CN104686804A - Method for preparing active peptide feed additive from aquatic product leftovers - Google Patents
Method for preparing active peptide feed additive from aquatic product leftovers Download PDFInfo
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- CN104686804A CN104686804A CN201510092514.9A CN201510092514A CN104686804A CN 104686804 A CN104686804 A CN 104686804A CN 201510092514 A CN201510092514 A CN 201510092514A CN 104686804 A CN104686804 A CN 104686804A
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- fermentation
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- bacillus subtilis
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- Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
- Y02—TECHNOLOGIES OR APPLICATIONS FOR MITIGATION OR ADAPTATION AGAINST CLIMATE CHANGE
- Y02A—TECHNOLOGIES FOR ADAPTATION TO CLIMATE CHANGE
- Y02A40/00—Adaptation technologies in agriculture, forestry, livestock or agroalimentary production
- Y02A40/80—Adaptation technologies in agriculture, forestry, livestock or agroalimentary production in fisheries management
- Y02A40/81—Aquaculture, e.g. of fish
- Y02A40/818—Alternative feeds for fish, e.g. in aquacultures
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Abstract
The invention discloses a method for preparing an active peptide feed additive from aquatic product leftovers, comprising the following main steps: mincing the fresh aquatic product leftovers; adding the minced aquatic product leftovers into a fermentation tank, further adding nutrients and water, and mixing together to obtain a fermentation culture medium; sterilizing and cooling the culture medium in the fermentation tank, and then inoculating Bacillus subtilis HL-1 capable of producing subtilin to carry out liquid state aerobic fermentation; and after fermentation, heating up fermentation liquid to 90 DEG C, deactivating enzyme for 5min, then, removing sediments by filtering to obtain a filtrate, and further carrying out vacuum concentration and spray drying on the filtrate to obtain the active peptide feed additive. The active peptide feed additive prepared by the method is rich in antibacterial peptide and nutritional small peptides, has good growth promoting performance and disease resistance, and has good application foreground and social significance.
Description
Technical field
The present invention relates to a kind of method utilizing aquatic product leftovers to prepare activated peptide forage additive, belong to processing of aquatic products and utilize field.
Background technology
Since the 1950's, in animal feeding and feed, the growth promotion health-care agents such as antibiotic, chemical synthetic drug, hormone, beta-stimulants, heavy metal, sedative are widely used.The applicating history of the growth promotion health-care agents such as antibiotic in feed is long-standing, it creates the revolution of aquaculture and feed industry, breeding scale is obtained in worldwide develop rapidly, production quantity and the feed conversion rate of animal are greatly improved.But the achievement of industrial civilization has its negative effect the same as in all, use the non-green feed addictives such as antibiotic for a long time, result in following problem: livestock and poultry and aquatic products product drug residue seriously, are directly detrimental to health; Livestock and poultry and aquatic products deterioration in quality, lack the market competitiveness and trust; Bacterial drug resistance strengthens, antibody-resistant bacterium increases, and threatens human security; Destroy ecological environment etc.Along with the fast development of feed industry and animal husbandry and fishery industry, the livestock products of current China and aquatic products have changed into interim superfluous from shortage, people are more concerned about food whether safety, health, thus green consumption has become development trend from now on, in addition China joins WTO, developed country requires more and more stricter to the agricultural product of China's export, current medicament residue problem becomes one of major obstacle that China's livestock products and aquatic products go abroad.Therefore accelerate development, promotion and application novel green feed addictive are imperative.
A large amount of documents shows, the active peptide adding separate sources in daily ration all can improve the production performance of different letting animals feed, and the growth promotion performance of active peptide has obtained generally acknowledging of feedstuff industry.Thus, as a kind of novel green feed addictive, activated peptide forage additive has good application prospect.Biologically active peptide can be divided into two large class, the i.e. little peptide of nutrition and Functional Polypeptides.The little peptide of trophism is only for protein synthesis provides nitrogen frame, does not have the active peptide of special physiological regulatory function.Functional Polypeptides is the peptide with some specific function, as antibacterial peptide, anti-oxidation peptide, immune peptide and Hormone Peptide etc.
