classification concentrated dry powder
The vinelandii fermented liquid that forms gemma is put into hold tank, through diameter 15cm, the 50000 molecular weight hollow-fibre membranes of long 100cm carry out concentration, the solution to be concentrated volume is reduced to about stoste 10%-15%, the light calcium carbonate that adds concentrated solution quality 150% adsorbs into the particle of diameter＜5-6mm, then the particle after the adsorption treatment is become water ratio≤8% through the warm air drying of 130 ℃ of pneumatic driers, fineness＜60 purpose bacterium powder, can package storage standby.The bacterial classification dry powder that is prepared into by this method should contain at least 30 hundred million of gemma numbers/gram, and dry storage active maintenance in 2 years is more than 80%.
The strain powder of making by the present invention can become particle bacterial manure or uses as the seed dressing of farm crop with strain powder for our factory or other factory with fertilizer matrix hybrid process.
3 become particle bacterial manure with fertilizer matrix hybrid process
The municipal sludge and the chicken manure that will become thoroughly decomposed are mixed in proportion pulverizing, cross 80 mesh sieves, add 10% ammonium sulfate then and are mixed into fertilizer matrix.Add the 1% vinelandii gemma powder thorough mixing made of step as described above in above-mentioned fertilizer matrix, spray adds 20% moisture and rolls and make diameter 3-5mm spherical granules in pan-pelletizer, promptly makes the particulate state azotobacteria fertilizer through 80 ℃ of hot blast roller dryings again.This product contains fixed nitrogen sporeformer at least 0.3 hundred million/gram, and dry storage active maintenance in 2 years is more than 80%.
Embodiment 2 (phosphorus bacteria gemma powder)
Cultivate the phosphorus bacteria fermented liquid by aforementioned phosphorus bacteria substratum and embodiment 1 described method, again through make the phosphorus bacteria gemma powder that concentrates processing with method tubular fibre membrane concentration and warm air drying.At least 30 hundred million/gram of the phosphorous bacterial spore of the phosphorus bacteria strain powder of making, can make bacterial classification for our factory or nonlocal bacterial manure factory, as described in embodiment 1, join fertilizer matrix in 1% ratio and promptly can be made into granular phosphorus bacterium fertilizer, every gram particle bacterial manure contains at least 0.3 hundred million of phosphorus bacteria.
Embodiment 3 (potassium bacterial spore powder)
Cultivate the potassium ferment product by aforementioned potassium bacteria culture medium and embodiment 1 described method, again through make the potassium bacterial spore powder that concentrates processing with method tubular fibre membrane concentration and spraying drying.The potassium bacteria culture powder of making contains at least 30 hundred million of potassium bacterial spores/gram, can make bacterial classification for our factory or nonlocal bacterial manure factory, method described in embodiment 1-3, join mixing granulation in the fertilizer matrix in 01% ratio, promptly can be made into particle potassium bacterium fertilizer, every gram finished product contains at least 0.3 hundred million on potassium bacterium.
Embodiment 4 (composite bacteria gemma powder)
The sporeformer powder of making by method described in previous example 1, example 2 and the example 3 is mixed in proportion, and promptly makes to contain vinelandii, phosphorus bacteria and potassium bacterium three bacterium blended sporeformer powder.Three kinds of bacterium compound gemma powder contain at least 30 hundred million of gemma/gram.
As execute as described in the routine 1-example 3, in fertilizer matrix, add 0.5-1% composite bacteria gemma powder and promptly can be made into the organic-inorganic microbial compound bacterial fertilizer granulated fertilizer material that contains vinelandii, phosphorus bacteria and potassium bacterium.This kind product contains at least 0.3 hundred million of three kinds of composite bacteria/gram, meets that the strong composite microbiological fertilizer of particle contains effective bacterium requirement in the speck fertilizer industry standard (NY227-94) that present China Ministry of Agriculture sends out.
