CN102326668A - Method for anaerobically degrading feather keratin with microbial strain 18D-TA - Google Patents
Method for anaerobically degrading feather keratin with microbial strain 18D-TA Download PDFInfo
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- CN102326668A CN102326668A CN201110245892A CN201110245892A CN102326668A CN 102326668 A CN102326668 A CN 102326668A CN 201110245892 A CN201110245892 A CN 201110245892A CN 201110245892 A CN201110245892 A CN 201110245892A CN 102326668 A CN102326668 A CN 102326668A
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Abstract
The invention relates to a method for anaerobically degrading feather keratin with a microbial strain 18D-TA, belonging to the technical field of microorganisms. The method comprises the following specific steps of: A, treating feathers: cleaning feathers with water, putting into a drying oven of which the temperature is 45-55 DEG C, and drying for 2-3 days for later use; and B, inoculating the strain: performing high-temperature moist-heat sterilization on a culture medium and the feathers treated in the step A, inoculating the strain 18D-TA in the amount of 2-10 percent, and anaerobically standing and culturing at the temperature of 45-60 DEG C for 8-10 hours till feather down falls off, wherein the feathers are completely separated from feather stems 18-24 hours later, and the feather stems are degraded in different degrees. Compared with other aerobic bacteria, the strain used in the method has the advantages: more products can be accumulated, so that ventilation equipment is not required to be provided, and the energy consumption is lowered greatly; and meanwhile, the efficiency is high, the fermenting period is short, racemization of most amino acids is avoided, and the absorption applicability of a feather hydrolysis product is enhanced.
Description
Technical field
The present invention relates to a kind of biodegrading process of feather keratin, particularly a kind of method of utilizing microbial strains 18D-TA anaerobic degradation feather keratin belongs to microbial technology field.
Background technology
At present; Technology with the feather keratin resource conversion mainly is to adopt the physicochemical method big, that be of low nutritive value of polluting, consume energy; Biological method is high because of the keratinase production cost, the not high reason of degradation of feather by using microorganism efficient, the universal utilization of influence technique.Patent application like application number 200810018317.2 discloses a kind of method with processing livestock hair keratin with ultrasonic, comprises that reductive water solution or oxydrolysis legal system are equipped with raw liquid of ceratin, ultrasonic Treatment, drying process step.This method needs reductive hydrolysis or oxydrolysis that the livestock and poultry feather keratin is processed albumen stoste, consumes a large amount of chemical reagent, possibly cause bigger pollution to environment; In addition, albumen stoste also needs ultrasonic wave further to handle could can arrive keratin liquid, has not only increased equipment cost, makes that the operating procedure complicacy of recycling of livestock and poultry feather keratin is loaded down with trivial details.Application number 200710171939.4 relates to a kind of bacterial strain and application thereof of producing keratinase; This bacterial strain is to have a liking for maltose oligotrophy monad (Stenotrophomonas maltophilia) DHHJ CGMCC No.2231, is applied to degradation of soft keratin chicken feather, pigeon hair, ox hair, wool, hair and hard keratin ox horn, goat's horn, sheep hoof angle, nail discarded object.This techniques make use is had a liking for maltose oligotrophy monad (Stenotrophomonas maltophilia) DHHJ needs 48h-96h at 40 ℃ of degraded feather keratins long period; This increases the time cost of industrial development probably; In addition; This Pseudomonas needs bigger aeration equipment in the aerobic bacteria feather keratin of degrading, and has improved fermentation costs; Be prone to cause microbiological contamination during ventilation.The present invention is through utilizing efficient anaerobic feather degradation bacteria 18D-TA; Form the anaerobic fermentation technology that efficient feather waste material transforms; Thereby making the microbial method degradation of feather by using produce this technology of useful resource is used widely; For the supply of China's feedstuff industry provides strong technical support, further improve China's feed product competitiveness in the world.
Summary of the invention
The present invention is intended to the defective to prior art, and a kind of method of utilizing microbial strains 18D-TA anaerobic degradation feather keratin is provided.This microbial strains can not only be handled insoluble structural proteins, the albumen of all right processing soluble, and this method under anaerobic can make feather that very high degradation efficiency is arranged, and fermentation period is short.
