CN111165652A - Fermented feed for river crabs as well as preparation method and application of fermented feed - Google Patents
Fermented feed for river crabs as well as preparation method and application of fermented feed Download PDFInfo
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- CN111165652A CN111165652A CN202010059865.0A CN202010059865A CN111165652A CN 111165652 A CN111165652 A CN 111165652A CN 202010059865 A CN202010059865 A CN 202010059865A CN 111165652 A CN111165652 A CN 111165652A
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- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23K—FODDER
- A23K10/00—Animal feeding-stuffs
- A23K10/10—Animal feeding-stuffs obtained by microbiological or biochemical processes
- A23K10/12—Animal feeding-stuffs obtained by microbiological or biochemical processes by fermentation of natural products, e.g. of vegetable material, animal waste material or biomass
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- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01K—ANIMAL HUSBANDRY; CARE OF BIRDS, FISHES, INSECTS; FISHING; REARING OR BREEDING ANIMALS, NOT OTHERWISE PROVIDED FOR; NEW BREEDS OF ANIMALS
- A01K61/00—Culture of aquatic animals
- A01K61/50—Culture of aquatic animals of shellfish
- A01K61/59—Culture of aquatic animals of shellfish of crustaceans, e.g. lobsters or shrimps
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- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23K—FODDER
- A23K10/00—Animal feeding-stuffs
- A23K10/20—Animal feeding-stuffs from material of animal origin
-
- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23K—FODDER
- A23K10/00—Animal feeding-stuffs
- A23K10/20—Animal feeding-stuffs from material of animal origin
- A23K10/26—Animal feeding-stuffs from material of animal origin from waste material, e.g. feathers, bones or skin
-
- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23K—FODDER
- A23K10/00—Animal feeding-stuffs
- A23K10/30—Animal feeding-stuffs from material of plant origin, e.g. roots, seeds or hay; from material of fungal origin, e.g. mushrooms
- A23K10/33—Animal feeding-stuffs from material of plant origin, e.g. roots, seeds or hay; from material of fungal origin, e.g. mushrooms from molasses
-
- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23K—FODDER
- A23K10/00—Animal feeding-stuffs
- A23K10/30—Animal feeding-stuffs from material of plant origin, e.g. roots, seeds or hay; from material of fungal origin, e.g. mushrooms
- A23K10/37—Animal feeding-stuffs from material of plant origin, e.g. roots, seeds or hay; from material of fungal origin, e.g. mushrooms from waste material
-
- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23K—FODDER
- A23K20/00—Accessory food factors for animal feeding-stuffs
- A23K20/20—Inorganic substances, e.g. oligoelements
- A23K20/30—Oligoelements
-
- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23K—FODDER
- A23K50/00—Feeding-stuffs specially adapted for particular animals
- A23K50/80—Feeding-stuffs specially adapted for particular animals for aquatic animals, e.g. fish, crustaceans or molluscs
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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
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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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- 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
- Y02P—CLIMATE CHANGE MITIGATION TECHNOLOGIES IN THE PRODUCTION OR PROCESSING OF GOODS
- Y02P60/00—Technologies relating to agriculture, livestock or agroalimentary industries
- Y02P60/80—Food processing, e.g. use of renewable energies or variable speed drives in handling, conveying or stacking
- Y02P60/87—Re-use of by-products of food processing for fodder production
Abstract
The invention discloses a fermented feed for river crabs as well as a preparation method and application thereof. The invention discloses a preparation method of fermented feed for river crabs, which comprises the following steps: inoculating a fermenting agent into the fermentation raw materials to obtain a fermentation system, and fermenting the fermentation system to obtain a fermentation product, namely the fermented feed for the river crabs; the fermentation raw materials comprise cottonseed meal, rapeseed meal, soybean meal, wheat bran, earthworm powder and shrimp shell powder; the active ingredients of the leaven consist of bacillus subtilis, bacillus pumilus and rhodotorula. After the fermented feed for the river crabs is used for feeding the river crabs, the survival rate, the average weight of live crab powder, the bait feeding amount and the relative growth rate are obviously increased, the hepatopancreas index of the fed river crabs is reduced, and the content of muscle amino acid is improved. The fermented feed for river crabs has good nutritive value, can improve the commodity value and the culture benefit of river crabs, and has wide application prospect.
Description
Technical Field
The invention relates to the field of biotechnology, and discloses a fermented feed for river crabs as well as a preparation method and application thereof.
Background
River crabs, also called 'crabs' and 'hairy crabs', are delicious in meat quality, contain rich proteins and trace elements, and are traditional cate of residents in freshwater watershed in China. The growth speed of the river crabs can be improved and the quality of the river crabs can be improved by properly increasing the proportion of animal protein feed in the mature period of the river crabs, the animal protein feed of a plurality of river crabs in the market at present mostly adopts fresh or frozen small trash fish or fish meal as main raw materials, however, the ice fresh small trash fish is fed in a large amount, so that the disease of the crabs is easy to frequently occur, and the water quality is easy to be polluted; in addition, the price of animal protein feed, especially fish meal, is high, and the use of the fish meal in a large proportion in the river crab culture causes the culture cost to be increased, thereby influencing the culture benefit; when the water resistance of the river crab feed is poor, the river crab feed is easy to scatter in water, cannot be ingested by river crabs and is easy to pollute the water quality. The search for an alternative resource of animal protein feed is a necessary development trend.
