CN1318582C - Aspergillus niger one-step fermentation production of multienzyme high-enzyme activity composite enzyme - Google Patents
Aspergillus niger one-step fermentation production of multienzyme high-enzyme activity composite enzyme Download PDFInfo
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- CN1318582C CN1318582C CNB021048703A CN02104870A CN1318582C CN 1318582 C CN1318582 C CN 1318582C CN B021048703 A CNB021048703 A CN B021048703A CN 02104870 A CN02104870 A CN 02104870A CN 1318582 C CN1318582 C CN 1318582C
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- 108090000790 Enzymes Proteins 0.000 title claims abstract description 32
- 102000004190 Enzymes Human genes 0.000 title claims abstract description 32
- 238000000855 fermentation Methods 0.000 title claims abstract description 24
- 230000004151 fermentation Effects 0.000 title claims abstract description 24
- 238000004519 manufacturing process Methods 0.000 title claims abstract description 10
- 241000228245 Aspergillus niger Species 0.000 title claims abstract description 9
- 230000000694 effects Effects 0.000 title claims abstract description 7
- 239000002131 composite material Substances 0.000 title description 2
- 229940088598 enzyme Drugs 0.000 claims abstract description 33
- 238000000034 method Methods 0.000 claims abstract description 8
- 239000003674 animal food additive Substances 0.000 claims abstract description 6
- 101710130006 Beta-glucanase Proteins 0.000 claims abstract description 4
- 108010059892 Cellulase Proteins 0.000 claims abstract description 4
- 108010059820 Polygalacturonase Proteins 0.000 claims abstract description 4
- 229940106157 cellulase Drugs 0.000 claims abstract description 4
- 239000007787 solid Substances 0.000 claims abstract description 4
- 238000010563 solid-state fermentation Methods 0.000 claims abstract description 3
- 235000019764 Soybean Meal Nutrition 0.000 claims abstract 3
- 239000004455 soybean meal Substances 0.000 claims abstract 3
- 108091005508 Acid proteases Proteins 0.000 claims abstract 2
- 101710121765 Endo-1,4-beta-xylanase Proteins 0.000 claims abstract 2
- 108010093305 exopolygalacturonase Proteins 0.000 claims abstract 2
- 238000009423 ventilation Methods 0.000 claims abstract 2
- 238000002360 preparation method Methods 0.000 claims description 9
- 239000000463 material Substances 0.000 claims description 7
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- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims description 5
- 239000000725 suspension Substances 0.000 claims description 4
- 239000004382 Amylase Substances 0.000 claims description 3
- 108010065511 Amylases Proteins 0.000 claims description 3
- 102000013142 Amylases Human genes 0.000 claims description 3
- BFNBIHQBYMNNAN-UHFFFAOYSA-N ammonium sulfate Chemical compound N.N.OS(O)(=O)=O BFNBIHQBYMNNAN-UHFFFAOYSA-N 0.000 claims description 3
- 229910052921 ammonium sulfate Inorganic materials 0.000 claims description 3
- 235000011130 ammonium sulphate Nutrition 0.000 claims description 3
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- 238000004321 preservation Methods 0.000 claims description 3
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- 229920001817 Agar Polymers 0.000 claims description 2
- 244000061456 Solanum tuberosum Species 0.000 claims description 2
- 235000002595 Solanum tuberosum Nutrition 0.000 claims description 2
- CZMRCDWAGMRECN-UGDNZRGBSA-N Sucrose Chemical compound O[C@H]1[C@H](O)[C@@H](CO)O[C@@]1(CO)O[C@@H]1[C@H](O)[C@@H](O)[C@H](O)[C@@H](CO)O1 CZMRCDWAGMRECN-UGDNZRGBSA-N 0.000 claims description 2
- 229930006000 Sucrose Natural products 0.000 claims description 2
- 239000008272 agar Substances 0.000 claims description 2
- 239000011521 glass Substances 0.000 claims description 2
- 239000000203 mixture Substances 0.000 claims description 2
- 229940085127 phytase Drugs 0.000 claims description 2
- 239000005720 sucrose Substances 0.000 claims description 2
- 239000001963 growth medium Substances 0.000 claims 3
- 150000001875 compounds Chemical class 0.000 claims 2
- 239000002609 medium Substances 0.000 claims 2
- 241000228212 Aspergillus Species 0.000 claims 1
