CN118203070A - Fermentation method for preparing vinasse biological feed - Google Patents

Abstract

The invention discloses a fermentation method for preparing vinasse biological feed, belongs to the field of fermentation technology, and comprises the following steps: compounding brewer's yeast powder, Candida utilis powder, Bacillus subtilis powder, Aspergillus niger powder and Lactobacillus plantarum powder to obtain a composite microbial agent, stirring and mixing vinasse, bran, soybean meal and the composite microbial agent to obtain a dry material, adding water to the dry material, fermenting for 2d, stirring and adding a preservative aid thereto, and drying after constant temperature stirring to obtain vinasse biological feed. The crude protein content in the vinasse biological feed prepared by the fermentation method of the invention is significantly improved, and the crude fiber content is significantly reduced, which helps to improve the feed taste, improve the bioavailability of nutrients, reduce anti-nutritional factors in vinasse, and make protein feed safer and easier to be absorbed and utilized by animals.

Description

Fermentation method for preparing distiller's grains biological feed

Technical Field

The invention belongs to the technical field of fermentation, and in particular relates to a fermentation method for preparing distiller's grains biological feed.

Background technique

Liquor lees, as a by-product of the liquor brewing process, are derived from grains such as sorghum, barley, and wheat, and are representative of lees resources. However, the current treatment method for fresh liquor lees is mostly to dry and pile them up, which is a relatively extensive and inefficient treatment method, which not only aggravates environmental pollution, but also leads to a huge waste of resources. Therefore, it is particularly important to seek efficient and environmentally friendly treatment methods and make full use of liquor lees resources.

Fresh liquor lees contain a wide variety of nutrients and microorganisms. If they are not handled in time or handled improperly, they are prone to corruption, which in turn causes secondary environmental pollution. At the same time, it is rich in nutrients such as protein, vitamins, minerals and amino acids, as well as bioactive substances such as organic acids, phenolic acids, flavonoids and alkaloids, showing its great potential in feed development. Converting this resource-rich and nutritionally comprehensive by-product into animal feed can not only effectively alleviate the current resource shortage, but also reduce environmental pollution from the source.

However, there are some technical problems in directly using fresh distiller's grains as feed: the high fiber content, high humidity and perishable nature of fresh distiller's grains may cause reproductive tract diseases in animals, thereby affecting their reproductive capacity; at the same time, harmful components such as acetic acid and ethanol in fresh distiller's grains may cause animal poisoning and diarrhea in long-term feeding. These technical problems seriously limit the application of fresh distiller's grains in feed, resulting in low utilization of distiller's grains resources.

Therefore, in order to overcome these technical problems, it is particularly important to effectively treat fresh white wine lees. The present invention aims to provide a fermentation method for preparing wine lees bio-feed.

Summary of the invention

The invention provides a fermentation method for preparing distiller's grains biological feed, aiming to solve one of the technical problems of high fiber content, high moisture, easy decay and high acetic acid content when fresh white distiller's grains are used as feed.

The purpose of the present invention can be achieved through the following technical solutions:

The fermentation method for preparing distiller's grains bio-feed comprises the following steps:

Compounding brewer's yeast powder, Candida utilis powder, Bacillus subtilis powder, Aspergillus niger powder and Lactobacillus plantarum powder to obtain a composite microbial agent, stirring and mixing distiller's grains, bran, soybean meal and the composite microbial agent to obtain a dry material, adding water to the dry material, adjusting the humidity of the system to 50%-60%, and then fermenting at 30-37° C. for 2 days to obtain a fermentation system, stirring and adding a preservative aid to the fermentation system, and then stirring at a constant temperature of 30-37° C. for 1-2 hours to obtain a fermentation product, drying the fermentation product to a water content of 10%-15%, and after the drying process is completed, obtaining a distiller's grains biological feed;

The antiseptic additive is prepared by the following steps:

Add modified chitosan to a 1% hydrochloric acid solution, stir and mix for 10-20 minutes, adjust the pH to 5.0-5.2 with sodium hydroxide, add montmorillonite powder and stir, stir for 12 hours at room temperature, centrifuge after stirring, dry the precipitate to constant weight, and obtain a preservative additive.

Furthermore, the brewer's yeast powder, Candida utilis powder, Bacillus subtilis powder, Aspergillus niger powder and Lactobacillus plantarum powder are compounded in a mass ratio of 2:2:1:1:1.