It is necessary to provide a feed which can replace the feeding proportion of river crab animal protein feed, especially fish meal, and simultaneously improve the water resistance of the river crab pellet feed, so that the river crab pellet feed can be better ingested by the river crab, the feed utilization rate is improved, the survival rate and the meat quality of the river crab are improved, and the breeding benefit is improved.
Disclosure of Invention
The invention provides a fermented feed which can replace animal protein feed in river crab feed, can improve the water resistance of river crab pellet feed, enables the river crab pellet feed to be better eaten by river crabs, improves the utilization rate of the feed, and can improve the production performance and the nutritional value of the river crabs after being fed.
In order to solve the technical problems, the invention firstly provides a preparation method of the fermented feed for river crabs, which comprises the following steps: inoculating a fermenting agent into the fermentation raw materials to obtain a fermentation system, and fermenting the fermentation system to obtain a fermentation product, namely the fermented feed for the river crabs; the fermentation raw materials comprise cottonseed meal, rapeseed meal, soybean meal, wheat bran, earthworm powder and shrimp shell powder;
the active ingredients of the leavening agent consist of Bacillus subtilis, Bacillus pumilus and Rhodotorula sp.
In the above method, the bacillus subtilis may be bacillus subtilis BFC 1601.
The Bacillus pumilus can be Bacillus pumilus BFEC 180101.
The rhodotorula may be rhodotorula BFC 170301.
In the above method, the leavening agent may be composed of the bacillus subtilis, the bacillus pumilus, the rhodotorula and a carrier, and the carrier may be rice hull powder or corn cob powder.
The content of viable count of Bacillus subtilis in the starter is 1 × 10 or more9CFU/g, the viable count content of the bacillus pumilus can be more than or equal to 2 multiplied by 1010CFU/g, the viable count content of the rhodotorula is more than or equal to 1 multiplied by 1010CFU/g。
Specifically, the leavening agent can be obtained according to a method comprising the following steps: culturing the bacillus subtilis to obtain a bacillus subtilis suspension, culturing the bacillus pumilus to obtain a bacillus pumilus suspension, culturing the rhodotorula to obtain a rhodotorula suspension, and mixing the bacillus subtilis suspension, the bacillus pumilus suspension and the rhodotorula suspension to obtain a composite thallus mixture; and mixing the composite thallus mixture with the carrier, and then drying at low temperature to obtain the leavening agent.
The ratio of viable count of the bacillus subtilis, the bacillus pumilus and the rhodotorula in the composite thallus mixture can be (3-6) CFU: (2-5) CFU: (2-5) CFU, specifically 4 CFU: 4 CFU: 2 CFU.
The mass ratio of the composite thallus mixture to the carrier when the composite thallus mixture is mixed with the carrier can be 3-5: 6-10. The mass ratio of the composite thallus mixture to the carrier can be 4: 8.
The method may further comprise: replacing the liquid in the bacillus subtilis suspension with physiological saline or phosphate buffer solution with pH6.8, and/or replacing the liquid in the bacillus pumilus suspension with physiological saline or phosphate buffer solution with pH6.8, and/or replacing the liquid in the rhodotorula rubra suspension with physiological saline or phosphate buffer solution with pH 6.8.
In the above method, the inoculating a fermentation agent into the fermentation raw material to obtain a fermentation system may include: activating the leavening agent to obtain a fermented seed liquid; and inoculating the fermentation seed liquid into the fermentation raw material to obtain the fermentation system.
Activating the starter comprises adding the starter to water in a mass ratio of 1:50 for activation. The activation may be carried out at 32 ℃. The activation time may be 12-24h, such as 12 h. The activation may be performed in a seed activation tank.
The inoculation amount of the fermentation seed liquid in the fermentation system meets the following requirements: the mass ratio of the leavening agent to the fermentation raw material is (0.1-0.3): 100, such as 0.25: 100.
In the method, the fermentation raw material can be composed of cottonseed meal, rapeseed meal, soybean meal, wheat bran, earthworm powder, shrimp shell powder, river crab trace element premixed feed and molasses. The mass percentages of the cottonseed meal, the rapeseed meal, the soybean meal, the wheat bran, the earthworm powder, the shrimp shell powder, the river crab microelement pre-mixed feed and the molasses in the fermentation raw materials can be 5-20% of the cottonseed meal, 5-20% of the rapeseed meal, 20-54% of the soybean meal, 10-30% of the wheat bran, 2-10% of the earthworm powder, 2-10% of the shrimp shell powder, 1-2% of the river crab microelement pre-mixed feed and 1-5% of the molasses. The mass percentages of the cottonseed meal, the rapeseed meal, the soybean meal, the wheat bran, the earthworm powder, the shrimp shell powder, the river crab microelement premix feed and the molasses in the fermentation raw materials can be 15% of the cottonseed meal, 10% of the rapeseed meal, 45% of the soybean meal, 15% of the wheat bran, 5% of the earthworm powder, 5% of the shrimp shell powder, 1.5% of the river crab microelement premix feed and 3.5% of the molasses.
The earthworm powder is a powdery product prepared by drying earthworms, the content of crude protein is more than or equal to 55 percent, and the water content is less than or equal to 8 percent. In one embodiment of the present invention, the earthworm powder is a product of zhengzhou puro animal pharmaceutical industry ltd.