- 239000004743 Polypropylene Substances 0.000 claims 1
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- 229910000402 monopotassium phosphate Inorganic materials 0.000 claims 1
- 235000019796 monopotassium phosphate Nutrition 0.000 claims 1
- 238000003199 nucleic acid amplification method Methods 0.000 claims 1
- PJNZPQUBCPKICU-UHFFFAOYSA-N phosphoric acid;potassium Chemical compound [K].OP(O)(O)=O PJNZPQUBCPKICU-UHFFFAOYSA-N 0.000 claims 1
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- -1 polypropylene Polymers 0.000 claims 1
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- 244000144972 livestock Species 0.000 abstract description 2
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- 102000035101 Aspartic proteases Human genes 0.000 description 2
- 108091005502 Aspartic proteases Proteins 0.000 description 2
- 229910019142 PO4 Inorganic materials 0.000 description 2
- ZLMJMSJWJFRBEC-UHFFFAOYSA-N Potassium Chemical compound [K] ZLMJMSJWJFRBEC-UHFFFAOYSA-N 0.000 description 2
- 241001052560 Thallis Species 0.000 description 2
- 238000001816 cooling Methods 0.000 description 2
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- 238000012262 fermentative production Methods 0.000 description 2
- 239000002054 inoculum Substances 0.000 description 2
- 244000005700 microbiome Species 0.000 description 2
- MBDIPBHBEVOYQB-UHFFFAOYSA-N nigerone Chemical compound O1C(C)=CC(=O)C2=C1C(C1=C3C(C(C=C(C)O3)=O)=C(O)C3=C(OC)C=C(C=C31)OC)=C1C=C(OC)C=C(OC)C1=C2O MBDIPBHBEVOYQB-UHFFFAOYSA-N 0.000 description 2
- 238000000643 oven drying Methods 0.000 description 2
- NBIIXXVUZAFLBC-UHFFFAOYSA-K phosphate Chemical compound [O-]P([O-])([O-])=O NBIIXXVUZAFLBC-UHFFFAOYSA-K 0.000 description 2
- 239000010452 phosphate Substances 0.000 description 2
- 229910052700 potassium Inorganic materials 0.000 description 2
- 239000011591 potassium Substances 0.000 description 2
- 238000010298 pulverizing process Methods 0.000 description 2
- IMQLKJBTEOYOSI-GPIVLXJGSA-N Inositol-hexakisphosphate Chemical compound OP(O)(=O)O[C@H]1[C@H](OP(O)(O)=O)[C@@H](OP(O)(O)=O)[C@H](OP(O)(O)=O)[C@H](OP(O)(O)=O)[C@@H]1OP(O)(O)=O IMQLKJBTEOYOSI-GPIVLXJGSA-N 0.000 description 1
- 241001465754 Metazoa Species 0.000 description 1
- IMQLKJBTEOYOSI-UHFFFAOYSA-N Phytic acid Natural products OP(O)(=O)OC1C(OP(O)(O)=O)C(OP(O)(O)=O)C(OP(O)(O)=O)C(OP(O)(O)=O)C1OP(O)(O)=O IMQLKJBTEOYOSI-UHFFFAOYSA-N 0.000 description 1
- 206010044565 Tremor Diseases 0.000 description 1
- NIXOWILDQLNWCW-UHFFFAOYSA-N acrylic acid group Chemical group C(C=C)(=O)O NIXOWILDQLNWCW-UHFFFAOYSA-N 0.000 description 1
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- 239000012467 final product Substances 0.000 description 1
- 239000004459 forage Substances 0.000 description 1
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- 229910017053 inorganic salt Inorganic materials 0.000 description 1
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- 229940068041 phytic acid Drugs 0.000 description 1
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- 235000020679 tap water Nutrition 0.000 description 1
- 239000002351 wastewater Substances 0.000 description 1
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- Micro-Organisms Or Cultivation Processes Thereof (AREA)
- Enzymes And Modification Thereof (AREA)
Abstract
The invention relates to a method for producing multienzyme high-enzyme-activity complex enzyme by one-time fermentation of aspergillus niger, belonging to the fields of biochemistry and enzyme engineering. A good strain suitable for solid state fermentation, namely Aspergillus niger HQ978, is obtained through mutagenesis and screening. The method is characterized in that a fermentation medium mainly containing agricultural and sideline products such as bran, soybean meal and the like is utilized, and a solid fermentation machine is used for ventilation fermentation in a thick layer to obtain the complex enzyme such as acid protease, cellulase, xylanase, beta-glucanase, pectinase and the like with high enzyme activity unit. The product is a green feed additive for livestock, fowl, aquatic product, etc.
