CN1236065C - Production of flavicidic acid by microorgan degradation - Google Patents
Production of flavicidic acid by microorgan degradation Download PDFInfo
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- CN1236065C CN1236065C CN 02158189 CN02158189A CN1236065C CN 1236065 C CN1236065 C CN 1236065C CN 02158189 CN02158189 CN 02158189 CN 02158189 A CN02158189 A CN 02158189A CN 1236065 C CN1236065 C CN 1236065C
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- coal
- natural plant
- humic acid
- fulvic acid
- yellow humic
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- 241000194110 Bacillus sp. (in: Bacteria) Species 0.000 claims abstract description 11
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- QJZYHAIUNVAGQP-UHFFFAOYSA-N 3-nitrobicyclo[2.2.1]hept-5-ene-2,3-dicarboxylic acid Chemical compound C1C2C=CC1C(C(=O)O)C2(C(O)=O)[N+]([O-])=O QJZYHAIUNVAGQP-UHFFFAOYSA-N 0.000 abstract description 10
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- Fertilizers (AREA)
- Micro-Organisms Or Cultivation Processes Thereof (AREA)
Abstract
The present invention relates to a method for degrading low-energy coal from microorganisms to produce yellow humic acid. Raw material coal and an auxiliary material are mixed and added into a degradation strain, and the coal is fermented to obtain the yellow humic acid; the auxiliary material comprises a natural plant carbon source, a natural plant protein source, urea and KH2PO4, and the degradation bacteria include penicillium sp. LNP6CGMCC0866 and/or bacillus sp. BJB2CGMCC0867. In the method, the yellow humic acid can be obtained from various sources of lignite, efflorescent coal, coal gangue, etc., and compared with a chemical method, the yield of the yellow humic acid is obviously enhanced; the obtained yellow humic acid has strong activity, molecules of the obtained yellow humic acid are obviously minified because of the biological degradation, the content of oxygen and nitrogen is increased, and the limit of flocculation is large; chemical solvents are not used in the process of fermentation and extraction, secondary wastes polluting the environment can not be generated, agriculture leftovers are used as the auxiliary material, residues can be used as organic fertilizer, and the sustainable use of energy resources can be realized.
Description
Technical field
The present invention relates to a kind of method of utilizing the low-yield coal of microbiological deterioration to produce xanthohumic acid.
Background technology:
Xanthohumic acid is molecular weight less (it is generally acknowledged less than 500D) in the humic acids, active high component, aspect such as can be widely used in worker, farming, cure, herd, as improve the plant drought ability, as chemical fertilizer, pesticide synergistic agent, plant growth stimulator, medicine industry raw material etc.
Chemical process is all adopted in xanthohumic acid production at present, mainly extracts from weathered coal:
(1) uses the sulfuric acid acetone method
Weathered coal behind the wet concentration and aqueous acetone are mixed, make it to suspend, add the sulfuric acid stirring reaction again through stirring.To react supernatant liquor after the natural subsidence and remove solvent, obtain the xanthohumic acid product.Because of weathered coal xanthohumic acid content is low, must xanthohumic acid content be brought up to more than 38% through wet concentration; For the pH that keeps extracting solution about 1.5, consume a large amount of sulfuric acid, and production equipment requires acidproof; Solvent is an acetone, and is volatile during recovery, and reaction vessel requires airtight.
(2) ion-exchange-resin process
The metal ion that replaces in the Xanthohumate with strong acid ion exchange resin discharges xanthohumic acid.
This method requires xanthohumic acid content＞30% in weathered coal, and granularity＜80 orders fully reacting with resin, and is separated with ion exchange resin by screening; The granularity that also requires resin is between the 30-60 order, and consumption is big; Needing with relatively large concentration during regeneration is about 8% hydrochloric acid that enter the mouth; Also to use soft water during reaction; Will feed pressurized air during reaction and stir, energy consumption is big; Ash content from coal dust is arranged in the xanthohumic acid that obtains, will keep higher temperature (70-80 ℃) during de-ash.
There is abundant lignite resource in China.Brown coal incendiary thermo-efficiency is low, and causes environmental pollution easily, therefore extracts xanthohumic acid highly significant from brown coal.Prior art is to extract from brown coal with chemical means.Method is: extract with alkali lye earlier, then with the supernatant liquor acidifying, be settled out xanthohumic acid wherein.But this method extraction yield is low, and no industrial production is worth.And the xanthohumic acid flocculation limit that chemical method is produced is low, and is active low, and produces the secondary refuse of contaminate environment,
Summary of the invention
The problem to be solved in the present invention is with the microbial method low-yield coal of degrading, and obtains xanthohumic acid.
