Method for preparing fish feed by secondary utilization of traditional Chinese medicine wastes

Abstract

The invention relates to a method for preparing fish feed by secondary utilization of traditional Chinese medicine wastes, and belongs to the technical field of fish feed. After the traditional Chinese medicine waste is pretreated to remove heavy metal, primary fermentation of bacillus subtilis and aspergillus niger is carried out, the bacillus subtilis can degrade cellulose, the aspergillus niger decomposes antinutritional factors, then fermentation of lactobacillus lactis and candida utilis is carried out, lactic acid bacteria produce acid to inhibit infectious microbes, flavor substances are synthesized, and palatability is improved. Through traditional Chinese medicine residue pretreatment, multi-strain synergistic fermentation and a scientific feed formula, efficient reutilization of traditional Chinese medicine waste is successfully realized, and the fish feed which is rich in nutrition, high in safety and remarkable in economic benefit is produced.

Description

Technical Field

[0001] The present invention belongs to the technical field of fish feed, and relates to a method for preparing fish feed by recycling Chinese medicine waste. Background Art

[0002] With the rapid development of the global aquaculture industry, fish farming has become an important industry to ensure the supply of human protein. The problems brought about by the intensive farming mode are becoming increasingly prominent: on the one hand, traditional fish feeds rely too much on raw materials such as fish meal and soybean meal, leading to rising feed costs and overexploitation of marine resources; on the other hand, antinutritional factors (such as tannins and phytic acid) commonly present in plant protein substitution programs seriously hinder the absorption of nutrients by fish, and the abuse of antibiotics has caused problems such as drug residues, water pollution and decreased fish immunity. Developing new fish feeds with both environmental protection and functionality has become an urgent need to promote the sustainable development of the aquaculture industry.

[0003] As a special type of biomass resource, the large-scale utilization of Chinese medicine waste has important ecological and economic value. The Chinese medicine industry generates millions of tons of solid waste residues every year. The traditional treatment methods mainly include landfill and incineration, which not only cause land occupation and greenhouse gas emissions, but also lead to the waste of functional components such as polysaccharides, flavonoids and alkaloids. Research shows that the active substances remaining in Chinese medicine residues have potential functions such as antibacterial, antioxidant and immunomodulatory effects. If they can be converted into fish feed raw materials, it can not only relieve the pressure of shortage of feed resources, but also improve fish health through functional components. However, directly using Chinese medicine residues as feed additives has significant defects: their heavy metal content (such as lead and cadmium) may exceed the feed safety standards, the lignin-cellulose composite structure hinders the release of active substances, and components such as tannins and saponins have antinutritional properties, and long-term use may inhibit fish growth and cause metabolic disorders. Therefore, how to achieve detoxification, efficiency improvement and resource conversion of Chinese medicine residues through green processes has become the key to breaking through the bottleneck of their feed application.

[0004] In the prior art for preparing fish feed from Chinese medicine residues, most methods only use simple crushing or single-strain solid-state fermentation, and it is difficult to simultaneously achieve heavy metal removal, antinutritional factor degradation and efficient release of active substances. For example, although the traditional acid-base treatment method can partially remove heavy metals, it will damage heat-sensitive components such as polysaccharides; while the single Bacillus subtilis fermentation has insufficient cellulose decomposition efficiency, resulting in most of the functional components in Chinese medicine residues still being embedded in the cell wall. In addition, the degradation rate of antinutritional factors such as tannins and phytic acid remaining in Chinese medicine residues is insufficient in conventional processes, significantly reducing protein digestibility and inducing intestinal inflammation in fish. These technical defects have restricted the large-scale application of Chinese medicine waste in the field of fish feed. Summary of the Invention

[0005] The present invention removes heavy metals through the pretreatment of traditional Chinese medicine waste, releases the active ingredients and nutrients in the traditional Chinese medicine waste through multi-strain step-by-step fermentation, and combines with a scientific feed formula to successfully achieve the efficient reuse of traditional Chinese medicine waste and produce fish feed rich in nutrients.

[0006] The object of the present invention can be achieved by the following technical solutions:

[0007] A method for reusing traditional Chinese medicine waste to prepare fish feed, comprising the following steps:

[0008] (1) Impregnate the traditional Chinese medicine residue with a citric acid-chitosan composite solution and then pulverize it;

[0009] (2) Perform step-by-step fermentation on the medicinal residue treated in step (1) with a composite strain composed of Bacillus subtilis, Aspergillus niger, Lactobacillus lactis, and Candida utilis;

[0010] (3) Add fish meal, soybean meal, wheat bran, rapeseed oil, calcium phosphate, and seaweed powder to the fermentation product obtained in step (2) and mix evenly to make the finished feed.