The technical scheme that the present invention adopts is:
Utilize the method for microbial strains 18D-TA anaerobic degradation feather keratin, it is characterized in that concrete steps are following:
The processing of A, feather
Feather washing is put into 45 ℃ of-55 ℃ of baking ovens after clean, dry 2-3 days subsequent use;
The inoculation of B, bacterial strain
1) preparation culture medium: add 500ml distilled water in advance in the triangular flask of 1L, add K more respectively
2HPO
43 g/L, KH
2PO
42.5 g/L, NaCl 3 g/L, MgSO
47H
2O 0.02 g/L, CaCO
30. 02 g/L, vitamin liquid 1% (v/v), liquid microelement 1% (v/v) adds water to 1L then to its dissolving, boils 5-8min, is cooled to room temperature, adds 0.1% (w/v) HClH
2O regulates pH to 7.0-7.5 with 5mol/l KOH, and adding mass and size concentration again is 0.1% resazurin solution 0.1% (v/v), presses 2.0m
3/ h leads to N
2, boil 15min, cooling back is divided and is filled in the anaerobism pipe or serum bottle that is added with 1% (w/v) feather after processing of step A in advance, and is subsequent use behind 121 ℃, 30min high-temperature heat sterilization.
The NaHCO of aseptic anaerobic
3Solution and Na
2S solution: get in the triangular flask that 500ml distilled water is added on 1L, boil about 10min, under Hungate copper post deaerating type of cycles anaerobic device, logical N
2(2.0m
3/ h), boil 20min, after the cooling, get 70ml moisture and be loaded on and be added with NaHCO in advance
3(10%--w/v) and Na
2In the serum bottle of S (3%--w/v), treat the good interior plug of whole dissolvings back plug, add the aluminium envelope.Add about 120ml N respectively with every bottle of 60ml syringe
2, 121 ℃, high-temperature heat sterilization 30min, for use.
2) inoculation: culture medium after sterilization is added 10% (w/v) NaHCO
3The Na of solution and 3% (w/v)
2S solution makes its final concentration reach 0.1% (w/v) and 0.3% (w/v), pH 7.0-9.5 respectively; Inoculation keratin anaerobic degrading bacteria 18D-TA, inoculum concentration 2%-10% (v/v), 45 ℃ of-60 ℃ of anaerobism leave standstill cultivated 8-10 hour; Feather down comes off; After 18-24 hour, feather comes off from the plumage stalk fully, and the plumage stalk also has degraded in various degree.
Consisting of of said culture medium: feather 1% (w/v), K
2HPO
43 g/L, KH
2PO
42.5 g/L, NaCl 3 g/L, MgSO
47H
2O 0. 02 g/L, CaCO
30. 02 g/L, vitamin liquid 1% (v/v), liquid microelement 1% (v/v), 0.1% (w/v) Cysteine-HClH
2O, 0.1% (w/v) resazurin solution, 0.1% (v/v), distilled water 1 L, pH 7.0-7.5.
Consisting of of said vitamin liquid: biotin 2.00 mg, folic acid 2.00 mg, pyridoxol-hydrochloric acid 10.00mg, two hydration thiamine-hydrochloric acid, 5.00 mg, riboflavin 5.00mg, nicotinic acid 5.00mg, D-calcium pantothenate 5.00mg, Cobastab
120.10mg, p-aminobenzoic acid 5.00mg, lipoic acid 5.00 mg, distilled water 1000.00ml.
Consisting of of said liquid microelement: MgSO
47 H
2O 3.00g, MnSO
4H
2O 0.50 g, NaCl 1.00 g, FeSO
47 H
2O 0.10 g, CoSO
47 H
2O 0.18 g, CaCl
22 H
2O 0.10 g, ZnSO
47 H
2O 0.18 g, CuSO
45 H
2O 0.01 g, KAl (SO
4)
212 H
2O 0.02g, H
3BO
30.01g, Na
2MoO
42 H
2O 0.01 g, NiCl
26 H
2O 0.03g, Na
2SeO
35 H
2O 0.30mg, distilled water 1000.00ml.
Said anaerobism pipe is selected from the specification that 10ml/ props up; Said serum bottle is selected from the specification of 120ml/ bottle.
Said feather is sole carbon source and the nitrogenous source of bacterial strain 18D-TA.
The culture of said inoculation back after 18-24 hour can be used as fermentation seed, carries out the enlarged culture of bacterial classification, carries out large scale fermentation.