The shrimp shell powder is a powdery product prepared by drying inedible residues (such as shrimp heads, shells, tail brains and the like) left in shrimp processing, and has a crude protein content of more than or equal to 35 percent and a water content of less than or equal to 10 percent. In one embodiment of the invention, the shrimp shell meal is a product of Qingdao Jinlong fodder Co.
Wherein, the trace element content in the river crab trace element pre-mixed feed is as follows: more than or equal to 6500mg/kg of potassium, 2600-12000mg/kg of sodium, more than or equal to 740mg/kg of magnesium, 900-75000mg/kg of iron, 4700-15000mg/kg of zinc, 1300-10000mg/kg of manganese, 340-2500mg/kg of copper, 125-200mg/kg of cobalt, 160-2000mg/kg of iodine and 32-50mg/kg of selenium.
In one embodiment of the invention, the river crab microelement premix feed is a product of Biotech limited of Beijing Coencho.
The water content of the fermentation system can be 30-35% (mass percent).
In the above method, the fermentation may be carried out at 25 to 35 ℃ (e.g., 32 ℃). The fermentation time can be 3-10 days (such as 5 days).
The fermented feed for river crabs prepared by the preparation method of the fermented feed for river crabs or the feed containing the fermented feed for river crabs also belongs to the protection scope of the invention.
The invention also provides a kit of parts having any of the following functions M1) -M7), said product comprising said fermentation feedstock and/or said starter culture:
m1) preparing products for improving the production performance of the river crabs;
m2) preparing a product for promoting the growth of river crabs;
m3) preparing a product for improving the survival rate of the river crabs;
m4) preparing a product for improving the body immunity of the river crabs;
m5) preparing products for reducing the hepatopancreas index of the river crabs;
m6) preparing products for improving the amino acid content of the river crabs;
m7) preparing the fermented feed for the river crabs.
The kit may consist of the fermentation raw material alone, the fermentation agent alone, or both the fermentation raw material and the fermentation agent.
Each substance in the kit can be packaged independently.
The invention also provides the following applications of I or II:
I. the fermented feed for the river crabs can be applied to any one of the following applications:
A1) improving the production performance of the river crabs;
A2) preparing products for improving the production performance of the river crabs;
A3) promoting the growth of river crabs;
A4) preparing a product for promoting the growth of river crabs;
A5) improving the survival rate of the river crabs;
A6) preparing a product for improving the survival rate of the river crabs;
A7) improving the organism immunity of the river crabs;
A8) preparing a product for improving the body immunity of the river crabs;
A9) reducing the hepatopancreas index of the river crab;
A10) preparing a product for reducing the hepatopancreas index of the river crabs;
A11) the amino acid content of the river crab is improved;
A12) preparing products for improving the amino acid content of the river crabs;
II. Any of the following uses of the kit:
A1) improving the production performance of the river crabs;
A2) preparing products for improving the production performance of the river crabs;
A3) promoting the growth of river crabs;
A4) preparing a product for promoting the growth of river crabs;
A5) improving the survival rate of the river crabs;
A6) preparing a product for improving the survival rate of the river crabs;
A7) improving the organism immunity of the river crabs;
A8) preparing a product for improving the body immunity of the river crabs;
A9) reducing the hepatopancreas index of the river crab;
A10) preparing a product for reducing the hepatopancreas index of the river crabs;
A11) the amino acid content of the river crab is improved;
A12) preparing products for improving the amino acid content of the river crabs;
A13) preparing the fermented feed for the river crabs.
In the application, the product can be a feed additive, a premix, a concentrated feed or a complete feed.
The invention also provides a feeding method of the river crabs, which comprises the following steps: and (3) taking the fermented feed for the river crabs as a protein raw material to replace all or part of fish meal and/or soybean meal in the basic daily ration to prepare the daily ration of the river crabs, and feeding the river crabs by using the daily ration of the river crabs.
In the method, the fermented feed for river crabs in the river crab daily ration can be 20-50 percent, such as 30 percent in percentage by mass.
In one embodiment of the invention, the daily feed for the river crabs comprises the following components in percentage by mass: 30% of fermented feed for river crabs, 10% of fish meal, 30% of soybean meal, 5% of rapeseed meal, 5% of cottonseed meal, 15% of wheat middling and 5% of composite premixed feed. The compound premix feed is a product of Dismann vitamin (Shandong) Co.
In the invention, the step of increasing the amino acid content of the river crab can be specifically increasing the total amino acid content of the muscle of the river crab.
Experiments prove that the fermented feed for river crabs has high protein content, contains functional probiotic components, degrades high molecular protein into small peptides, improves the nutritive value, has high stability in water and low dissolution rate, and can replace high-protein raw materials such as fish meal, bean pulp and the like to be used for feeding river crabs. After the fermented feed for river crabs is used for feeding river crabs, the survival rate, the average weight of live crab powder, the bait feeding amount and the relative growth rate are all obviously increased, and the hepatopancreas index of the fed river crabs is reduced, and the content of muscle amino acid is improved. The fermented feed for river crabs has good nutritive value, can improve the commodity value and the culture benefit of river crabs, and has wide application prospect.