The method of macro-molecular protein chemistry or enzyme is mainly hydrolyzed into micromolecular peptide by the preparation of active peptide.The preparation method of current active peptide has chemical method, enzymatic isolation method and microbe fermentation method.Chemical hydrolysis adopts sour, that alkali chemical reagent impels protein molecule at a certain temperature Fragmentation to form small-molecular peptides.Although chemical hydrolysis is simple, become wood low, its shortcoming to carry out production control regularly, and amino acid can suffer damage and reduce its nutritive value, and big for environment pollution.Current chemical hydrolysis is used for test, uses less in production practices.It is take protein raw materials as substrate that enzymatic isolation method produces active peptide, uses the enzymolysis of protease, produces small-molecular peptides.Enzymatic isolation method is easy to control, and action condition gentleness is applicable to the production of active peptide.But enzymatic isolation method needs a large amount of protease, and production cost is high.Therefore, enzymatic isolation method is prepared active peptide and is used for food and pharmaceuticals industry, and the feeding active peptide product of low side seldom adopts the method for enzymolysis.Microbe fermentation method take macro-molecular protein as raw material, and the microorganism of protease is produced in access, the enzymolysis of the protease utilizing fermentation process to produce, and generates small-molecular peptides.Microbe fermentation method organically combines the fermenting and producing of protease and enzymolysis process, reduces the manufacturing cost of active peptide, is more suitable for the production of active peptide.Patent CN200410029787.0 " a kind of activated peptide forage additive and preparation method and purposes " discloses a kind of fermentation and sends out the method preparing activated peptide forage additive, it is the mixture of one or more with vegetable protein and animal protein, through fermentative degradations such as bacillus, make its degree of hydrolysis control 20 ~ 35%, then can obtain activated peptide forage additive through post processing.But the uncontrollability of the concentration due to sweat protease, adds the randomness of enzymolysis, the active peptide that thus prepared by above method mostly is the little peptide of nutrition, and lacks Functional Polypeptides, particularly the antibacterial peptide of feed industry urgent need.
As a kind of substitute of feeding antibiotic, antibacterial peptide usually has heat-resisting, has a broad antifungal spectrum, good water solubility and not easily produces the characteristics such as drug resistance, receives much concern in recent years.Antibacterial peptide derive from animal, plant and microorganism, but animal and plant source antibacterial peptide is due to limited source, and application is restricted; And microbial antibacterial peptide can large-scale production, advantage of lower cost, thus microbial source antibacterial peptide more has application prospect.Current microbial source antibacterial peptide most study be the antibacterial peptide of lactic acid bacteria, and in the world to bacillus subtilis (
bacillus subtilis) this novel fine bacteriums of the subtilin antibacterial peptide research that produces is then little, domestic rare especially.
Summary of the invention
The activated peptide forage additive prepared for current fermentation method mostly is the little peptide of nutrition, and lacks antibacterial peptide, makes its effect outstanding not.The object of the invention is to prepare a kind of activated peptide forage additive being rich in antibacterial peptide, improving on the basis of activated peptide forage additive growth promotion performance further, also giving its resistance against diseases.Thus significantly improve effect of activated peptide forage additive, to promote that activated peptide forage additive is in the extensive use of feed industry.
In order to solve the problem, the technical solution adopted in the present invention is as follows: a kind of method utilizing aquatic product leftovers to prepare activated peptide forage additive, comprises the steps:
(1) by slant preservation can produce subtilin bacillus subtilis (
bacillus subtilis) in the LB fluid nutrient medium that cools after being linked into sterilizing of HL-1, in 37 DEG C, 150r/min cultivates 16h, be prepared into bacillus subtilis (
bacillus subtilis) HL-1 seed liquor is for subsequent use;
(2) for subsequent use after fresh aquatic product leftovers being rubbed;
(3) aquatic product leftovers of rubbing obtained for step (2) is put in fermentation tank, and add water and nutrients in fermentation tank, after mixing, obtain fermentation medium;
(4) fermented and cultured in fermentation tank step (3) obtained is based on 115 DEG C of sterilizing 20min;
(5) fermentation medium after sterilizing step (4) obtained is cooled to room temperature, then access bacillus subtilis prepared by step (1) (
bacillus subtilis) HL-1 seed liquor;
(6) control of sweat: fermentation temperature is 28 DEG C ~ 30 DEG C, and filtrated air intake is 0.5vvm ~ 2.5vvm, the rotating speed of fermentation tank is 150rpm ~ 350rpm, and fermentation time is 35h ~ 45h;
(7) product handling system: fermentation ends after fermentation liquid is warming up to 90 DEG C of insulations and goes out enzyme 5min; After the zymotic fluid after enzyme that goes out is cooled to room temperature, removes precipitation after filtration and obtain filtrate; Filtrate obtains activated peptide forage additive finished product again after Vacuum Concentration and spraying dry.