[0011] As a preferred technical solution of the present invention, the preparation method of the citric acid-chitosan composite solution in step (1) is: drop 1-2% w / v citric acid solution into 1-2% w / v chitosan solution and stir to dissolve; the mass ratio of the citric acid-chitosan composite solution to the traditional Chinese medicine residue is 1:5-10.

[0012] As a preferred technical solution of the present invention, the fermentation process in step (2) includes the following steps:

[0013] (21) Activate and expand Bacillus subtilis, Aspergillus niger, Lactobacillus lactis, and Candida utilis respectively to form seed solutions;

[0014] (22) Add the fermentation medium to the fermenter, sterilize it, and then cool it to a predetermined temperature;

[0015] (23) Inoculate the Bacillus subtilis seed solution and the Aspergillus niger seed solution into the fermenter and perform primary fermentation for 20-28h;

[0016] (24) Add the Lactobacillus lactis seed solution and the Candida utilis seed solution and perform secondary fermentation for 24-48h;

[0017] (25) Perform post-fermentation for 20-28h on the basis of step (24).

[0018] As a preferred technical solution of the present invention, the mass ratio of the freeze-dried powder of the fermentation broth, fish meal, soybean meal, wheat bran, rapeseed oil, calcium phosphate, and seaweed powder in step (3) is 40-60:8-12:6-10:6-10:5-8:2-3:2-3.

[0019] As a preferred technical solution of the present invention, the preparation of the Bacillus subtilis seed liquid in step (21) is as follows: inoculate a single colony of Bacillus subtilis into 5 mL of LB medium, culture it in a shaker at 37 °C and 220 rpm for 12 - 16 hours, and then expand it to the LB medium at an inoculation amount of 1% until the OD 600 reaches 1.0 - 1.2; the preparation of the Aspergillus niger seed liquid is as follows: add 0.1 - 0.5 mL of a spore suspension with a concentration of 10 5 -10 7 spores / mL to 5 mL of Czapek - Dox medium, activate it at 35 °C for 48 - 72 h, and then add it to the Czapek - Dox medium at an inoculation amount of 5% and expand the culture at 35 °C until the OD 600 reaches 0.8 - 1.0; the preparation of the Lactobacillus lactis seed liquid is as follows: inoculate a single colony of Lactobacillus lactis into 5 mL of MRS medium, culture it statically at 37 °C for 12 - 16 h for activation, and then expand it to the MRS medium at an inoculation amount of 1% and culture it statically at 37 °C until the OD 600 reaches 1.0 - 1.2; the preparation of the Candida utilis seed liquid is as follows: inoculate a single colony of Candida utilis into 5 mL of YPD medium, activate and culture it in a shaker at 30 °C and 220 rpm for 12 - 16 h, and then add it to the YPD medium at an inoculation amount of 1% and expand the culture in a shaker at 30 °C and 220 rpm until the OD of the bacterial liquid 600 reaches 1.0 - 1.2.

[0020] As a preferred technical solution of the present invention, the fermentation medium in step (22) comprises the following components: 20 - 30% w / v Chinese medicine residue powder, 1 - 2% w / v yeast extract, 1 - 2% w / v glucose, 0.1 - 0.3% w / v sodium chloride, 0.05 - 0.15% w / v potassium chloride, 0.05 - 0.15% w / v potassium dihydrogen phosphate, and the balance is deionized water.

[0021] As a preferred technical solution of the present invention, the inoculation amount of Bacillus subtilis in step (23) is 2 - 4%, and the inoculation amount of Aspergillus niger is 1 - 3%; the primary fermentation conditions are: temperature 33 - 37 °C, pH 5.5 - 6.0, stirring speed 150 - 200 rpm, aeration rate: the ratio of the air volume flow rate to the liquid volume is 1 - 1.5 vvm.

[0022] As a preferred technical solution of the present invention, the inoculation amount of Lactobacillus lactis in step (24) is 4 - 6%, and the inoculation amount of Candida utilis is 4 - 6%; the secondary fermentation conditions are: temperature 30 - 35 °C, pH 4.5 - 5.0, stirring speed 120 - 150 rpm, aeration rate: the ratio of the air volume flow rate to the liquid volume is 0.5 - 1.0 vvm.