Furthermore, the effective live bacterial counts of the brewer's yeast powder, the Candida utilis powder, the Bacillus subtilis powder, the Aspergillus niger powder and the Lactobacillus plantarum powder are all ≥1.0×10 ⁸ /g.

Furthermore, the dry material comprises, by weight, 80 parts of distiller's grains, 20-30 parts of bran, 15-20 parts of soybean meal and 0.5-1 part of composite microbial agent.

Furthermore, the amount of the preservative agent used is 2%-5% of the mass of the fermentation system.

Furthermore, the usage ratio of the hydrochloric acid solution with a volume fraction of 1%, the modified chitosan and the montmorillonite powder is 1L:15-18g:100-150g.

Furthermore, the modified chitosan is prepared by the following steps:

Step A1, adding tocopherol and succinic anhydride to anhydrous pyridine with stirring, stirring for 48 hours to obtain a product, dropping the product into a hydrochloric acid solution with a pH value of 1-2, precipitating a precipitate in the hydrochloric acid solution to form a layered system, refrigerating the layered system at 0° C. for 24 hours and then filtering to obtain a solid component, washing the solid component three times with deionized water, and then recrystallizing with ethanol, filtering, and vacuum drying the crystals to obtain carboxylated tocopherol; wherein the preparation process of carboxylated tocopherol is as follows:

Step A2, soaking chitosan in isopropanol, adding sodium hydroxide to the chitosan until the chitosan swells, alkalizing and gelatinizing at 55-60° C. for 1-2 hours, adding chloroacetic acid to the chitosan, stirring and reacting at 65-70° C. for 2-3 hours, cooling to room temperature after the reaction, adding glacial acetic acid to the chitosan to adjust the pH to neutral, filtering, washing with anhydrous ethanol and drying to obtain carboxymethyl chitosan;

Step A3, add carboxylated tocopherol to anhydrous dimethyl sulfoxide with stirring, stir for 10-20 minutes, then add N-hydroxysuccinimide and 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride thereto, cool the system to 0°C, stir at a constant temperature for 2-3 hours, then add carboxymethyl chitosan thereto with stirring, heat the system to 60-65°C, stir to react for 4-5 hours, cool to room temperature, and then filter and dry to obtain modified chitosan.

Furthermore, the tocopherol in step A1 is α-tocopherol.

Furthermore, in step A1, the usage ratio of anhydrous pyridine, tocopherol and succinic anhydride is 100 mL: 0.01 mol: 0.05-0.06 mol.

Furthermore, the deacetylation degree of the chitosan in step A2 is greater than 85%.

Furthermore, in step A2, the usage ratio of isopropanol, chitosan, sodium hydroxide and chloroacetic acid is 200 mL: 15-18 g: 22-25 g: 40-50 g.

Furthermore, the amount ratio of anhydrous dimethyl sulfoxide, carboxylated tocopherol, N-hydroxysuccinimide, 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride and carboxymethyl chitosan in step A3 is 200 mL: 0.01-0.012 mol: 0.15-0.20 mol: 0.18-0.20 mol: 1.8-2.2 g.

Beneficial effects of the present invention:

The present invention provides a fermentation method for preparing distiller's grains biological feed. The crude protein content in the distiller's grains biological feed prepared by the fermentation method of the present invention is significantly increased, and the crude fiber content is significantly reduced, which is helpful to improve the taste of the feed, increase the bioavailability of nutrients, reduce anti-nutritional factors in the distiller's grains, and make the protein feed safer and easier to be absorbed and utilized by animals.

At the same time, the fermentation method of the present invention utilizes a composite microbial agent to guide the fermentation process of distiller's grains, etc., which cleverly solves the problem of easy decay when distiller's grains are directly used as feed. Among them, the composite microbial agent includes probiotics (Candida utilis, Bacillus subtilis and Lactobacillus plantarum). The probiotics can inhibit the reproduction of harmful bacteria in the animal intestine, maintain the ecological balance of the intestine, and thus promote the development of the intestine.