Biological material preservation instructions
Classification nomenclature of biological materials: bacillus subtilis (Bacillus subtilis)
Strain number of biological material: BFC1601
Deposit name of biological material: china general microbiological culture Collection center
The preservation unit of the biological material is abbreviated as: CGMCC (China general microbiological culture Collection center)
Deposit unit address of biological material: west road No.1, north west of the township, beijing, ministry of sciences, china, institute of microbiology, zip code: 100101
Preservation date of biological material: 2016 (10 months) and 21 (days)
Accession number to the collection of biological materials: CGMCC No.13131
Biological material preservation instructions
Classification nomenclature of biological materials: bacillus pumilus (Bacillus pumilus)
Strain number of biological material: BFEC180101
Deposit name of biological material: china general microbiological culture Collection center
The preservation unit of the biological material is abbreviated as: CGMCC (China general microbiological culture Collection center)
Deposit unit address of biological material: west road No.1, north west of the township, beijing, ministry of sciences, china, institute of microbiology, zip code: 100101
Preservation date of biological material: 7 month and 5 days 2018
Accession number to the collection of biological materials: CGMCC No.16061
Biological material preservation instructions
Classification nomenclature of biological materials: rhodotorula sp (Rhodotorula sp.)
Strain number of biological material: BFC170301
Deposit name of biological material: china general microbiological culture Collection center
The preservation unit of the biological material is abbreviated as: CGMCC (China general microbiological culture Collection center)
Deposit unit address of biological material: west road No.1, north west of the township, beijing, ministry of sciences, china, institute of microbiology, zip code: 100101
Preservation date of biological material: 26/12/2017
Accession number to the collection of biological materials: CGMCC No.15125
Detailed Description
The present invention is described in further detail below with reference to specific embodiments, which are given for the purpose of illustration only and are not intended to limit the scope of the invention. The experimental procedures in the following examples are conventional unless otherwise specified. Materials, reagents, instruments and the like used in the following examples are commercially available unless otherwise specified. The quantitative tests in the following examples, all set up three replicates and the results averaged.
The earthworm powder is a powdery product prepared by drying earthworms, the content of crude protein is more than or equal to 55 percent, and the water content is less than or equal to 8 percent. The earthworm powder used in the following examples was Zhengzhou puro animal pharmaceutical Co., Ltd.
The shrimp shell powder is residue (shrimp head, shell, tail brain, etc.) left after processing shrimp, and is dried to obtain powdered product with crude protein content not less than 35% and water content not more than 10%. The shrimp shell powder used in the following examples is a product of Qingdao Jinlong fodder Co.
The river crab trace element premix feed comprises: the trace element pre-mixed feed for river crab contains potassium not less than 6500mg, sodium 2600-12000mg, magnesium not less than 740mg, iron 900-75000mg, zinc 4700-15000mg, manganese 1300-10000mg, copper 340-2500mg, cobalt 125-200mg, iodine 160-2000mg and selenium 32-50 mg. The river crab microelement pre-mixed feed used in the following examples is a product of Biotech limited of Beijing Coencho.
Example 1 preparation of fermented feed for river crab and its use in improving production performance of river crab
Preparation of fermented feed for river crabs
1. Preparation of leavening agent
1) Preparation of Bacillus subtilis suspension
B, bacillus subtilis strain: bacillus subtilis BFC1601 is preserved in China general microbiological culture Collection center with the preservation number of CGMCC NO. 13131.
Inoculating the activated bacillus subtilis BFC1601 into a 500ml triangular flask filled with 100ml of fermentation medium, standing and culturing at 33 ℃ for 24 hours, and collecting fermentation liquor to obtain bacillus subtilis seed liquid.
Inoculating the bacillus subtilis seed solution into a 500L fermentation tank filled with 200L fermentation medium according to the inoculation amount of 6% (volume ratio), culturing for 24 hours at 33 ℃ under the condition that the stirring speed is 100rpm, and collecting fermentation liquor; and centrifuging the fermentation liquor to collect wet bacteria, and resuspending the bacteria by using normal saline or phosphate buffer solution with pH of 6.8 to obtain the bacillus subtilis suspension.
Each liter of fermentation medium is prepared according to the following method: 10g glucose, 10g peptone, 1.5g KH2PO4，0.80gMnSO4，1.00g MgSO41.0g of NaCl, and the volume is constant to l000 mL by using distilled water, and the pH value is 6.5; sterilizing at 121 deg.C for 20 min.
2) Preparation of Bacillus pumilus suspension
Bacillus pumilus strains: bacillus pumilus (Bacillus pumilus) BFEC180101 is preserved in China general microbiological culture Collection center with the preservation number of CGMCC NO. 16061.
Inoculating the activated bacillus pumilus BFEC180101 into a 500ml triangular flask filled with 100ml of fermentation medium, standing and culturing at 33 ℃ for 24 hours, and collecting fermentation liquor to obtain bacillus pumilus seed liquid.
Inoculating Bacillus pumilus seed solution into 500L fermentation tank containing 200L fermentation medium at 6% (volume ratio), culturing at 33 deg.C and stirring speed of 100rpm for 24 hr, and collecting fermentation liquid; and centrifuging the fermentation liquor to collect wet bacteria, and resuspending the bacteria by using normal saline or phosphate buffer solution with pH of 6.8 to obtain the bacillus pumilus suspension.