Said bacterial strain belongs to
TepidimicrobiumThe bacterial classification of Pseudomonas, the classification called after of this bacterial strain
TepidimicrobiumSp., on April 28th, 2011 was preserved in China Committee for Culture Collection of Microorganisms common micro-organisms center, and the deposit number of bacterial classification is CGMCC NO
. 4800
Compare with tradition of having reported or bioanalysis, the invention has the advantages that:
1, the metabolizable energy of microorganism of the present invention is carried out under temperate condition, and pollution, the energy consumption that can avoid physico-chemical process processing feather to bring are big, and product nutrient is worth low shortcoming.
2, the bacterial strain that the present invention utilized belongs to the anaerobic fermentation bacterium, compares other aerobic bacterias, can add up more product, because of need not to provide aeration equipment, energy consumption and cost is reduced greatly.
3, the present invention carries out fermentation process under higher temperature, thereby has guaranteed that biodegradation process can not guaranteed the safe and sanitary of fermented product by microbiological contamination; In addition, higher temperature also can obtain better substrate dissolubility, and keratinase more is prone to substrate is had an effect.
4, the present invention degrades to feather under anaerobic condition, and its efficient is high, and fermentation period is short, and in known aerobic and anaerobic bacteria degradation bacteria, its degradation of feather by using speed is the fastest, thereby has opened up new approach for the degradation of feather by using microorganism large-scale production.
5, the present invention can be useful saves preliminary treatment; Like hot-pressing processing, particularly greater than the preliminary treatment under 130 ℃ of temperature, and such preliminary treatment is continued to use when the hydrolysis feather always; Thereby effectively avoided the racemization of Most amino-acids, improved the absorbing property of feather hydrolysate.
Description of drawings
Fig. 1 is that the interior feather of different time is by the variation diagram of 18D-TA bacterial strain anaerobic degradation.
The specific embodiment
Embodiment 1
Utilize the method for microbial strains 18D-TA anaerobic degradation feather keratin, it is characterized in that concrete steps are following:
The processing of A, feather
Feather washing is put into 45 ℃ of-55 ℃ of baking ovens after clean, dry 2-3 days subsequent use;
The inoculation of B, bacterial strain
1) preparation culture medium: add 500ml distilled water in advance in the triangular flask of 1L, add K respectively
2HPO
43 g/L, KH
2PO
42.5 g/L, NaCl 3 g/L, MgSO
47H
2O 0. 02 g/L, CaCO
30. 02 g/L vitamin liquid 1% (v/v), liquid microelement 1% (v/v) adds water to 1L then to its dissolving, boils 5-8min, is cooled to room temperature, adds 0.1% (w/v) Cysteine-HClH
2O regulates pH to 7.0-7.5 with 5mol/ L KOH, and adding concentration again is the resazurin solution 0.1% (v/v) of 0.1% (w/v), presses 2.0m
3/ h leads to N
2, boil 15min, cooling back is divided and is filled in the anaerobism pipe or serum bottle that is added with 1% (w/v) feather after processing of step A in advance, and is subsequent use behind 121 ℃, 30min high-temperature heat sterilization.
2) inoculation: culture medium after sterilization is added 10% (w/v) NaHCO
3The Na of solution and 3% (w/v)
2S solution makes its final concentration reach 0.1% (w/v) and 0.3% (w/v), pH 7.0-9.5 respectively; Inoculation keratin anaerobic degrading bacteria 18D-TA, inoculum concentration 2%-10% (v/v), 45 ℃ of-60 ℃ of anaerobism leave standstill cultivated 8-10 hour; Feather down comes off; After 18-24 hour, feather comes off from the plumage stalk fully, and the plumage stalk also has degraded in various degree.
Consisting of of said culture medium: feather 1% (w/v), K
2HPO
43 g/L, KH
2PO
42.5 g/L, NaCl 3 g/L, MgSO
47H
2O 0. 02 g/L, CaCO
30. 02 g/L, vitamin liquid 1% (v/v), liquid microelement 1% (v/v), 0.1% Cysteine-HClH
2O, 0.1% (w/v) resazurin solution 0.1%, distilled water 1 L.