Description
One. technical field
The invention belongs to biological chemistry and enzyme engineering field.Enzyme extensively is present in all organisms, is a kind of active substance that organism produces, and is the catalyzer of various biochemical reactions in the body.The digestion of various nutritive substances absorbs and utilizes the effect of all necessary dependence enzyme.The present invention utilizes black mold HQ978 single culture to produce by biological engineering method and specific production technique to have highly active enzyme product.
Two. background technology
Fodder enzyme preparation is new a member that emerges in the fodder additives.Since the U.S. in 1975 uses the microorganism fodder enzyme preparation first, significant progress has been arranged.The development and application of fodder enzyme preparation is that biotechnology is used the most successful example in Animal nutrition and fodder industry; it is improving breeding performonce fo animals and efficiency of feed utilization; develop new feed resource; produce many-sides such as the natural livestock product of natural, nontoxic, noresidue and environment protection, demonstrate great potential gradually.The existing zymin of having used more than 80% of the feed of developed country.China's fodder industry starting is late, but development is very fast, produces about 7,000 ten thousand tons per year, and the market space is very big.
At present external complex enzyme preparation for feeding and the domestic minority unit of producing is that, dilution composite by certain prescription with several single enzyme preparations of liquid submerged fermentation forms, and enzyme system is incomplete, and the cost height is difficult to satisfy the needs of forage component complexity.
Purpose of the present invention just provides that production cost is low, that the enzyme of product system is complete, enzyme is lived is high, the method for the green feed additive of compliance with environmental protection requirements.
Advantage of the present invention:
1. adopt the outstanding aspergillus niger single culture (Aspergillusniger HQ978) that obtains through mutagenesis screening; Preserving number is CGMCC NO:0696.Depositary institution: China Committee for Culture Collection of Microorganisms common micro-organisms center, preservation place: China, Beijing, Zhong Guan-cun, preservation time: on January 23rd, 2002.
2. applying solid fermentation machine thick-layer ventilating fermentation, these facility have that automatic temperature-control, stand are drawn together, stirring, discharging and sterilized humidifying equipment; Yet available shallow tray fermentation mode is not produced when not possessing the fermentation machine.
3. fermention medium is based on wheat bran, dregs of beans and small amounts of inorganic salt, the agricultural byproducts that its raw material sources are wide, price is lower, and production cost is lower;
4.One time fermentation can obtain the multienzyme kind, and the leaven material that high enzyme is lived through oven drying at low temperature, pulverizing, is prepared complex enzyme feed additive;
5. production process does not produce waste water and dregs, causes secondary environmental pollution.