Each liter of fermentation medium is prepared according to the following method: 10g glucose, 10g peptone, 1.5g KH2PO4，0.80gMnSO4，1.00g MgSO41.0g of NaCl, and the volume is constant to l000 mL by using distilled water, and the pH value is 6.5; sterilizing at 121 deg.C for 20 min.
3) Preparation of Rhodotorula suspension
Rhodotorula strain: rhodotorula sp BFC170301 is preserved in China general microbiological culture Collection center with the preservation number of CGMCC NO. 15125.
Inoculating activated rhodotorula glutinis BFC170301 into a 500ml triangular flask containing 100ml of fermentation medium, standing and culturing at 33 ℃ for 24 hours, and collecting fermentation liquor to obtain rhodotorula glutinis seed liquid.
Inoculating Rhodotorula glutinis seed solution into 500L fermentation tank containing 200L fermentation medium according to inoculation amount of 6% (volume ratio), culturing at 33 deg.C and stirring speed of 100rpm for 24 hr, and collecting fermentation liquid; and centrifuging the fermentation liquor to collect wet bacteria, and resuspending the bacteria by using normal saline or phosphate buffer solution with pH of 6.8 to obtain the rhodotorula suspension.
Each liter of fermentation medium is prepared according to the following method: 10g glucose, 10g peptone, 1.5g KH2PO4，0.80gMnSO4，1.00g MgSO41.0g of NaCl, and the volume is constant to l000 mL by using distilled water, and the pH value is 6.5; sterilizing at 121 deg.C for 20 min.
4) Preparation of leavening agent
Mixing the bacillus subtilis suspension, the bacillus pumilus suspension and the rhodotorula rubra suspension obtained in the steps 1) to 3) to obtain a composite thallus mixture, wherein the number of viable bacteria of the bacillus subtilis, the bacillus pumilus and the rhodotorula rubra in the composite thallus mixture is 4:4: 2; mixing the composite thallus mixture with carrier (rice hull powder or corn cob powder), and drying at low temperature to obtain starter, wherein the viable count of Bacillus subtilis is not less than 1.0 × 109CFU/g, the number of viable bacillus pumilus is more than or equal to 2.0 multiplied by 1010CFU/g, the number of live rhodotorula is more than or equal to 1 multiplied by 1010CFU/g, and the balance of carrier.The low-temperature drying condition is that the drying is carried out at 65 ℃ until the moisture content is below 15 percent.
2. Preparation of fermented feed for river crabs
Adding the leavening agent obtained in the step 1 into clean water according to the mass ratio of 1:50, and activating for 12 hours in a strain activation tank at 32 ℃ to serve as fermentation seed liquid for later use;
crushing cottonseed meal, rapeseed meal, bean pulp and wheat bran, screening the crushed cottonseed meal, rapeseed meal, bean pulp, wheat bran, earthworm powder, shrimp shell powder and river crab trace element premixed feed for later use by using a 1.5mm pore size sieve, premixing the crushed cottonseed meal, rapeseed meal, bean pulp, wheat bran, earthworm powder, shrimp shell powder and river crab trace element premixed feed for 2-5 minutes according to a certain proportion to obtain a mixed meal mixture, wherein the mass ratio of the cottonseed meal, the rapeseed meal, the bean pulp, the wheat bran, the earthworm powder, the shrimp shell powder and the river crab trace element premixed feed in the mixed meal mixture is 15: 10: 45: 15: 5: 5: 1.5; then transferring the mixed meal mixture into a sterilization tank, sterilizing for 30min at 121 ℃ by high-pressure steam, and standing to 30 ℃ for later use; sequentially adding clean water, molasses and fermentation seed liquid into a liquid preparation tank, and uniformly mixing for 0.5 minute to serve as a bacterial liquid for later use; spraying the bacterial liquid into the sterilized and stirred mixed meal mixture to obtain a fermentation system, wherein the amount of the molasses, the fermentation seed liquid and the mixed meal mixture in the added bacterial liquid meets the mass ratio of the molasses, the starter and the mixed meal mixture in the bacterial liquid of 3.5:0.25:96.5, and the water content in the fermentation system is 30% -35%; continuously stirring for 2-5min after the bacterial liquid is added; discharging, fermenting for 5 days at 32 deg.C, and turning and throwing for 1-2 times per day; the fermentation product obtained after the fermentation is the fresh-base fermented feed for the river crabs; or further drying the fermented feed for river crabs in a fresh base state at 85-135 ℃ until the water content is about 20 percent, and then drying the fermented feed for river crabs at the temperature of below 65 ℃ until the weight is constant (the water content is about 15 percent) to obtain the fermented feed for river crabs in a dry base state, wherein the obtained fermented feed for river crabs has special fermentation fragrance.
Wherein, the cottonseed meal, rapeseed meal, soybean meal, wheat bran, earthworm powder, shrimp shell meal, the river crab microelement pre-mixed feed and molasses constitute the fermentation raw materials for preparing the fermented feed for the river crabs.
Preparation of other fermented feeds
Firstly) preparation of fermented feed A
1. Preparation of leavening agent
1) Preparation of Bacillus subtilis suspension
Replacing the bacillus subtilis BFC1601 by bacillus subtilis (Tianjin Bo Fei Germany Co., Ltd., product number 920) according to the method of 1) in the step one to obtain a bacillus subtilis suspension.
2) Preparation of Bacillus pumilus suspension
The same as step 2) of step 1 in step one).
3) Preparation of Rhodotorula suspension
Same as step 1 in step one, step 3).