The invention provides a kind of accessory formula mixes with feed coal, add the efficient degrading bacterial strain of seed selection of the present invention, coal is fermented, the xanthohumic acid that is combined in the coal is discharged, become the component of water soluble, and can make part macromole humic acids become the small molecules xanthohumic acid.
Concrete grammar is as follows:
1, batching:
Feed coal is mixed with auxiliary material, add entry and stir;
2, fermentation:
Insert Penicillium notatum Penicillium sp.LNP6 CGMCC 0866 or bacillus sp.BJB2 CGMCC 0867 in above-mentioned batching, inoculum size is l0%, and solid koji unit is W/W, and liquid culture is V/W, ferments 7-9 days; Or in above-mentioned batching, inserting Penicillium notatum Penicillium sp.LNP6 CGMCC0866, inoculum size is 5-10%, cultivates and inserts bacillus sp.BJB2 CGMCC0867 after 2 days again, inoculum size is 5-10%, solid koji unit is W/W, and liquid culture is V/W, ferments 6-7 days.
3, extract:
Add entry, separate and contain the xanthohumic acid supernatant.
This supernatant can directly dilute utilization, will can obtain the xanthohumic acid powder after this supernatant drying; Residue can further extract or directly be used as humic acid fertilizer.
Described feed coal comprises brown coal, weathered coal and coal gangue;
Described auxiliary material comprise natural phant carbon source from agricultural byproduct (as in sawdust, rice bran, the straw (wheat, corn or rice straw etc.) one or more) and natural plant protein source (as soya-bean cake, peanut cake), and urea, KH
2PO
4
Feed coal and auxiliary material parts by weight proportioning are: coal: auxiliary material (natural phant carbon source): auxiliary material (natural plant protein source): urea: KH
2PO=40-60: 20-40: 10-20: 2: 0.1
Described degradation bacteria is Penicillium notatum Penicillium sp.LNP6 CGMCC 0866 and/or bacillus sp.BJB2 CGMCC 0867
Preferred fermentation process is to plant Penicillium notatum Penicillium sp.LNP6 CGMCC0866 in batching earlier, inoculum size is 5-10%, cultivate and plant bacillus sp.BJB2 CGMCC0867 after 2 days again, inoculum size is 5-10%, fermentation to 6～7 day, solid koji unit is W/W, and liquid culture is V/W;
The present invention's beneficial effect against existing technologies is:
1, can obtain xanthohumic acid from multiple sources such as brown coal, weathered coal and coal gangues, and the rate ratio chemical method improves obviously;
2, the gained xanthohumic acid is active strong, because biological Degradation, gained xanthohumic acid molecule obviously diminishes, and oxygen and nitrogen content increase, and flocculation limit is big;
3, fermentation, leaching process do not use chemical solvents, can not produce the secondary refuse of contaminate environment;
4, auxiliary material is an agricultural byproduct, and residue can be made fertilizer and use, and can realize the sustainable use of the energy.
The explanation of related microorganism material
1, microbial name
(1) Penicillium notatum
Be Penicillium (Penicillium sp.), bacterial strain called after: Penicillium sp.LNP6
(2) genus bacillus
Be bacillus (Bacillus sp.), bacterial strain called after: Bacillus sp.BJB2
2, microbial preservation number
(1) Penicillium notatum Penicillium sp.LNP6 CGMCC 0866
(2) bacillus sp.BJB2 CGMCC 0867
3, microorganism mycology feature
(1) morphological specificity
Penicillium notatum:
Bacterium colony is round, bacterium colony is surperficial is fine and close fine and soft shape, is seeded on the czapek's solution, can form obvious bacterium colony in one day, cultivates a few days ago that bacterium colony is white in color, and the 3rd day begins to become yellow, and the substratum back side also produces tangible yellow soluble pigment.Along with the prolongation of incubation time, yellow is deepened gradually, after seven days, produces yellow to the safran conidium.
The inserted sheet culture method is observed and is found that the conidiophore of this bacterium is tangible single-wheel broom shape branch, and the conidium ellipse is so be defined as Penicillium.
Genus bacillus:
The bacterium colony circle, white, refractive index is strong.Gram-positive, the thalline size is 1-1.5 μ m * 4.0-5.0 μ m, cultivates on bacteria culture medium, and beginning in 16 hours can produce gemma, and gemma is middle giving birth to.
So be defined as genus bacillus.
(2) spawn culture separation method
Penicillium notatum separates from coal washery, colliery, Fuxin, Liaoning, and genus bacillus separates in the coal gangue of Mentougou, is normal temperature type bacterium (the mould growth temperature is 25-28 ℃, and genus bacillus is 28-30 ℃).Mould can be with general liquid fermenting or solid koji enlarged culturing, and genus bacillus is cultivated with general bacterial liquid fermentation method and gets final product.
