CN1318582C - Once aspergillus niger fermentation process of obtaining complex enzyme of several kinds of enzyme and with high enzyme activity - Google Patents

Once aspergillus niger fermentation process of obtaining complex enzyme of several kinds of enzyme and with high enzyme activity Download PDF

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Publication number
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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China
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enzyme
fermentation
aspergillus niger
substratum
several kinds
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CNB021048703A
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Chinese (zh)
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CN1374402A (en
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黄文彩
戚秀芳
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东莞市豪发生物工程开发有限公司
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Abstract

The present invention relates to a once Aspergillus niger fermentation process of obtaining complex enzyme of several kinds of enzyme and with high enzyme activity, which belongs to the field of biochemistry and enzyme engineering. An excellent bacterium strain-Aspergillus niger HQ978 which is suitable for solid fermentation is obtained through induced mutagenesis screening. In a fermentation medium with bran, bean dreg and other agricultural side products as main components and through thick-layer ventilated fermentation in a solid fermentation machine, a high-activity complex enzyme comprising acid proteinase, cellulase, xylanase, beta-glucanase, pectinase, etc. is obtained. The product is a green feed additive for domestic animals, poultry, marine products and other feeding animals.

Description

The production of aspergillus niger one time fermentation obtains complex enzyme of several kinds of enzyme and with high enzyme activity

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)

1. complex enzyme feed additive, it is characterized in that aspergillus niger (Aspergillus nigerHQ978) 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.
2. the described complex enzyme feed additive production method of claim 1 comprises strain preparation technology and zymotechnique:
Strain preparation comprises one, two, three bacterial classification technology of amplification culture bacterial classification step by step:
The first class inoculum substratum is conventional potato sucrose agar slant, and the method for scoring inoculation is cultivated 96~120h for 28 ± 1 ℃;
The second class inoculum substratum is wheat bran and dregs of beans, and its ratio is 8: 2, every 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 ℃;
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, insert the secondary seed spore suspension, every bag of 15ml, 29~30 ℃ of constant temperature culture, 24h falls bag once, and being about to lumps to tremble pinches into fritter, continues to cultivate; 48~60h respectively turns over bag once again, generally cultivates 72~75h, and black in a large number spore maturation can or refrigerate standby for the fermentative production use;
Zymotechnique: fermention medium is that wheat bran 60~80%, dregs of beans are 15~35%, ammonium sulfate 3~4%, potassium primary phosphate 1~2%, natural pH value; Adopt solid fermentation machine thick-layer ventilating fermentation or shallow tray fermentation mode to produce; 28~30 ℃ of culture temperature, fermentation time 60~62h.
CNB021048703A 2002-02-26 2002-02-26 Once aspergillus niger fermentation process of obtaining complex enzyme of several kinds of enzyme and with high enzyme activity CN1318582C (en)

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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

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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

Patent Citations (2)

* Cited by examiner, † Cited by third party
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

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