4) Preparation of leavening agent
Same as step 1 in step one, 4).
2. Preparation of fermented feed A
And (3) recording the obtained fermented feed for the river crabs as a fermented feed A in the step 2 in the same step.
II) preparation of fermented feed B
1. Preparation of leavening agent
1) Preparation of Bacillus subtilis suspension
Same as step 1 in step one).
2) Preparation of Bacillus pumilus suspension
The same as step 2) of step 1 in step one).
3) Preparation of Rhodotorula suspension
Same as step 1 in step one, step 3).
4) Preparation of leavening agent
Mixing the bacillus subtilis suspension, the bacillus pumilus suspension and the rhodotorula rubra suspension obtained in the steps 1) to 3) to obtain a composite thallus mixture, wherein the number of viable bacteria of the bacillus subtilis, the bacillus pumilus and the rhodotorula rubra in the composite thallus mixture is 2:6: 1; mixing the composite thallus mixture with carrier (rice hull powder or corn cob powder), and drying at low temperature to obtain the starter. The low-temperature drying condition is that the drying is carried out at 65 ℃ until the moisture content is below 15 percent.
2. Preparation of fermented feed B
And (3) recording the obtained fermented feed for the river crabs as a fermented feed B in the step 2 in the same step.
Thirdly) preparation of fermented feed C
1. Preparation of leavening agent
And (3) preparing the leavening agent in the same step 1 of the step one.
2. Preparation of fermented feed
Replacing the mass ratio of the cottonseed meal, the rapeseed meal, the soybean meal, the wheat bran, the earthworm powder, the shrimp shell powder and the river crab trace element premixed feed in the mixed meal mixture by 20: 10: 45: 15: 0: 5: 1.5, keeping other steps unchanged, and recording the obtained fermented feed for the river crabs as fermented feed C.
Hereinafter, the tests were carried out using the fermented feed for river crabs and the fermented feed A-C in a dry state.
Water resistance evaluation of fermented feed for river crab
20 250mL beakers were taken, an equal amount of water was added to each beaker, and then randomly divided into a control group and test 1-4 groups, each of which had 4 beakers, to perform a water resistance evaluation test of the feed, and the test was repeated three times. Wherein, the feed comprises the following components:
the control feed comprises fish meal 20%, soybean meal 50%, rapeseed meal 5%, cotton seed meal 5%, secondary powder 15% and composite premixed feed (Dismann vitamin (Shandong) Co., Ltd.) 5%;
the feed of the group 1 in the experiment comprises 30% of the fermented feed for the river crabs obtained in the step one, 10% of fish meal, 30% of soybean meal, 5% of rapeseed meal, 5% of cottonseed meal, 15% of wheat middling and 5% of composite premixed feed;
the feed of the test 2 group comprises 30% of the fermented feed A obtained in the step two, 10% of fish meal, 30% of soybean meal, 5% of rapeseed meal, 5% of cottonseed meal, 15% of wheat middling and 5% of compound premix feed;
the feed of the test 3 group comprises 30% of the fermented feed B obtained in the step two, 10% of fish meal, 30% of soybean meal, 5% of rapeseed meal, 5% of cottonseed meal, 15% of wheat middling and 5% of compound premix feed;
the feed of test 4 group comprises 30% of fermented feed obtained in step two, 10% of fish meal, 30% of soybean meal, 5% of rapeseed meal, 5% of cottonseed meal, 15% of wheat middling and 5% of composite premixed feed. % is mass percentage.
Weighing 15g of the feeds of the control group and the test groups 1-4 respectively, putting the feeds into prepared cylindrical mesh screens (with the aperture of 1.5mm, the diameter of 5cm and the height of 7cm), soaking the feeds in beakers of corresponding groups with the water depth of 6cm, standing the beakers in water for 8 hours at normal temperature (25 ℃) for water resistance evaluation, observing the feeds once every 2 hours, slowly lifting the mesh screens from the water to the water surface, and slowly sinking the mesh screens into the water; and randomly drawing 1 sample every 2 hours in groups, draining water, and drying in an oven at 105 ℃ to constant weight to determine the feed dissolution rate.
And (4) observing results: when the interval is 2 hours, the feed granules of the control group are greatly dissolved and sink; test 1 the feed of the group is proper in stability, dissolved but not scattered and can be clamped; the feed of the group 2 in the test is slightly dissolved and can be clipped up reluctantly; test 3 groups of feed had a solubility of greater than 50% and could not be picked up; the loose of the feed of the test 4 groups is more than 50 percent and the feed can not be clamped.
When the interval is 4 hours, the control group feed is completely dissolved, and the smell of the feed is deteriorated; test 1 group of feeds swelled but did not disperse, and were fragrant in smell; experiment 2, the feed has high solubility and can clamp particles; most of the feed of the 3 groups of the test is dissolved and sunk, and no peculiar smell exists; most of the feed of experiment 4 was dissolved and settled.
When the interval is 6 hours, the feed of the control group has bad smell of dissolved water; the feed of the test 1 group still has more than 50 percent of non-dissolubility and fermented smell; test 2 the feed of most part is dissolved and sunk, and no peculiar smell exists; test 3 groups of feed were substantially completely dissolved with a slight off-flavor; the feed of test 4 groups was substantially completely dissolved with a slight off-flavour.