The strain separating method
Sampling is made bacteria suspension with 8.5% salt solution, is that the minimum medium of sole carbon source is coated with separation in order to brown coal, waits to grow picking dominant bacteria behind single bacterium colony, and further checks the ability of its brown coal of degrading.Method be with inoculation on solid plate, 28-30 ℃ of cultivation, (mould three days behind the formation bacterium colony, genus bacillus 1 day) on bacterium colony, adds through (121 ℃ of high-temperature sterilizations, brown coal 30min), continue to cultivate observe brown coal and be degraded and form the situation of black drop, speed that occurs with drop and formation amount are as screening index.
Find that through screening a strain mould can form obvious drop in three days, a bacillus can obviously form drop in one day.Penicillium notatum is carried out ultraviolet mutagenesis twice, obtain a plant mutant strain, be numbered P6 (Penicillium sp.).The ability of this mutant strain degraded brown coal obviously improves, and beginning degradable brown coal generated the black drop in 36 hours, and its cell-free filtrate of cultivating through 3 days shaking tables brown coal of also degrading produce the black soluble material.
4, effect
This two strains bacterium has the effect of stronger degraded brown coal.Dull and stereotyped this mould of cultivating can will place the Inner Mongol brown coal degraded on its bacterium colony to produce the black drop since 36 hours.Genus bacillus can begin to place the brown coal degraded on its bacterium colony to produce the black soluble material at 20 hours, and atrament infiltration substratum, and tangible black is dyed at the substratum back side.
With these two strains bacterium mixed fermentation degraded Inner Mongol brown coal, water-soluble xanthohumic acid content reaches more than 30% in the tunning.
Embodiment
Embodiment 1
1) batching
45% brown coal+14% sawdust+14% soya-bean cake+25% straw (corn or rice straw)+2% urea+0.1%KH
2PO
4
Brown coal (from Huolin Gol colliery, Inner Mongol) are crushed to about 50 orders, and other ingredient powders are broken to about 2mm;
2) fermentation
Various material are mixed, stir, add water (urea and the KH of 2 times of weight
2PO
4Dissolve in the water and add), leave standstill after 30 minutes and stir, insert Penicillium notatum (solid koji or liquid culture: W/W or V/W) with 10% inoculum size, cultivated 2 days for 28 ℃-30 ℃, once since stirring in the 12nd hour per 5～6 hours.Insert sporeformer (inoculum size is the same) after 2 days again, standing for fermentation to 6～7 day.
3) extraction step: fermentation finishes, by expecting (doing): water=1: 20 adding tap water, room temperature (or keeping 60 ℃) two hours, intermittently stir, sedimentation two hours separates supernatant, and residue adds the equal-volume flooding once again, merge twice supernatant and be xanthohumic acid, can drying obtain xanthohumic acid powder or directly dilution utilization.If can extract (xanthohumic acid) 30-35%, if can extract (xanthohumic acid) 40-44% at 60 ℃ with room temperature (about 20 ℃).Humic acid fertilizer is further extracted or directly be used as to the residue available bases.
4) to the evaluation and the specificity analysis of product xanthohumic acid
1919, Oden at first proposed humic acids is divided into water miscible xanthohumic acid, soluble ulmic acid of ethanol and the soluble Ulmic acids of alkali.1984, Xue Hongji etc. pointed out: xanthohumic acid is that alkali is solvable, the solvable and water soluble of acid humic acid substance.1996, Wang Tianli etc. discussed with regard to the condition that defines of xanthohumic acid, and defining of xanthohumic acid of proposition should be with water-soluble humic acids as unique conditional.When being equipped with xanthohumic acid, limit the pH value (2) after the acidifying with the alkali-soluble acid analysis legal system.
Leavened prod is directly to use water extraction in this research, so be the xanthohumic acid product.Because auxiliary material is an organism, measures in the product and contain 5-6% amino acid, 3-5% nucleic acid.And gained xanthohumic acid product carried out following specificity analysis: Infrared spectroscopy shows: this product is at 1720cm
-1, 1620cm
-1, 1400cm
-1, 1250cm
-1There is absorption peak at the place, conforms to the absorption spectrum of Gongxian (now being the Gong Yi city) xanthohumic acid.All solvable under diluted acid (0.01mol/L HCl), neutrality and diluted alkaline (0.01mol/L NaOH) condition.So show it is xanthohumic acid.