Three. summary of the invention
The present invention is a kind of complex enzyme feed additive, the production bacterial classification is aspergillus niger (Aspergillus niger HQ978) bacterial classification, culture presevation number is: CGMCCNO:0696, promptly obtain the product that enzyme of several kinds of enzyme and with high is lived through a solid state fermentation, the composition of this product and enzyme work thereof comprise: aspartic protease 10000～11000u/g, cellulase 18000～20000u/g, zytase 60000～80000u/g, beta-glucanase 50000～60000u/g, polygalacturonase 18000～20000u/g, amylase and phytase.
(1) strain preparation
Strain preparation is divided into one, two, three strain preparation:
1. the first class inoculum substratum is conventional potato sucrose agar slant, and the method for scoring inoculation is cultivated 96～120h for 28 ± 1 ℃.
2. the second class inoculum substratum is wheat bran and dregs of beans, and its ratio is 8: 2, each 300ml glass bottle 10g, and material: water is 1: 0.6,121 ℃ of sterilization 30min, cooling inserts one-level kind spore suspension, and every bottle of 5ml cultivates 72～74h for 28 ± 1 ℃.
3. the same secondary of three-class strain substratum.The packed substratum 90g that goes into through profit water of the acrylic plastering of each 22 * 30cm, 121 ℃ of sterilization 30min, cooling inserts the secondary seed spore suspension, every bag of 15ml, 29～30 ℃ of constant temperature culture, 24h falls bag once, is about to caking and trembles and pinches into fritter, continues to cultivate; 48～60h respectively turns over bag once.General 72～the 75h that cultivates, black in a large number spore maturation can or refrigerate standby for the fermentative production use.
(2) fermentative production
1. fermentation machine solid ventilating fermentation machine, (butt meter) 1000kg feeds intake at every turn.
2. fermention medium preparation substratum is a wheat bran 60～80%, dregs of beans 15～35%, ammonium sulfate 3～4%, potassium primary phosphate 1～2%, natural pH mixes wheat bran and dregs of beans thoroughly earlier, ammonium sulfate and potassium primary phosphate are dissolved in the tap water that is equivalent to siccative 70%, evenly spray in the material, autoclaving 40～50min cools off standby.
3. inoculate the spore that washs 60～80 bags three-class strain (being equivalent to siccative 0.3～0.8%) with the sterilized water that is equivalent to substratum siccative 50%, evenly spray into the fermention medium of sterilizing.
4. the material input that will connect bacterial classification of fermenting is cultivated through the fermentor tank of slack tank sterilization.29～30 ℃ of design temperatures generally are cultured to about 24h, and thalli growth well lumps, and carry out the stirring first time, carry out second and third time stirring with backsight thalli growth caking situation, are advisable with the agglomerate that keeps 3～5cm.When first and second time stirring, if moisture＜30% o'clock can take the circumstances into consideration to spray into sterilized water, help producing enzyme, fermentation time generally is controlled at 60-62h and finishes fermentation.
(3) oven dry and pulverizing
After going out jar, bent material is through crusher in crushing, oven drying at low temperature, general＜45 ℃ dry material and contain moisture 8～10% and get final product, pulverize then, detect, get finished product.The enzyme of product and enzyme unit alive: aspartic protease 10000～11000u/g, cellulase 18000～20000u/g, zytase 60000～80000u/g, beta-glucanase 50000～60000u/g, polygalacturonase 18000～20000u/g, amylase and phytic acid.