When the interval is 8 hours, the feed of the control group dissolves water and smells; test 1 group feed was dissolved but still had particles that could be entrapped and the water was not smelly; test 2 the feed was completely dissolved and no off-flavor was observed; test 3 groups of feed had deteriorated in flavor; test 4 groups of feeds had deteriorated in smell.
The feed dissolution rate (%) - (feed weight before non-immersion-feed weight after immersion and drying)/feed weight before non-immersion x 100%.
Table 2 results of detection of feed solution loss rate (%)
The results of the feed leaching rates are shown in Table 2. The loss rate of the feed of test 1 at each time was significantly lower than that of the control and 2-4 feeds.
The test result shows that the feed of the test 1 group shows good stability in water no matter from the aspects of solubility, smell and dissolution rate, and is obviously superior to the feed of the control group and the feed of the test 2-4 groups. The fermented feed for river crabs prepared by the invention can avoid waste and adverse effects on water quality caused by dissolution and loss of the feed and residual feed, ensure that the river crabs have longer ingestion time and reduce water quality pollution.
Influence of fermented feed for river crabs on production performance of river crabs
1. Design of test protocol
The cultivation test is carried out by 15 glass water tanks, the specification of the water tanks is 75cm long, 45cm wide and 65cm high, 155L of water is discharged into each tank, 105 male river crabs which are originally weighted by 85g are randomly divided into a control group, a test 1 group, a test 2 group, a test 3 group and a test 4 group, each group comprises 3 water tanks, and 7 male river crabs are placed into each tank. The daily ration of the control group and the daily ration of the test 1-4 groups are respectively the feed of the control group, the feed of the test 1 group, the feed of the test 2 group, the feed of the test 3 group and the feed of the test 4 group in the step three. The test period is 60 days from late 7 months to late 9 months, the river crabs are put into a box for cultivation for 1 week, the control group is fed with the daily ration, and the test is formally started by feeding each group of the daily ration after the control group is adapted.
2. Feeding management
In the test process, daily feed is respectively fed twice at 8:30 in the morning and 5:00 in the evening, the daily feed is fed at a fixed point, the number of the next feed is finely adjusted according to the standard of no residual feed intake within 30 minutes, 1/3 of water is changed every other day, and other feeding management is carried out according to the normal feeding of the river crabs. The total daily ration (bait) input, the number of dead river crabs and the weight of the dead river crabs during the raising period of each water tank are recorded during the test period. After the test is finished, weighing and calculating the final weight of the male crabs one by one, and randomly selecting 6 male crabs with similar specifications from each group for measuring the composition and nutritional indexes of three edible parts of hepatopancreas, gonads and muscles.
3. Data processing
Statistical analysis was performed on the results using the SPSS statistical software ANOVA procedure. Results are expressed as mean ± standard deviation. The significance criterion was P < 0.05.
4. Test results
The test result shows that the survival rate of the river crabs in the test 1 group is obviously higher than that of the control group and the test 2-4 groups (P is less than 0.05), and the difference between the test 2-4 groups and the control group is not obvious; the average weight of the live crabs at the end and the total weight (including dead crabs) of the live crabs in the test 1 group are obviously higher than those in the test 2 group to the test 4 group and the control group; the relative growth rate of the live crabs in the test 1 group is remarkably higher than that of the control group and the test 2 group to 4 group (P is less than 0.01), the relative growth rate of the live crabs in the test 2 group has no significant difference compared with that of the control group, and the relative growth rates of the live crabs in the test 3 group and the test 4 group are remarkably lower than that of the control group; the difference between the feed efficiency of each test group and that of the control group is not significant, but the feed efficiency of the test group 1 is reduced compared with that of the control group. In conclusion, the survival rate, the relative growth rate and the feed efficiency of the river crab fermented feed used in the test 1 group are all superior to those of the control group, and 10 percent of fish meal and 20 percent of soybean meal in the same proportion in the compound feed can be replaced in practical application. The application effects of the tests 2-4 are obvious and the effect of the test 1 is not good, which shows that the technical effect of the invention can be achieved only when the strain, the fermentation inoculum ratio and the fermentation raw materials disclosed in the invention are combined.
TABLE 3 influence of fermented feed for river crab on the Productivity
Note: the data in the same row shows significant difference (P < 0.05) in the lower case of the shoulder mark letters, significant difference (P < 0.01) in the upper case and insignificant difference (P > 0.05) in the letters with or without letters. The relative growth rate (%/day) is (live crab average weight-initial average weight)/day x 100%, and the feed efficiency is the feeding amount/(end total amount-initial weight).
When the test is finished, the edible part composition and the nutritional index content of the male crabs are detected and analyzed, and the detection method comprises the following steps:
the detection of moisture in the liver, pancreas and muscle was performed according to GB 18394-2001; total amino acid detection in the liver, pancreas and muscle was performed according to GB 5009.124-2016.
The results show that the hepatopancreas index of the male crabs in the test 1 group is obviously lower than that of the control group (P is less than 0.05), and is not obviously different from that of the test groups 2 to 4; the total amino acid content in muscle in the test 1 group is obviously higher than that in the control group and the test 2-4 groups (P is less than 0.05), and the difference between the test 2-4 groups and the control group is not obvious; the sex gland index, the moisture content of the liver and the pancreas, the total amino acid content of the liver and the pancreas and the moisture content of the muscle are not obviously different among groups. The fermented feed for river crabs disclosed by the invention is shown to reduce the hepatopancreas index of male crabs and improve the total amino acid content of the muscle of the male crabs (Table 4).