Then, a homemade preservative is added in the fermentation method of the present invention. The addition of the preservative is mainly used to prevent further corruption of the fermented wine lees biological feed. The preservative prepared by the present invention is a modified chitosan-montmorillonite powder composite material. Adding the preservative prepared by the present invention has the following advantages:

(1) Anticorrosion: Chitosan itself has excellent anticorrosive properties and can reduce the spoilage rate of feed. Montmorillonite powder has good adsorption and mold removal effects and can effectively remove mycotoxins in feed. Modifying montmorillonite powder with chitosan can further reduce the toxic effects of spoiled feed on animals on the basis of achieving anticorrosion. At the same time, chitosan and montmorillonite powder are both natural and relatively low-cost additives. Through modification, the advantages of both can be combined to improve feed quality at a lower cost, thereby improving breeding efficiency. Finally, since chitosan is not easily decomposed by animal digestive enzymes, it can directly reach the intestines of animals after being eaten and finally excreted from the body. Modifying montmorillonite powder with chitosan can achieve efficient elimination of mycotoxins from the animal body after montmorillonite powder adsorbs mycotoxins, reducing the probability of mycotoxins infecting the animal body, thereby improving the safety of the final prepared distiller's grains biofeed.

(2) Buffering: The addition of montmorillonite powder can reduce the acetic acid content in the wine lees and buffer the acidity generated during fermentation. The wine lees feed produced by the present invention is mainly used as feed for ruminants (cattle, sheep, etc.). The rumen of ruminants is neutral and weakly acidic. The present invention uses montmorillonite powder to buffer the acidity in the feed and reduce the probability of animal poisoning and diarrhea caused by the feed. At the same time, montmorillonite powder itself can absorb moisture in the intestine and combine with intestinal mucus, which has a certain antidiarrheal effect.

(3) Balance: Montmorillonite powder contains some calcium, which can balance the calcium-phosphorus ratio of the final prepared distiller's grains biological feed to a certain extent. At the same time, montmorillonite powder also contains rich trace elements, which can comprehensively improve the nutritional value of the feed.

(4) Dehumidification: The present invention links carboxylated tocopherol and carboxymethyl chitosan via an amide group, and utilizes the hydrophobic structure of tocopherol to reduce the hygroscopicity of chitosan, thereby helping to reduce the hygroscopicity of the produced distiller's grains bio-feed, and helping to reduce the rate of increase in humidity when the distiller's grains bio-feed is stacked for a long time. Furthermore, the low humidity state can reduce the degree of feed corruption; at the same time, tocopherol has a certain antioxidant property, which can reduce the probability of mildew of the distiller's grains bio-feed under humid conditions.

Detailed ways

The following will be combined with the embodiments of the present invention to clearly and completely describe the technical solutions in the embodiments of the present invention. Obviously, the described embodiments are only part of the embodiments of the present invention, not all of the embodiments. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without creative work are within the scope of protection of the present invention.

The present invention provides a fermentation method for preparing distiller's grains biological feed, wherein the distiller's grains are purchased from Luzhou distiller's grains of Chengdu Xilingyuan Wine Co., Ltd. The purchased distiller's grains are the residues left after rice, wheat and sorghum are brewed, and are rich in crude protein (20.83%-24.15%) and crude fat (3.21%-4.58%), and also contain relatively high crude fiber (19.05%-22.41%) and crude ash (9.11%-10.78%). In addition, due to the large-scale reproduction and accumulation of microorganisms during the fermentation of distiller's koji, the composition and types of amino acids in the distiller's grains are relatively balanced, and the mineral content is also relatively rich.

The present invention uses the above-mentioned wine lees to develop and study a biological feed fermentation method, wherein the average crude protein content of the sampled wine lees is 22.10%; the average crude fat content is 3.55%; the average crude fiber content is 20.72%; and the average crude ash content is 9.43%.

Example 1

Preparation of modified chitosan:

Step A1, adding tocopherol (α-tocopherol) and succinic anhydride to anhydrous pyridine with stirring, stirring for 48 hours to obtain a product, dropping the product into a hydrochloric acid solution with a pH value of 1, precipitating a precipitate in the hydrochloric acid solution to form a layered system, refrigerating the layered system at 0° C. for 24 hours and then filtering to obtain a solid component, washing the solid component three times with deionized water, and then recrystallizing with ethanol, filtering, and vacuum drying the crystals to obtain carboxylated tocopherol; wherein the amount ratio of anhydrous pyridine, tocopherol and succinic anhydride is 100 mL: 0.01 mol: 0.05 mol;