The bacillus subtilis of access in described step (1) (
bacillus subtilis) HL-1 can produce subtilin, this bacterium is preserved in China Committee for Culture Collection of Microorganisms's common micro-organisms center on January 20th, 2015, deposit number is CGMCC No.10379, and preservation address is No. 3, Yard 1, BeiChen xi Road, Chaoyang District, Beijing City Institute of Microorganism, Academia Sinica.
Aquatic product leftovers in described step (2) is shrimp head, scallop body, the leftovers of tilapia removing internal organ and fish-skin, the cod leftover bits and pieces removing internal organ and fish-skin, the carp leftover bits and pieces removing internal organ and fish-skin, the black carp leftover bits and pieces removing internal organ and fish-skin, the silver carp leftover bits and pieces removing internal organ and fish-skin, the grass carp leftover bits and pieces removing internal organ and fish-skin and the one removed in the stripped tuna leftover bits and pieces of internal organ and fish-skin.In described step (3), in fermentation tank, the addition of aquatic product leftovers accounts for water gross mass percentages with the quality of the aquatic product leftovers added, and is 10% ~ 20%.
The mixture that nutrients in described step (3) is glucose, frerrous chloride and dipotassium hydrogen phosphate form; Nutraceutical addition in described step (3) accounts for water gross mass percentages with the nutraceutical quality of adding, and is respectively: glucose 0.01% ~ 5%, frerrous chloride 0.01% ~ 5%, dipotassium hydrogen phosphate 0.01% ~ 5%.
In described step (5), the inoculum concentration of bacillus subtilis (Bacillus subtilis) HL-1 seed liquor accounts for fermentation medium gross mass percentages with the quality of the seed liquor of access, is 0.1% ~ 5%.
The present invention compares with existing technology, the present invention adopt can produce subtilin bacillus subtilis (
bacillus subtilis) carry out purpose fermentation, while obtaining the little peptide of nutrition, also obtain this bacteriocin class of subtilin antibacterial peptide, the activated peptide forage additive that thus prepared by the inventive method is rich in antibacterial peptide.Be rich in antibacterial peptide, feed addictive prepared by the present invention, improving on the basis of activated peptide forage additive growth promotion performance further, is also given its resistance against diseases, is significantly improved effect of activated peptide forage additive.
The invention provides a kind of method utilizing aquatic product leftovers to prepare activated peptide forage additive.The culture experiment of the pig of the activated peptide forage additive that the present invention obtains in South China, chicken, prawn shows, the feed that the activated peptide forage additive produced with the present invention coordinates, not only can improve the growth result of letting animals feed, but also immunity and the premunition of letting animals feed can be improved, directly can be used for preparing green feed, have a good application prospect and social effect.
Detailed description of the invention
In order to understand the present invention better, with regard to the selection of the kind of aquatic product leftovers, fermentation parameter and fermentation mode, the invention will be further described in conjunction with the embodiments below, but protection scope of the present invention is not limited thereto.
Embodiment 1: by slant preservation can produce subtilin bacillus subtilis (
bacillus subtilis) in the LB fluid nutrient medium that cools after being linked into sterilizing of HL-1, in 37 DEG C, 150r/min cultivates 16h, be prepared into bacillus subtilis (
bacillus subtilis) HL-1 seed liquor is for subsequent use.For subsequent use after the fresh leftovers of tilapia removing internal organ and fish-skin is rubbed.10 kilograms of leftovers of tilapia rubbed and 100 kg of water are joined in fermentation tank, and in fermentation tank, adds 800 grams of glucose, 300 grams of frerrous chlorides and 500 grams of dipotassium hydrogen phosphates, obtain fermentation medium; Fermentation cylinder for fermentation is cultivated based on 115 DEG C of sterilizing 20min; Then add in the fermentation raw material cooled after sterilizing 2 kilograms of bacillus subtilises (
bacillus subtilis) HL-1 seed liquor; Fermentation temperature is 30 DEG C, and filtrated air intake is 1.5vvm ~ 2.5vvm, and the rotating speed of fermentation tank is 250rpm ~ 350rpm, and fermentation time is 35h; Fermentation ends after fermentation liquid is warming up to 90 DEG C of enzyme 5min that go out, after the zymotic fluid after enzyme that goes out is cooled to room temperature, remove precipitation after filtration and obtain filtrate, again by filtrate after Vacuum Concentration and spraying dry, namely obtained a kind of activated peptide forage additive finished product utilizing aquatic product leftovers to prepare of the present invention.