TABLE 4 influence of fermented feed for river crab on main nutrient components of edible parts of male crab
Note: the data in the same row shows significant difference (P < 0.05) in the lower case of the shoulder mark letters, significant difference (P < 0.01) in the upper case and insignificant difference (P > 0.05) in the letters with or without letters. The hepatopancreas index/%, hepatopancreas weight/body weight × 100%, and the gonadal index/%, gonadal weight/body weight × 100%.
In conclusion, the fermented feed for river crabs is used for preparing the granulated feed, so that the survival rate, the bait feeding amount, the relative growth rate and the crab weight of the river crabs can be improved, the hepatopancreas index of the river crabs is reduced, the muscle amino acid content of the river crabs is improved, and the effect of improving the commodity value and the culture benefit of the river crabs is achieved.
Claims (10)
1. The preparation method of the fermented feed for river crabs comprises the following steps: inoculating a fermenting agent into the fermentation raw materials to obtain a fermentation system, and fermenting the fermentation system to obtain a fermentation product, namely the fermented feed for the river crabs;
the fermentation raw materials comprise cottonseed meal, rapeseed meal, soybean meal, wheat bran, earthworm powder and shrimp shell powder;
the active ingredients of the leaven consist of bacillus subtilis, bacillus pumilus and rhodotorula.
2. The method of claim 1, further comprising: the bacillus subtilis is bacillus subtilis BFC 1601;
and/or the bacillus pumilus is bacillus pumilus BFEC 180101;
and/or the rhodotorula is rhodotorula BFC 170301.
3. The method according to claim 1 or 2, characterized in that: the leavening agent consists of the bacillus subtilis, the bacillus pumilus, the rhodotorula and a carrier, wherein the carrier is rice hull powder or corncob powder;
further, the number of viable bacteria of the bacillus subtilis in the leavening agent is more than or equal to 1 multiplied by 109CFU/g, the viable count of the bacillus pumilus is more than or equal to 2 multiplied by 1010CFU/g, the number of viable bacteria of the rhodotorula is more than or equal to 1 multiplied by 1010CFU/g。
4. A method according to any one of claims 1-3, characterized in that: the method for inoculating the fermenting agent into the fermentation raw material to obtain the fermentation system comprises the following steps: activating the leavening agent to obtain a fermented seed liquid; and inoculating the fermentation seed liquid into the fermentation raw material to obtain the fermentation system.
5. The method according to any one of claims 1-4, wherein: the fermentation raw materials comprise cottonseed meal, rapeseed meal, soybean meal, wheat bran, earthworm powder, shrimp shell powder, river crab trace element premixed feed and molasses.
6. A fermented feed for river crabs prepared by the method according to any one of claims 1 to 5.
7. A kit of parts having any of the following functions M1) -M7), comprising the fermentation feedstock and/or the starter culture of any of claims 1-5:
m1) preparing products for improving the production performance of the river crabs;
m2) preparing a product for promoting the growth of river crabs;
m3) preparing a product for improving the survival rate of the river crabs;
m4) preparing a product for improving the body immunity of the river crabs;
m5) preparing products for reducing the hepatopancreas index of the river crabs;
m6) preparing products for improving the amino acid content of the river crabs;
m7) preparing the fermented feed for the river crabs.
8. The following uses of I or II:
I. the fermented feed for river crabs according to claim 6, which has any one of the following applications:
A1) improving the production performance of the river crabs;
A2) preparing products for improving the production performance of the river crabs;
A3) promoting the growth of river crabs;
A4) preparing a product for promoting the growth of river crabs;
A5) improving the survival rate of the river crabs;
A6) preparing a product for improving the survival rate of the river crabs;
A7) improving the organism immunity of the river crabs;
A8) preparing a product for improving the body immunity of the river crabs;
A9) reducing the hepatopancreas index of the river crab;
A10) preparing a product for reducing the hepatopancreas index of the river crabs;
A11) the amino acid content of the river crab is improved;
A12) preparing products for improving the amino acid content of the river crabs;
II. Use of a kit according to claim 7 for any one of the following applications:
A1) improving the production performance of the river crabs;
A2) preparing products for improving the production performance of the river crabs;
A3) promoting the growth of river crabs;
A4) preparing a product for promoting the growth of river crabs;
A5) improving the survival rate of the river crabs;
A6) preparing a product for improving the survival rate of the river crabs;
A7) improving the organism immunity of the river crabs;
A8) preparing a product for improving the body immunity of the river crabs;
A9) reducing the hepatopancreas index of the river crab;
A10) preparing a product for reducing the hepatopancreas index of the river crabs;
A11) the amino acid content of the river crab is improved;
A12) preparing products for improving the amino acid content of the river crabs;
A13) preparing the fermented feed for the river crabs.
9. A method for feeding river crabs comprises the following steps: the fermented feed for river crabs according to claim 6 is used as a protein raw material to replace all or part of fish meal and/or soybean meal in a basic ration to prepare a daily ration for river crabs, and the obtained daily ration for river crabs is used for feeding river crabs to finish the feeding of the river crabs.
10. A feed comprising the fermented feed for river crabs according to claim 6.
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