The composite microbiological fertilizer that the present invention makes has carried out a large amount of tests through the land-reclaimable economic center of China from 2003 on various crop such as the rubber in reclamation areas such as Yunnan, Hainan, Guangdong, sugarcane, tealeaves, tropical fruit, the result shows that this fertilizer makes crop contrast volume increase 4.8-21.8%, can significantly improve crop quality, and the reclamation area soil of fostering and apply fertilizer is had promoter action.
Claims (7)
1. the preparation method of a strain powder comprises that a process, that spawn culture is fermented into the process of gemma, a spissated process, an absorption pulverizes and the exsiccant process; In the described process that spawn culture is fermented into gemma, reach more than 80% and the gemma that accounts for overall number 20% stops fermentation when deviating from from sporangiocyst in the gemma rate; It is characterized in that: in described spissated process, utilizing diameter is that 12～18cm, length are that the hollow-fibre membrane of 4 of 90～110cm～80,000 molecular weight is concentrated to 10%～15% of stoste volume with the circulation of the fermented liquid of bacterial classification; In the process of described absorption, utilize the fermented liquid after sorbent material will concentrate to adsorb into the particle of diameter less than 5～6mm; In described pulverizing and exsiccant process, utilize pneumatic drier with the particle drying of the hot blast under 120～160 ℃ of temperature after with adsorption treatment become water ratio smaller or equal to 8%, fineness contains the bacterium powder of 3,000,000,000 of gemma at least less than 60 orders, every gram.
2. the preparation method of strain powder as claimed in claim 1 is characterized in that: described sorbent material employing light calcium carbonate.
3. the preparation method of strain powder as claimed in claim 1, it is characterized in that: described bacterial classification is fixed nitrogen sporeformer (Bacillus azotofixans) CCTCC M95040.
4. the preparation method of strain powder as claimed in claim 1, it is characterized in that: described bacterial classification is bacillus megaterium (Bacillus megatherium) ACCC 10010.
5. the preparation method of strain powder as claimed in claim 1, it is characterized in that: described bacterial classification is bacillusmusilaginosiengineering (Bacillus mucilaginosus) ACCC 10013.
6. the preparation method of strain powder as claimed in claim 1 is characterized in that: in the process of absorption, sorbent material and concentrate after the weight percent of fermented liquid be 3: 1.5～2.5.
7. the preparation method of strain powder as claimed in claim 6 is characterized in that: in the process of absorption, sorbent material and concentrate after the weight percent of fermented liquid be 3: 2.
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|---|---|---|---|
| CNB2006101184629A CN100532541C (en) | 2006-11-17 | 2006-11-17 | Method for preparing bacterial powder |
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| CN105255785A (en) * | 2015-11-17 | 2016-01-20 | 山东省农业科学院农业资源与环境研究所 | Fermentation method of bacillus megatherium with high rate of sporation |
| CN109496683B (en) * | 2018-11-30 | 2021-07-30 | 宜章县福城生物科技开发有限公司 | Method for producing homogeneous dispersion type edible mushroom strain |
| CN113186184B (en) * | 2021-04-17 | 2022-11-01 | 沃地丰生物肥料科技(山东)股份有限公司 | Production process of general bacillus concentrated strain |
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Non-Patent Citations (2)
| Title |
|---|
| 中空纤维膜过滤法高密度培养凝结芽孢杆菌TQ33. 戚薇等.食品与发酵工业，第29卷第4期. 2003 |
| 中空纤维膜过滤法高密度培养凝结芽孢杆菌TQ33. 戚薇等.食品与发酵工业，第29卷第4期. 2003 * |
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Address after: 410005 Changsha, Wuyi Road, Hunan, No. 2, First Avenue, room 909 Patentee after: Hunan Yu Garden biological Polytron Technologies Inc Address before: 410001 Hunan province Changsha Furong district near east Kam Business Building Room 309 Patentee before: Hunan Yuyuan Bio-technological Co., Ltd. |