Embodiment 2: by slant preservation can produce subtilin bacillus subtilis (
bacillus subtilis) in the LB fluid nutrient medium that cools after being linked into sterilizing of HL-1, in 37 DEG C, 150r/min cultivates 16h, be prepared into bacillus subtilis (
bacillus subtilis) HL-1 seed liquor is for subsequent use.It is for subsequent use after fresh shrimp head is rubbed.By 15 kilograms rub shrimp heads and 100 kg of water join in fermentation tank, and add 900 grams of glucose, 300 grams of frerrous chlorides mix together with 550 grams of dipotassium hydrogen phosphates, obtain mixing after fermentation medium; Fermentation cylinder for fermentation is cultivated based on 115 DEG C of sterilizing 20min; Then add in the fermentation raw material cooled after sterilizing 2 kilograms of bacillus subtilises (
bacillus subtilis) HL-1 seed liquor; Fermentation temperature is 28 DEG C, and filtrated air intake is 0.5vvm ~ 1.5vvm, and the rotating speed of fermentation tank is 150rpm ~ 250rpm, and fermentation time is 45h; Fermentation ends after fermentation liquid is warming up to 90 DEG C of enzyme 5min that go out, after the zymotic fluid after enzyme that goes out is cooled to room temperature, remove precipitation after filtration and obtain filtrate, again by filtrate after Vacuum Concentration and spraying dry, namely obtained a kind of activated peptide forage additive finished product utilizing aquatic product leftovers to prepare of the present invention.
Embodiment 3: by slant preservation can produce subtilin bacillus subtilis (
bacillus subtilis) in the LB fluid nutrient medium that cools after being linked into sterilizing of HL-1, in 37 DEG C, 150r/min cultivates 16h, be prepared into bacillus subtilis (
bacillus subtilis) HL-1 seed liquor is for subsequent use.It is for subsequent use after fresh scallop body is rubbed.By 20 kilograms rub scallop bodies and 100 kg of water join in fermentation tank, and add 600 grams of glucose, 300 grams of frerrous chlorides mix together with 550 grams of dipotassium hydrogen phosphates, obtain mixing after fermentation medium; Fermentation cylinder for fermentation is cultivated based on 115 DEG C of sterilizing 20min; Then add in the fermentation raw material cooled after sterilizing 2 kilograms of bacillus subtilises (
bacillus subtilis) HL-1 seed liquor; Fermentation temperature is 29 DEG C, and filtrated air intake is 1.0vvm ~ 2.0vvm, and the rotating speed of fermentation tank is 180rpm ~ 280rpm, and fermentation time is 40h; Fermentation ends after fermentation liquid is warming up to 90 DEG C of enzyme 5min that go out, after the zymotic fluid after enzyme that goes out is cooled to room temperature, remove precipitation after filtration and obtain filtrate, again by filtrate after Vacuum Concentration and spraying dry, namely obtained a kind of activated peptide forage additive finished product utilizing aquatic product leftovers to prepare of the present invention.
Claims (7)
1. utilize aquatic product leftovers to prepare a method for activated peptide forage additive, it is characterized in that: the method comprises the steps:
(1) by slant preservation can produce subtilin bacillus subtilis (
bacillus subtilis) in the LB fluid nutrient medium that cools after being linked into sterilizing of HL-1, in 37 DEG C, 150r/min cultivates 16h, be prepared into bacillus subtilis (
bacillus subtilis) HL-1 seed liquor is for subsequent use;
(2) for subsequent use after fresh aquatic product leftovers being rubbed;
(3) aquatic product leftovers of rubbing obtained for step (2) is put in fermentation tank, and add water and nutrients in fermentation tank, after mixing, obtain fermentation medium;
(4) fermented and cultured in fermentation tank step (3) obtained is based on 115 DEG C of sterilizing 20min;
(5) fermentation medium after sterilizing step (4) obtained is cooled to room temperature, then access bacillus subtilis prepared by step (1) (
bacillus subtilis) HL-1 seed liquor;
(6) control of sweat: fermentation temperature is 28 DEG C ~ 30 DEG C, and filtrated air intake is 0.5vvm ~ 2.5vvm, the rotating speed of fermentation tank is 150rpm ~ 350rpm, and fermentation time is 35h ~ 45h;
(7) product handling system: fermentation ends after fermentation liquid is warming up to 90 DEG C of insulations and goes out enzyme 5min; After the zymotic fluid after enzyme that goes out is cooled to room temperature, removes precipitation after filtration and obtain filtrate; Filtrate obtains activated peptide forage additive finished product again after Vacuum Concentration and spraying dry.