Consisting of of said vitamin liquid: biotin 2.00 mg, folic acid 2.00 mg, pyridoxol-hydrochloric acid 10.00mg, two hydration thiamine-hydrochloric acid, 5.00 mg, riboflavin 5.00mg, nicotinic acid 5.00mg, D-calcium pantothenate 5.00mg, Cobastab
120.10mg, p-aminobenzoic acid 5.00mg, lipoic acid 5.00 mg, distilled water 1000.00ml.
Consisting of of said liquid microelement: MgSO
47 H
2O 3.00g, MnSO
4H
2O 0.50 g, NaCl 1.00 g, FeSO
47 H
2O 0.10 g, CoSO
47 H
2O 0.18 g, CaCl
22 H
2O 0.10 g, ZnSO
47 H
2O 0.18 g, CuSO
45 H
2O 0.01 g, KAl (SO
4)
212 H
2O 0.02g, H
3BO
30.01g, Na
2MoO
42 H
2O 0.01 g, NiCl
26 H
2O 0.03g, Na
2SeO
35 H
2O 0.30mg, distilled water 1000.00ml.
Said bacterial strain belongs to
TepidimicrobiumThe bacterial classification of Pseudomonas, the classification called after of this bacterial strain
TepidimicrobiumSp., on April 28th, 2011 was preserved in China Committee for Culture Collection of Microorganisms common micro-organisms center, and the deposit number of bacterial classification is CGMCC NO
. 4800
Claims (5)
1. utilize the method for microbial strains 18D-TA anaerobic degradation feather keratin, it is characterized in that concrete steps are following:
The processing of A, feather
Feather washing is put into 45 ℃ of-55 ℃ of baking ovens after clean, dry 2-3 days subsequent use;
The inoculation of B, bacterial strain
1) preparation culture medium: add 500ml distilled water in advance in the triangular flask of 1L, add K respectively
2HPO
43 g/L, KH
2PO
42.5 g/L, NaCl 3 g/L, MgSO
47H
2O 0. 02 g/L, CaCO
30. 02 g/L, vitamin liquid 1% (v/v), liquid microelement 1% (v/v) adds water to 1L then to its dissolving, boils 5-8min, is cooled to room temperature, adds 0.1% (w/v) Cysteine-HClH
2O regulates pH to 7.0-7.5 with 5mol/ L KOH, and adding concentration again is the resazurin solution 0.1% (v/v) of 0.1% (w/v), presses 2.0m
3/ h leads to N
2, boil 15min, cooling back is divided and is filled in the anaerobism pipe or serum bottle that is added with 1% (w/v) feather after processing of step A in advance, and is subsequent use behind 121 ℃, 30min high-temperature heat sterilization;
2) inoculation: culture medium after sterilization is added 10% (w/v) NaHCO
3The Na of solution and 3% (w/v)
2S solution makes its final concentration reach 0.1% (w/v) and 0.3% (w/v), pH 7.0-9.5 respectively; Inoculation keratin anaerobic degrading bacteria 18D-TA, inoculum concentration 2%-10% (v/v), 45 ℃ of-60 ℃ of anaerobism leave standstill cultivated 8-10 hour; Feather down comes off; After 18-24 hour, feather comes off from the plumage stalk fully, and the plumage stalk also has degraded in various degree.
2. utilize the method for microbial strains 18D-TA anaerobic degradation feather keratin according to claim 1, it is characterized in that: the consisting of of said culture medium: feather 1% (w/v), K
2HPO
43 g/L, KH
2PO
42.5 g/L, NaCl 3 g/L, MgSO
47H
2O 0. 2 g/L, CaCO
30. 02 g/L, vitamin liquid 1% (v/v), liquid microelement 1% (v/v), 0.1% (w/v) Cysteine-HClH
2O, 0.1% (w/v) resazurin solution 0.1%, distilled water 1 L.
3. like the said method of utilizing microbial strains 18D-TA anaerobic degradation feather keratin of claim 2, it is characterized in that: the consisting of of said vitamin liquid: biotin 2.00 mg, folic acid 2.00 mg; Pyridoxol-hydrochloric acid 10.00mg; Two hydration thiamine-hydrochloric acid, 5.00 mg, riboflavin 5.00mg, nicotinic acid 5.00mg; D-calcium pantothenate 5.00mg, Cobastab
120.10mg, p-aminobenzoic acid 5.00mg, lipoic acid 5.00 mg, distilled water 1000.00ml.