Step A2, soaking chitosan (with a deacetylation degree of 90%) in isopropanol, adding sodium hydroxide to the chitosan until the chitosan swells, alkalizing and gelatinizing at 55°C for 1 hour, adding chloroacetic acid to the chitosan, stirring and reacting at 65°C for 2 hours, cooling to room temperature after the reaction, adding glacial acetic acid to the chitosan to adjust the pH to neutral, filtering, washing with anhydrous ethanol and drying to obtain carboxymethyl chitosan; wherein the amount ratio of isopropanol, chitosan, sodium hydroxide and chloroacetic acid is 200mL:15g:22g:40g;

Step A3, add carboxylated tocopherol to anhydrous dimethyl sulfoxide with stirring, stir for 10 minutes, then add N-hydroxysuccinimide (NHS) and 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride (EDC), cool the system to 0°C, stir at constant temperature for 2 hours, then add carboxymethyl chitosan with stirring, heat the system to 60°C, stir for 4 hours, cool to room temperature, filter and dry to obtain modified chitosan; wherein the amount ratio of anhydrous dimethyl sulfoxide, carboxylated tocopherol, N-hydroxysuccinimide, 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride and carboxymethyl chitosan is 200mL:0.01mol:0.15mol:0.18mol:1.8g.

Example 2

Preparation of modified chitosan:

Step A1, adding tocopherol (α-tocopherol) and succinic anhydride to anhydrous pyridine with stirring, stirring for 48 hours to obtain a product, dropping the product into a hydrochloric acid solution with a pH value of 1.5, precipitating a precipitate in the hydrochloric acid solution to form a layered system, refrigerating the layered system at 0° C. for 24 hours and then filtering to obtain a solid component, washing the solid component three times with deionized water, and then recrystallizing with ethanol, filtering, and vacuum drying the crystals to obtain carboxylated tocopherol; wherein the amount ratio of anhydrous pyridine, tocopherol and succinic anhydride is 100 mL: 0.01 mol: 0.055 mol;

Step A2, soaking chitosan (with a deacetylation degree of 85%) in isopropanol, adding sodium hydroxide to the chitosan until the chitosan swells, alkalizing and gelatinizing at 60°C for 2h, adding chloroacetic acid to the chitosan, stirring and reacting at 65°C for 3h, cooling to room temperature after the reaction, adding glacial acetic acid to the chitosan to adjust the pH to neutral, filtering, washing with anhydrous ethanol and drying to obtain carboxymethyl chitosan; wherein the amount ratio of isopropanol, chitosan, sodium hydroxide and chloroacetic acid is 200mL:17g:24g:45g;

Step A3, add carboxylated tocopherol to anhydrous dimethyl sulfoxide with stirring, stir for 20 minutes, then add N-hydroxysuccinimide (NHS) and 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride (EDC), cool the system to 0°C, stir at constant temperature for 3 hours, then add carboxymethyl chitosan with stirring, heat the system to 65°C, stir for 5 hours, cool to room temperature, filter and dry to obtain modified chitosan; wherein the amount ratio of anhydrous dimethyl sulfoxide, carboxylated tocopherol, N-hydroxysuccinimide, 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride and carboxymethyl chitosan is 200mL:0.011mol:0.18mol:0.20mol:2.0g.

Example 3

Preparation of modified chitosan:

Step A1, adding tocopherol (α-tocopherol) and succinic anhydride to anhydrous pyridine with stirring, stirring for 48 hours to obtain a product, dropping the product into a hydrochloric acid solution with a pH value of 2, precipitating a precipitate in the hydrochloric acid solution to form a layered system, refrigerating the layered system at 0° C. for 24 hours and then filtering to obtain a solid component, washing the solid component three times with deionized water, and then recrystallizing with ethanol, filtering, and vacuum drying the crystals to obtain carboxylated tocopherol; wherein the amount ratio of anhydrous pyridine, tocopherol and succinic anhydride is 100 mL: 0.01 mol: 0.06 mol;

Step A2, soaking chitosan (with a deacetylation degree of 85%) in isopropanol, adding sodium hydroxide to the chitosan until the chitosan swells, alkalizing and gelatinizing at 60°C for 2h, adding chloroacetic acid to the chitosan, stirring and reacting at 70°C for 3h, cooling to room temperature after the reaction, adding glacial acetic acid to the chitosan to adjust the pH to neutral, filtering, washing with anhydrous ethanol and drying to obtain carboxymethyl chitosan; wherein the amount ratio of isopropanol, chitosan, sodium hydroxide and chloroacetic acid is 200mL:18g:25g:50g;