2. a kind of method utilizing aquatic product leftovers to prepare activated peptide forage additive according to claim 1, is characterized in that: bacillus subtilis used in described step (1) (
bacillus subtilis) HL-1 is CGMCC No.10379 at the deposit number at China Committee for Culture Collection of Microorganisms's common micro-organisms center.
3. a kind of method utilizing aquatic product leftovers to prepare activated peptide forage additive according to claim 1, is characterized in that: the aquatic product leftovers in described step (2) is shrimp head, scallop body, the leftovers of tilapia removing internal organ and fish-skin, the cod leftover bits and pieces removing internal organ and fish-skin, the carp leftover bits and pieces removing internal organ and fish-skin, the black carp leftover bits and pieces removing internal organ and fish-skin, the silver carp leftover bits and pieces removing internal organ and fish-skin, the grass carp leftover bits and pieces removing internal organ and fish-skin and the one removed in the stripped tuna leftover bits and pieces of internal organ and fish-skin.
4. a kind of method utilizing aquatic product leftovers to prepare activated peptide forage additive according to claim 1, it is characterized in that: in described step (3), the addition of aquatic product leftovers accounts for water gross mass percentages with the quality of the aquatic product leftovers added, and is 10% ~ 20%.
5. a kind of method utilizing aquatic product leftovers to prepare activated peptide forage additive according to claim 1, is characterized in that: the mixture that described step (3) Middle nutrition thing is glucose, frerrous chloride and dipotassium hydrogen phosphate form.
6. a kind of method utilizing aquatic product leftovers to prepare activated peptide forage additive according to claim 1, it is characterized in that: the nutraceutical addition in described step (3) accounts for water gross mass percentages with the nutraceutical quality of adding, be respectively: glucose 0.01% ~ 5%, frerrous chloride 0.01% ~ 5%, dipotassium hydrogen phosphate 0.01% ~ 5%.
7. a kind of method utilizing aquatic product leftovers to prepare activated peptide forage additive according to claim 1, is characterized in that: the middle bacillus subtilis of described step (5) (
bacillus subtilis) inoculum concentration of HL-1 seed liquor accounts for fermentation medium gross mass percentages with the quality of seed liquor of access, is 0.1% ~ 5%.
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Cited By (3)
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|---|---|---|---|---|
| CN105231000A (en) * | 2015-09-25 | 2016-01-13 | 广东海洋大学 | Method for producing compound antibacterial nutrient feed additive by utilizing offal of aquatic products |
| CN109123726A (en) * | 2018-10-08 | 2019-01-04 | 何秋宏 | A kind of preparation method of feed fermentation fish meal |
| CN109953176A (en) * | 2017-12-23 | 2019-07-02 | 青岛麦迪尔生物工程有限公司 | A method of peptone is prepared using aquatic products processing leftover bits and pieces |
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Cited By (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN105231000A (en) * | 2015-09-25 | 2016-01-13 | 广东海洋大学 | Method for producing compound antibacterial nutrient feed additive by utilizing offal of aquatic products |
| CN105231000B (en) * | 2015-09-25 | 2020-01-07 | 广东海洋大学 | Method for producing antibacterial nutritional compound feed additive by using aquatic product leftovers |
| CN109953176A (en) * | 2017-12-23 | 2019-07-02 | 青岛麦迪尔生物工程有限公司 | A method of peptone is prepared using aquatic products processing leftover bits and pieces |
| CN109123726A (en) * | 2018-10-08 | 2019-01-04 | 何秋宏 | A kind of preparation method of feed fermentation fish meal |
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Effective date of registration: 20200131 Address after: 524394 Gas Station on 374 Provincial Road, Yangqing Town, Suixi County, Zhanjiang City, Guangdong Province Patentee after: Zhanjiang silver Biotechnology Co., Ltd. Address before: 524088 Guangdong city of Zhanjiang province sea Mazhang District Road No. 1 Patentee before: Guangdong Ocean University |