5) strain separating method
From the sampling of limit, ditch, coal washery, colliery, Fuxin, Liaoning, make bacteria suspension with 8.5% salt solution, in order to brown coal the minimum medium (K of sole carbon source
2HPO
40.5, KH
2PO
40.5, MgSO
4.2H
2O 0.5, (NH
4)
2SO
41, CaCl
2.2H
2O 0.1, brown coal 2g, H
2O 1L) is coated with separation, waits to grow picking dominant bacteria behind single bacterium colony, and further check the ability of its degraded brown coal.Method be with inoculation on solid plate, 28-30 ℃ of cultivation, (mould three days behind the formation bacterium colony, genus bacillus 1 day) on bacterium colony, adds through (121 ℃ of high-temperature sterilizations, brown coal 30min), continue to cultivate observe brown coal and be degraded and form the situation of black drop, speed that occurs with drop and formation amount are as screening index.
Find that through screening a strain mould can form obvious drop in three days, a bacillus can obviously form drop in one day.Penicillium notatum is carried out ultraviolet mutagenesis twice, obtain a plant mutant strain, be numbered P6 (Penicillium sp.).The ability of this mutant strain degraded brown coal obviously improves, and beginning degradable brown coal generated the black drop in 36 hours, and its cell-free filtrate of cultivating through 3 days shaking tables brown coal of also degrading produce the black soluble material.
6) strain identification
According to colony characteristics, mycelia have every and [determine that P6 is mould (Penicilliumsp.); According to bacterium colony, thalli morphology and can produce features such as gemma and determine that bacterium is genus bacillus (Bacillus sp).
Embodiment 2
1) batching
56% brown coal (from the Sanmenxia Gorge, Henan)+20% soya-bean cake+12% rice bran+10% sawdust+2% urea+0.1%KH
2PO
4
2) operation steps
Various material are mixed, stir, and (moistening to expect, loose is principle, urea and KH to add suitable quantity of water
2PO
4Dissolve in the water and add), leave standstill after 30 minutes and stir, insert Penicillium notatum (solid koji or liquid culture: W/W or V/W) with 10% inoculum size, cultivated 2 days for 28 ℃～30 ℃, once since stirring in the 12nd hour per 5～6 hours.Insert sporeformer (inoculum size is the same) after 2 days again, standing for fermentation to 6～7 day.Fermenting finishes extracts xanthohumic acid 18-25% with method described in the embodiment 1 at 20 ℃, and the residue water further extracts or directly be used as humic acid fertilizer.
The evaluation of strain separating cultural method and product xanthohumic acid and characteristic analysis method are with embodiment 1.
Claims (5)
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN 02158189 CN1236065C (en) | 2002-12-24 | 2002-12-24 | Production of flavicidic acid by microorgan degradation |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN 02158189 CN1236065C (en) | 2002-12-24 | 2002-12-24 | Production of flavicidic acid by microorgan degradation |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| CN1510142A CN1510142A (en) | 2004-07-07 |
| CN1236065C true CN1236065C (en) | 2006-01-11 |
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| Application Number | Title | Priority Date | Filing Date |
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| CN 02158189 Expired - Fee Related CN1236065C (en) | 2002-12-24 | 2002-12-24 | Production of flavicidic acid by microorgan degradation |
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Families Citing this family (10)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101913950A (en) * | 2010-08-19 | 2010-12-15 | 范鸿基 | Organic fertilizer preparation method and multi-strain breeding of biodegradable coal gangue |
| CN103588977B (en) * | 2013-11-08 | 2015-10-28 | 云南联合药业有限责任公司 | Method and the medicinal application of the yellow humic acid of peat are prepared in a kind of extraction |
| CN103724060B (en) * | 2013-12-31 | 2015-09-09 | 新疆科林思德新能源有限责任公司 | A kind of brown coal bio-fertilizer and production method thereof |
| CN107815433B (en) * | 2017-12-08 | 2021-03-12 | 青岛益佰农肥业有限公司 | Compound microbial agent and application thereof in producing fulvic acid by fermenting humic acid lignite |
| CN108033860A (en) * | 2018-01-09 | 2018-05-15 | 遵义粒满丰肥业有限责任公司 | A kind of method of multiplicity reactivation weathered coal humic acid |
| CN109006364A (en) * | 2018-10-10 | 2018-12-18 | 中国科学院沈阳应用生态研究所 | A kind of planting lawn matrix and preparation method thereof |
| CN110747132B (en) * | 2019-10-31 | 2021-03-23 | 内蒙古科技大学 | White paecilomyces and application thereof in preparation of phthalic acid ester by dissolving lignite |
| CN110999753B (en) * | 2019-12-26 | 2021-07-27 | 北京林业大学 | A kind of artificial ecological substrate based on coal gangue and preparation method thereof |
| CN110951626B (en) * | 2019-12-27 | 2021-05-25 | 北京林业大学 | A strain of Penicillium oxalate that decomposes coal gangue |
| CN116589700A (en) * | 2023-05-19 | 2023-08-15 | 内蒙古蒙土生态环保科技有限公司 | Method for purifying fulvic acid |
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| CN1510142A (en) | 2004-07-07 |
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