Step A3, add carboxylated tocopherol to anhydrous dimethyl sulfoxide with stirring, stir for 20 minutes, then add N-hydroxysuccinimide (NHS) and 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride (EDC), cool the system to 0°C, stir at constant temperature for 3 hours, then add carboxymethyl chitosan with stirring, heat the system to 65°C, stir and react for 5 hours, cool to room temperature, filter and dry to obtain modified chitosan; wherein the amount ratio of anhydrous dimethyl sulfoxide, carboxylated tocopherol, N-hydroxysuccinimide, 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride and carboxymethyl chitosan is 200mL:0.012mol:0.20mol:0.20mol:2.2g.

Example 4

Preparation of preservative additives:

The modified chitosan prepared in Example 1 was added to a 1% hydrochloric acid solution by volume, and the mixture was stirred for 10 minutes. The pH was adjusted to 5.0 with sodium hydroxide, and montmorillonite powder (purchased from Dayu Bentonite Mining Co., Ltd., Santai County, Sichuan Province) was added thereto by stirring. The mixture was stirred for 12 hours at room temperature. After stirring, the mixture was centrifuged and the precipitate was dried to constant weight to obtain a preservative additive, wherein the amount ratio of the 1% hydrochloric acid solution by volume, the modified chitosan prepared in Example 1 and the montmorillonite powder was 1L:15g:100g.

Example 5

Preparation of preservative additives:

The modified chitosan prepared in Example 2 was added to a 1% hydrochloric acid solution by volume, and the mixture was stirred for 20 minutes. The pH was adjusted to 5.2 with sodium hydroxide, and montmorillonite powder (purchased from Dayu Bentonite Mining Co., Ltd., Santai County, Sichuan Province) was added thereto by stirring. The mixture was stirred for 12 hours at room temperature. After stirring, the mixture was centrifuged and the precipitate was dried to constant weight to obtain a preservative additive, wherein the amount ratio of the 1% hydrochloric acid solution by volume, the modified chitosan prepared in Example 2 and the montmorillonite powder was 1L:18g:100g.

Example 6

Preparation of preservative additives:

The modified chitosan prepared in Example 3 was added to a 1% hydrochloric acid solution by volume, and the mixture was stirred for 20 minutes. The pH was adjusted to 5.2 with sodium hydroxide, and montmorillonite powder (purchased from Dayu Bentonite Mining Co., Ltd., Santai County, Sichuan Province) was added thereto by stirring. The mixture was stirred for 12 hours at room temperature. After stirring, the mixture was centrifuged and the precipitate was dried to constant weight to obtain a preservative additive, wherein the amount ratio of the 1% hydrochloric acid solution by volume, the modified chitosan prepared in Example 3 and the montmorillonite powder was 1L:18g:150g.

Example 7

The fermentation method for preparing distiller's grains bio-feed comprises the following steps:

S1. Compounding saccharomyces cerevisiae powder, Candida utilis powder, Bacillus subtilis powder, Aspergillus niger powder and Lactobacillus plantarum powder in a mass ratio of 2:2:1:1:1 to obtain a composite microbial agent; wherein the effective viable counts of saccharomyces cerevisiae powder, Candida utilis powder, Bacillus subtilis powder, Aspergillus niger powder and Lactobacillus plantarum powder are all 1.0×10 ⁸ cells/g;

S2. By weight, 80 parts of distiller's grains, 20 parts of bran, 15 parts of soybean meal and 0.5 parts of the composite microbial agent prepared above were stirred and mixed to obtain dry material, water was added to the dry material, the humidity of the system was regulated to 50%, and then fermented at 30° C. for 2 days to obtain a fermentation system, and the preservative aid prepared in Example 4 was stirred and added to the fermentation system, wherein the amount of the preservative aid prepared in Example 4 was 2% of the mass of the fermentation system, and then placed at 30° C. and stirred at a constant temperature for 1 hour to obtain a fermentation product, and the fermentation product was dried to control the water content of the fermentation product to 15%. The drying treatment can simultaneously reduce the ethanol content in the fermentation product after fermentation. After the drying treatment is completed, a distiller's grains biological feed is obtained.