4. like the said method of utilizing microbial strains 18D-TA anaerobic degradation feather keratin of claim 2, it is characterized in that: the consisting of of said liquid microelement: MgSO
47 H
2O 3.00g, MnSO
4H
2O 0.50 g, NaCl 1.00 g, FeSO
47 H
2O 0.10 g, CoSO
47 H
2O 0.18 g, CaCl
22 H
2O 0.10 g, ZnSO
47 H
2O 0.18 g, CuSO
45 H
2O 0.01 g, KAl (SO
4)
212 H
2O 0.02g, H
3BO
30.01g, Na
2MoO
42 H
2O 0.01 g, NiCl
26 H
2O 0.03g, Na
2SeO
35 H
2O 0.30mg, distilled water 1000.00ml.
5. utilize the method for microbial strains 18D-TA anaerobic degradation feather keratin according to claim 1 or claim 2, it is characterized in that: said bacterial strain belongs to
TepidimicrobiumThe bacterial classification of Pseudomonas, the classification called after of this bacterial strain
TepidimicrobiumSp., on April 28th, 2011 was preserved in China Committee for Culture Collection of Microorganisms common micro-organisms center, and the deposit number of bacterial classification is CGMCC NO
. 4800
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Cited By (6)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102726596A (en) * | 2012-06-18 | 2012-10-17 | 杭州启明星生物营养有限公司 | Feather protein peptide feed additive and application thereof |
| CN104171413A (en) * | 2014-08-01 | 2014-12-03 | 农业部沼气科学研究所 | Meat poultry feed containing anaerobic bacterium K.paraultunense KD-1 fermentation liquid and feather meal |
| CN109590306A (en) * | 2018-11-09 | 2019-04-09 | 农业部沼气科学研究所 | A kind of processing method for poultry anaerobic biodegradation of dying of illness |
| CN109628534A (en) * | 2018-11-09 | 2019-04-16 | 农业部沼气科学研究所 | A kind of method of bacterium enzyme Combined Treatment degradation cyokeratin waste resource |
| CN109619268A (en) * | 2018-11-09 | 2019-04-16 | 农业部沼气科学研究所 | A kind of amino acid polypeptide feed and preparation method thereof |
| CN109704819A (en) * | 2018-11-09 | 2019-05-03 | 农业部沼气科学研究所 | A kind of amino acid polypeptide Water soluble fertilizer and preparation method thereof |
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Cited By (7)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102726596A (en) * | 2012-06-18 | 2012-10-17 | 杭州启明星生物营养有限公司 | Feather protein peptide feed additive and application thereof |
| CN104171413A (en) * | 2014-08-01 | 2014-12-03 | 农业部沼气科学研究所 | Meat poultry feed containing anaerobic bacterium K.paraultunense KD-1 fermentation liquid and feather meal |
| CN109590306A (en) * | 2018-11-09 | 2019-04-09 | 农业部沼气科学研究所 | A kind of processing method for poultry anaerobic biodegradation of dying of illness |
| CN109628534A (en) * | 2018-11-09 | 2019-04-16 | 农业部沼气科学研究所 | A kind of method of bacterium enzyme Combined Treatment degradation cyokeratin waste resource |
| CN109619268A (en) * | 2018-11-09 | 2019-04-16 | 农业部沼气科学研究所 | A kind of amino acid polypeptide feed and preparation method thereof |
| CN109704819A (en) * | 2018-11-09 | 2019-05-03 | 农业部沼气科学研究所 | A kind of amino acid polypeptide Water soluble fertilizer and preparation method thereof |
| CN109628534B (en) * | 2018-11-09 | 2020-07-24 | 农业部沼气科学研究所 | Method for degrading keratin waste resources through combined treatment of bacteria and enzymes |
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|---|---|
| CN102326668B (en) | 2012-11-28 |
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Application publication date: 20120125 Assignee: Chengdu Bai Xinwei Technology Co. Ltd. Assignor: Methane Scientific Research Inst., Ministry of Agriculture Contract record no.: 2015510000137 Denomination of invention: Method for anaerobically degrading feather keratin with microbial strain 18D-TA Granted publication date: 20121128 License type: Exclusive License Record date: 20151029 |
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