[0023] As a preferred technical solution of the present invention, the later fermentation conditions in step (25) are as follows: temperature 30 - 32 °C, pH 4.0 - 4.5, stirring speed 150 - 180 rpm, aeration rate: the ratio of the air volume flow rate to the liquid volume is 1.5 - 2.0 vvm.

[0024] Advantages of the present invention:

[0025] (1) After impregnating the Chinese medicine residue with the citric acid-chitosan composite solution, the organic acid in the present invention can form stable chelates with heavy metal ions. The natural polymer material chitosan has good adsorption properties, can further adsorb and remove harmful metal ions, and prevent the influence of the residual heavy metal ions in the Chinese medicine residue on the subsequent composite microbial fermentation.

[0026] (2) By using multiple microorganisms such as Bacillus subtilis, Aspergillus niger, Lactobacillus lactis, and Candida utilis for co-fermentation, the present invention makes full use of their respective metabolic characteristics to promote the conversion of organic matter in the Chinese medicine residue into nutrient components that are easily absorbed by fish. Bacillus subtilis and Aspergillus niger can degrade the complex organic matter in the Chinese medicine residue, and then further ferment with Lactobacillus lactis and Candida utilis to generate beneficial metabolites, making full use of the active ingredients and nutrients of the Chinese medicine waste. Specific embodiments

[0027] To further illustrate the technical means and effects adopted by the present invention to achieve the predetermined invention purpose, the following examples are used to describe in detail the specific embodiments, structures, features and their effects according to the present invention.

[0028] The sources of raw materials involved in the following examples and comparative examples are as follows:

[0029] The deacetylation degree of the chitosan is 85 - 95%, purchased from Weimao Biotechnology Co., Ltd.; fish meal is purchased from Shanghai Chuanyong Trading Co., Ltd., with the product number 4562; soybean meal is purchased from Angli Biotechnology Co., Ltd., with the product number 00021; wheat bran is purchased from Xiangnian Natural Health Technology Co., Ltd.; rapeseed oil is purchased from Shandong Chanjixifu Food Co., Ltd.; seaweed powder is purchased from Tianben Biotechnology Co., Ltd.; yeast extract is purchased from Angel Yeast Co., Ltd.; Bacillus subtilis is purchased from Huiqingyuan Biotechnology Co., Ltd.; Aspergillus niger is purchased from Qingdao Haibo Biotechnology Co., Ltd.; Lactobacillus lactis is purchased from Shenghe Bioengineering Co., Ltd.; Candida utilis is purchased from Wuhan Huizao Biotechnology Co., Ltd.; LB medium is purchased from Beyotime Biotechnology Co., Ltd.; Czapek-Dox medium is purchased from Shanghai Maokang Biotechnology Co., Ltd.; YPD medium is purchased from Qingdao Haibo Biotechnology Co., Ltd.

[0030] Example 1

[0031] For juvenile fish, the Chinese medicine waste selected is the residue of wild jujube seeds, coix seeds, Chinese yam, and houttuynia cordata produced by Guangdong Yifang Pharmaceutical Co., Ltd., with a mass ratio of 25:30:20:25, which helps to improve the digestive system of juvenile fish and enhance its immunity to quickly adapt to the growth environment.

[0032] A method for the secondary utilization of Chinese medicine waste to prepare fish feed includes the following steps:

[0033] (1) Immerse the Chinese medicine residue in a citric acid-chitosan composite solution for 10 h and then pulverize it;

[0034] (2) Perform step-by-step fermentation on the residue treated in step (1) with a composite strain composed of Bacillus subtilis, Aspergillus niger, Lactobacillus lactis, and Candida utilis;

[0035] (3) Add fish meal, soybean meal, wheat bran, rapeseed oil, calcium phosphate, and seaweed powder to the fermentation product obtained in step (2) and mix evenly to prepare the finished feed.

[0036] The preparation method of the citric acid-chitosan composite solution in step (1) is: drop a 1% w / v citric acid solution into a 1% w / v chitosan solution and stir to dissolve; the mass ratio of the citric acid-chitosan composite solution to the Chinese medicine residue is 1:5.