Example 8

The fermentation method for preparing distiller's grains bio-feed comprises the following steps:

S1. Compounding saccharomyces cerevisiae powder, Candida utilis powder, Bacillus subtilis powder, Aspergillus niger powder and Lactobacillus plantarum powder in a mass ratio of 2:2:1:1:1 to obtain a composite microbial agent; wherein the effective viable counts of saccharomyces cerevisiae powder, Candida utilis powder, Bacillus subtilis powder, Aspergillus niger powder and Lactobacillus plantarum powder are all 2.0×10 ⁸ cells/g;

S2. By weight, 80 parts of vinasse, 25 parts of bran, 20 parts of soybean meal and 1 part of the composite microbial agent prepared above were stirred and mixed to obtain dry material, water was added to the dry material, the humidity of the system was regulated to 50%, and then fermented at 35° C. for 2 days to obtain a fermentation system, and the preservative aid prepared in Example 5 was stirred and added to the fermentation system, wherein the amount of the preservative aid prepared in Example 5 was 5% of the mass of the fermentation system, and then placed at 35° C. and stirred for 2 hours at a constant temperature to obtain a fermentation product, and the fermentation product was dried to control the water content of the fermentation product to 12%. The drying treatment can simultaneously reduce the ethanol content in the fermentation product after fermentation. After the drying treatment is completed, a vinasse biological feed is obtained.

Example 9

The fermentation method for preparing distiller's grains bio-feed comprises the following steps:

S1. Compounding saccharomyces cerevisiae powder, Candida utilis powder, Bacillus subtilis powder, Aspergillus niger powder and Lactobacillus plantarum powder in a mass ratio of 2:2:1:1:1 to obtain a composite microbial agent; wherein the effective viable counts of saccharomyces cerevisiae powder, Candida utilis powder, Bacillus subtilis powder, Aspergillus niger powder and Lactobacillus plantarum powder are all 1.0×10 ⁸ cells/g;

S2. By weight, 80 parts of vinasse, 30 parts of bran, 20 parts of soybean meal and 1 part of the composite microbial agent prepared above were stirred and mixed to obtain dry material, water was added to the dry material, the humidity of the system was regulated to 60%, and then fermented at 37° C. for 2 days to obtain a fermentation system, and the preservative aid prepared in Example 6 was stirred and added to the fermentation system, wherein the amount of the preservative aid prepared in Example 6 was 4% of the mass of the fermentation system, and then placed at 37° C. and stirred for 2 hours at a constant temperature to obtain a fermentation product, and the fermentation product was dried to control the water content of the fermentation product to 10%. The drying treatment can simultaneously reduce the ethanol content in the fermentation product after fermentation. After the drying treatment is completed, a vinasse biological feed is obtained.

Comparative Example 1

Comparative Example 1 discloses a fermentation method for preparing distiller's grains bio-feed. Comparative Example 1 is the control group of Example 8. First, the modified chitosan prepared in Example 2 of Example 5 is replaced with the carboxymethyl chitosan prepared in step A2 of Example 2, and the other raw materials and the preparation process remain unchanged from Example 5 to obtain the preservative aid D1. Then, the preservative aid prepared in Example 5 of Example 8 is replaced with the preservative aid D1, and the other raw materials and the fermentation process remain unchanged from Example 8 to finally obtain the distiller's grains bio-feed.

Comparative Example 2

Comparative Example 2 discloses a fermentation method for preparing distiller's grains bio-feed. Comparative Example 2 is the control group of Example 8, in which the preservative prepared in Example 5 in Example 8 is replaced with montmorillonite powder (purchased from Dayu Bentonite Mining Co., Ltd., Santai County, Sichuan Province), and the remaining raw materials and fermentation process remain unchanged from Example 8, and finally distiller's grains bio-feed is obtained.

Comparative Example 3

Comparative Example 3 discloses a fermentation method for preparing distiller's grains bio-feed. Comparative Example 3 is the control group of Example 8, in which the preservative prepared in Example 5 of Example 8 is replaced by the modified chitosan prepared in Example 2, and the other raw materials and the fermentation process remain unchanged from Example 8, and finally the distiller's grains bio-feed is obtained.

Comparative Example 4

Comparative Example 4 is a distiller's grains raw material, namely, Luzhou distiller's grains purchased from Chengdu Xilingyuan Wine Industry Co., Ltd.