Claims (2)
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CNB021048703A CN1318582C (en) | 2002-02-26 | 2002-02-26 | Aspergillus niger one-step fermentation production of multienzyme high-enzyme activity composite enzyme |
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| Application Number | Priority Date | Filing Date | Title |
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| CNB021048703A CN1318582C (en) | 2002-02-26 | 2002-02-26 | Aspergillus niger one-step fermentation production of multienzyme high-enzyme activity composite enzyme |
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| CN1374402A CN1374402A (en) | 2002-10-16 |
| CN1318582C true CN1318582C (en) | 2007-05-30 |
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| CNB021048703A Expired - Fee Related CN1318582C (en) | 2002-02-26 | 2002-02-26 | Aspergillus niger one-step fermentation production of multienzyme high-enzyme activity composite enzyme |
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Families Citing this family (15)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN100448979C (en) * | 2005-12-09 | 2009-01-07 | 山东省中协食品添加剂研究开发中心 | Production method for xylose by enzyme process |
| CN101117628B (en) * | 2006-08-01 | 2010-12-08 | 中国农业科学院北京畜牧兽医研究所 | Method for producing phytase |
| CN1919044B (en) * | 2006-09-20 | 2010-10-06 | 南京林业大学 | Animal feedstuff additive and preparation method thereof |
| CN101326967B (en) * | 2008-08-01 | 2011-05-11 | 重庆市畜牧科学院 | Composite pre-mix material for pig and preparation thereof |
| CN101591619B (en) * | 2009-07-03 | 2011-05-18 | 中国农业科学院饲料研究所 | Aspergillus niger strain and use thereof |
| CN102242091B (en) * | 2011-04-21 | 2012-11-28 | 河南省科学院生物研究所有限责任公司 | Method for preparing beta-glucanase for feeding by utilizing solid-state fermentation of peanut hull meal |
| CN102732433B (en) * | 2012-06-15 | 2014-08-27 | 河南省科学院生物研究所有限责任公司 | Strain capable of producing compound enzyme and method of using same to prepare compound enzyme for weaned piglets |
| CN104630069B (en) * | 2013-11-12 | 2017-11-28 | 天津工业大学 | The aspergillus niger of one plant height cellulase-producing |
| CN103695396B (en) * | 2013-12-23 | 2015-09-02 | 湖南鸿鹰生物科技有限公司 | A kind of high vigor complex enzyme for feed |
| CN103740601B (en) * | 2013-12-23 | 2015-07-29 | 湖南鸿鹰生物科技有限公司 | A kind of aspergillus niger of high yield complex enzyme for feed and application thereof |
| CN103805582B (en) * | 2013-12-27 | 2017-02-01 | 湖南鸿鹰生物科技有限公司 | Beta-glucanase containing special enzyme for growing pigs and preparation method thereof |
| CN109576245A (en) * | 2018-12-27 | 2019-04-05 | 北京华美源生物科技有限公司 | Barley diet barley enzyme preparation and its zymotechnique |
| CN111642639B (en) * | 2020-03-02 | 2024-07-26 | 漳州大北农农牧科技有限公司 | Biological fermentation feed for piglets |
| CN111593036B (en) * | 2020-06-08 | 2023-01-20 | 杭州保安康生物技术有限公司 | Preparation of enzyme preparation mainly containing acid protease, strain and application thereof |
| CN111607574B (en) * | 2020-06-08 | 2021-05-14 | 杭州保安康生物技术有限公司 | Enzyme preparation mainly comprising xylanase and acid protease, and strain and application thereof |
Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| JPH03188093A (en) * | 1989-12-18 | 1991-08-16 | Gun Ei Chem Ind Co Ltd | Novel saccharide and production thereof |
| CN1187312A (en) * | 1997-12-30 | 1998-07-15 | 中国科学技术大学研究生院 | Method for producing biologic inorganic composite feed additive for animals and fowls |
| JP3188093B2 (en) * | 1994-04-06 | 2001-07-16 | 三洋電機株式会社 | Cassette mounting mechanism and magnetic recording / reproducing apparatus provided with the mechanism |
-
2002
- 2002-02-26 CN CNB021048703A patent/CN1318582C/en not_active Expired - Fee Related
Patent Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| JPH03188093A (en) * | 1989-12-18 | 1991-08-16 | Gun Ei Chem Ind Co Ltd | Novel saccharide and production thereof |
| JP3188093B2 (en) * | 1994-04-06 | 2001-07-16 | 三洋電機株式会社 | Cassette mounting mechanism and magnetic recording / reproducing apparatus provided with the mechanism |
| CN1187312A (en) * | 1997-12-30 | 1998-07-15 | 中国科学技术大学研究生院 | Method for producing biologic inorganic composite feed additive for animals and fowls |
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