[0037] The fermentation process in step (2) includes the following steps:

[0038] (21) Activate and expand Bacillus subtilis, Aspergillus niger, Lactobacillus lactis, and Candida utilis respectively to form seed solutions;

[0039] (22) After adding the fermentation medium to the fermenter and sterilizing it, cool it to 35 °C;

[0040] (23) Inoculate the Bacillus subtilis seed solution and the Aspergillus niger seed solution into the fermenter and conduct primary fermentation for 28 h;

[0041] (24) Add the Lactobacillus lactis seed solution and the Candida utilis seed solution and conduct secondary fermentation for 48 h;

[0042] (25) Conduct post-fermentation for 28 h on the basis of step (24).

[0043] The mass ratio of the freeze-dried powder of the fermentation broth, fish meal, soybean meal, wheat bran, rapeseed oil, calcium phosphate, and seaweed powder in step (3) is 50:12:10:6:5:3:2.

[0044] The preparation of the Bacillus subtilis seed solution in step (21) is: inoculate a single colony of Bacillus subtilis into 5 mL of LB medium, culture it in a shaker at 37 °C and 220 rpm for 14 hours, and then expand it to the LB medium at an inoculation amount of 1% until OD 600is 1.2; The preparation of Aspergillus niger seed liquid is as follows: Add 0.3 mL of spore suspension with a concentration of 10 6 spores / mL to 5 mL of Czapek-Dox medium, activate at 29 °C for 60 h, and then add it to Czapek-Dox medium at an inoculation amount of 5% and expand the culture at 29 °C until the OD 600 is 1.0; The preparation of Lactobacillus lactis seed liquid is as follows: Inoculate a single colony of Lactobacillus lactis into 5 mL of MRS medium, statically culture at 37 °C for 16 h for activation, and then expand it to MRS medium at an inoculation amount of 1% and statically culture at 37 °C until the OD 600 reaches 1.2; The preparation of Candida utilis seed liquid is as follows: Inoculate a single colony of Candida utilis into 5 mL of YPD medium, activate and culture on a shaker at 30 °C and 220 rpm for 16 h, and then add it to YPD medium at an inoculation amount of 1% and expand the culture on a shaker at 30 °C and 220 rpm until the OD of the bacterial liquid 600 reaches 1.2.

[0045] The fermentation medium described in step (22) includes the following components: 20% w / v Chinese medicine residue powder, 2% w / v yeast extract, 2% w / v glucose, 0.1% w / v sodium chloride, 0.05% w / v potassium chloride, 0.05% w / v potassium dihydrogen phosphate, and the balance is deionized water.

[0046] The inoculation amount of Bacillus subtilis described in step (23) is 3%, and the inoculation amount of Aspergillus niger is 2%; The primary fermentation conditions are: temperature 35 °C, pH 6.0, stirring speed 180 rpm, aeration rate: the ratio of air volume flow to liquid volume is 1.5 vvm.

[0047] The inoculation amount of Lactobacillus lactis described in step (24) is 5%, and the inoculation amount of Candida utilis is 5%; The secondary fermentation conditions are: temperature 32 °C, pH 5.0, stirring speed 150 rpm, aeration rate: the ratio of air volume flow to liquid volume is 1.0 vvm.

[0048] The later fermentation conditions described in step (25) are: temperature 32 °C, pH 4.5, stirring speed 180 rpm, aeration rate: the ratio of air volume flow to liquid volume is 2.0 vvm.

[0049] Example 2

[0050] For adult fish, the Chinese medicine waste selected is the Chinese medicine residues of Coix lacryma-jobi, Plantago asiatica, Chrysanthemum morifolium, and Perilla frutescens from Guangdong Yifang Pharmaceutical Co., Ltd., with a mass ratio of 25:20:25:30, which helps adult fish better absorb the nutrients in the feed.

[0051] The object of the present invention can be achieved by the following technical solutions:

[0052] A method for the secondary utilization of traditional Chinese medicine waste to prepare fish feed, comprising the following steps:

[0053] (1) Immerse the traditional Chinese medicine residue in a citric acid-chitosan composite solution for 8 h and then pulverize it;

[0054] (2) Perform step-by-step fermentation on the residue treated in step (1) with a composite strain composed of Bacillus subtilis, Aspergillus niger, Lactobacillus lactis, and Candida utilis;

[0055] (3) Add fish meal, soybean meal, wheat bran, rapeseed oil, calcium phosphate, and seaweed powder to the fermentation product obtained in step (2) and mix evenly to prepare the finished feed.