Test Example 1

The distiller's grains bio-feed prepared by the fermentation methods of Examples 7 to 9 and Comparative Examples 1 to 4 were tested. The testing process is as follows. The test results are shown in Table 1:

1. Acetic acid content detection:

The HPLC method was used to detect the acetic acid content (g/kg) in the distiller's grains bio-feed prepared by the fermentation methods of Examples 7 to 9 and Comparative Examples 1 to 4; specifically, the chromatographic conditions were: a reverse phase chromatographic column, a mobile phase of 0.02 mol/L potassium dihydrogen phosphate (pH=2.70) and methanol in a volume ratio of 97, a flow rate of 1.0 mL/min, a column temperature of 25°C, and a detection wavelength of 214 nm.

2. Anti-mildew performance test:

The vinasse bio-feed prepared by the fermentation method of Examples 7 to 9 and Comparative Examples 1 to 4 were placed under constant temperature, humidity, air permeability and light-proof conditions at 35°C and 80% humidity. Each group of Examples/Comparative Examples was placed into 5 equal piles, for a total of 35 piles. The sensory evaluation of each pile of vinasse bio-feed was performed on the 0th day, 0.5th day, 1st day, 2nd day, 4th day, 7th day, 10th day and 15th day. The evaluation adopted a 5-point system. The specific evaluation criteria are as follows. The scores of each group of Examples/Comparative Examples were calculated and the average score of the six piles was taken;

4-5 points: the feed has no color change, fermentation aroma, and no mold spots or mold blocks observed by naked eyes;

3-4 points: the color of the feed is slightly changed, there is a fermented aroma, and there are very few mold spots and no mold blocks when observed with the naked eye;

2-3 points: The color of the feed is slightly changed, with a faint sour smell. There are a few mold spots and no mold blocks when observed with the naked eye.

1-2 points: The feed is dark in color, has a strong musty smell, and has a small amount of mold spots and mold blocks when observed with the naked eye;

0-1 points: The feed is dark in color, has a strong rotten and moldy smell, and a large number of mold spots and lumps can be seen with the naked eye.

3. Nutritional component testing:

The vinasse bio-feed prepared by the fermentation methods of Examples 7 to 9 and Comparative Examples 1 to 4 was dried (dried at 105° C. to constant mass) and then tested for nutritional components:

Crude protein: Determine the crude protein content by referring to the Kjeldahl method in GB5009.5-2016.

Crude fat: Determine the crude fat content by referring to the Soxhlet extraction method in GB5009.6-2016.

Crude fiber: Determine the crude fiber content by referring to the filtration method in GB/T6434-2006.

Crude ash: Determine the crude ash content according to the method in GB/T6438-2007.

Table 1

It can be seen from Table 1 that, relative to Comparative Examples 1 to 4, the distiller's grains bio-feed produced by the fermentation methods of Examples 7 to 9 of the present invention has the advantages of lower acetic acid content, excellent mildew resistance, high crude protein content and low crude fiber content.

It should be noted that, in this article, terms such as "include", "comprises" or any other variations thereof are intended to cover non-exclusive inclusion, so that a process, method, article or apparatus that includes a series of elements includes not only those elements, but also other elements not explicitly listed, or also includes elements inherent to such process, method, article or apparatus.

Although embodiments of the present invention have been shown and described, it will be appreciated by those skilled in the art that various changes, modifications, substitutions and variations may be made to the embodiments without departing from the principles and spirit of the present invention, and the scope of the present invention is defined by the appended claims and their equivalents.

Claims (10)

1. A fermentation process for preparing a distillers' grain biological feed, comprising the steps of:

Compounding Saccharomyces cerevisiae powder, candida utilis powder, bacillus subtilis powder, aspergillus niger powder and lactobacillus plantarum powder to obtain a compound microbial agent, stirring and mixing vinasse, bran, bean pulp and the compound microbial agent to obtain a dry material, adding water into the dry material, regulating the humidity of the system to be 50% -60%, fermenting for 2d at 30-37 ℃ to obtain a fermentation system, stirring and adding an antiseptic auxiliary agent into the fermentation system, stirring at a constant temperature of 30-37 ℃ for 1-2h to obtain a fermentation product, drying the fermentation product until the water content is 10% -15%, and obtaining the vinasse biological feed after the drying treatment is finished;

the anti-corrosion auxiliary agent is prepared by the following steps:

adding modified chitosan into hydrochloric acid solution with volume fraction of 1%, stirring and mixing for 10-20min, regulating pH to 5.0-5.2 with sodium hydroxide, stirring and adding montmorillonite powder, stirring at room temperature for 12 hr, centrifuging after stirring, and drying the precipitate to constant weight to obtain the antiseptic auxiliary agent.

2. The fermentation method for preparing distillers' grain biological feed according to claim 1, wherein the saccharomyces cerevisiae powder, candida utilis powder, bacillus subtilis powder, aspergillus niger powder and lactobacillus plantarum powder are compounded according to a mass ratio of 2:2:1:1:1.

3. The fermentation method for producing a distillers' grain biological feed according to claim 1, wherein the number of effective viable bacteria of the Saccharomyces cerevisiae powder, the candida utilis powder, the bacillus subtilis powder, the aspergillus niger powder and the lactobacillus plantarum powder is equal to or more than 1.0 x 10 ⁸/g.

4. The fermentation method for producing a distillers 'grain biological feed according to claim 1, wherein the dry ingredients comprise, by weight, 80 parts of distillers' grains, 20-30 parts of bran, 15-20 parts of soybean meal, and 0.5-1 part of a composite microbial agent.

5. The fermentation method for producing a distillers' grain biological feed according to claim 1, wherein the amount of the preservative aid is 2% -5% of the mass of the fermentation system.

6. The fermentation method for producing a distillers' grain biological feed according to claim 1, wherein the volume fraction of the hydrochloric acid solution, the modified chitosan and the montmorillonite powder is 1l:15-18g:100-150g.

7. The fermentation method for producing a distillers' grain biological feed according to claim 1, wherein the modified chitosan is produced by:

Step A1, adding tocopherol and succinic anhydride into anhydrous pyridine under stirring, stirring for 48 hours to obtain a product, dripping the product into hydrochloric acid solution with pH=1-2, precipitating the hydrochloric acid solution to form a layered system, refrigerating the layered system at 0 ℃ for 24 hours, filtering to obtain a solid component, washing the solid component with deionized water for three times, recrystallizing with ethanol, filtering, and drying the crystal in vacuum to obtain carboxylated tocopherol;

Step A2, soaking chitosan in isopropanol, adding sodium hydroxide into the solution after the chitosan swells, alkalizing and gelatinizing the solution for 1-2 hours at 55-60 ℃, adding chloroacetic acid into the solution, stirring the solution at 65-70 ℃ for reaction for 2-3 hours, cooling the solution to room temperature after the reaction is finished, adding glacial acetic acid into the solution to adjust the pH to be neutral, and filtering, washing the solution with absolute ethyl alcohol and drying the solution to obtain carboxymethyl chitosan;

And A3, adding carboxylated tocopherol into anhydrous dimethyl sulfoxide under stirring, stirring for 10-20min, adding N-hydroxysuccinimide and 1- (3-dimethylaminopropyl) -3-ethylcarbodiimide hydrochloride into the mixture, cooling the system to 0 ℃, stirring the mixture at constant temperature for 2-3h, adding carboxymethyl chitosan into the mixture under stirring, heating the system to 60-65 ℃, stirring the mixture for 4-5h, cooling the mixture to room temperature, filtering the mixture, and drying the mixture to obtain the modified chitosan.

8. The fermentation process for producing a distillers grain biological feed according to claim 7, wherein the anhydrous pyridine, tocopherol and succinic anhydride are used in the ratio of 100ml to 0.01mol to 0.05-0.06mol in step A1.

9. The fermentation process for producing distillers' grain biological feed according to claim 7, wherein the amount ratio of isopropyl alcohol, chitosan, sodium hydroxide and chloroacetic acid in step A2 is 200ml:15-18g:22-25g:40-50g.

10. The fermentation method for producing distillers' grain biological feed according to claim 7, wherein the anhydrous dimethyl sulfoxide, carboxylated tocopherol, N-hydroxysuccinimide, 1- (3-dimethylaminopropyl) -3-ethylcarbodiimide hydrochloride and carboxymethyl chitosan in the amount ratio of 200mL:0.01-0.012mol:0.15-0.20mol:0.18-0.20mol:1.8-2.2g in step A3.