[0056] The preparation method of the citric acid-chitosan composite solution described in step (1) is: drop 1% w / v citric acid solution into 1% w / v chitosan solution and stir to dissolve; the mass ratio of the citric acid-chitosan composite solution to the traditional Chinese medicine residue is 1:5.

[0057] The fermentation process in step (2) includes the following steps:

[0058] (21) Activate and expand Bacillus subtilis, Aspergillus niger, Lactobacillus lactis, and Candida utilis respectively to form seed solutions;

[0059] (22) Add the fermentation medium to the fermentation tank, sterilize it, and then cool it to a predetermined temperature;

[0060] (23) Inoculate the Bacillus subtilis seed solution and the Aspergillus niger seed solution into the fermentation tank and perform primary fermentation for 20 h;

[0061] (24) Add the Lactobacillus lactis seed solution and the Candida utilis seed solution and perform secondary fermentation for 24 h;

[0062] (25) Perform post-fermentation for 20 h on the basis of step (24).

[0063] The mass ratio of the freeze-dried powder of the fermentation broth, fish meal, soybean meal, wheat bran, rapeseed oil, calcium phosphate, and seaweed powder described in step (3) is 60:8:10:10:8:2:2.

[0064] The preparation of the Bacillus subtilis seed solution described in step (21) is: inoculate a single colony of Bacillus subtilis into 5 mL of LB medium, culture it in a shaker at 37 °C and 220 rpm for 12 hours, and then expand it to the LB medium at an inoculation amount of 1% until the OD 600 is 1.0; the preparation of the Aspergillus niger seed solution is: add 0.1 mL of a spore suspension with a concentration of 10 6 spores / mL to 5 mL of Czapek-Dox medium, activate it at 28 °C for 48 h, and then add it to the Czapek-Dox medium at an inoculation amount of 5% and expand the culture at 28 °C until the OD600 is 0.8; The preparation of the Lactobacillus casei seed solution is as follows: inoculate a single colony of Lactobacillus casei into 5 mL of MRS medium, statically culture at 37 °C for 12 h for activation, and then expand it into MRS medium at an inoculation amount of 1% and statically culture at 37 °C until the OD 600 reaches 1.0; The preparation of the Candida utilis seed solution is as follows: inoculate a single colony of Candida utilis into 5 mL of YPD medium, activate and culture it on a shaker at 30 °C and 220 rpm for 12 h, and then add it to YPD medium at an inoculation amount of 1% and expand the culture on a shaker at 30 °C and 220 rpm until the OD of the bacterial liquid 600 reaches 1.0.

[0065] The fermentation medium described in step (22) includes the following components: 30% w / v Chinese medicine residue powder, 2% w / v yeast extract, 2% w / v glucose, 0.3% w / v sodium chloride, 0.15% w / v potassium chloride, 0.15% w / v potassium dihydrogen phosphate, and the balance is deionized water.

[0066] The inoculation amount of Bacillus subtilis described in step (23) is 4%, and the inoculation amount of Aspergillus niger is 3%; The primary fermentation conditions are: temperature 33 °C, pH 5.8, stirring speed 180 rpm, aeration rate: the ratio of air volume flow to liquid volume is 1.2 vvm.

[0067] The inoculation amount of Lactobacillus casei described in step (24) is 6%, and the inoculation amount of Candida utilis is 6%; The secondary fermentation conditions are: temperature 30 °C, pH 4.8, stirring speed 140 rpm, aeration rate: the ratio of air volume flow to liquid volume is 0.7 vvm.

[0068] As a preferred technical solution of the present invention, the later fermentation conditions described in step (25) are: temperature 31 °C, pH 4.2, stirring speed 160 rpm, aeration rate: the ratio of air volume flow to liquid volume is 1.8 vvm.

[0069] Example 3

[0070] For slightly eutrophic aquaculture, the Chinese medicine waste is selected from the Chinese medicine residues of Ziziphus jujuba seed, Houttuynia cordata, Coix lacryma-jobi, Cornus officinalis, and Plantago asiatica of Guangdong Yifang Pharmaceutical Co., Ltd., and the mass ratio is 15:25:20:15:25, which helps to inhibit harmful microorganisms in water, enhance immunity, regulate the growth state of fish groups, and improve the stress resistance of fish.

[0071] A method for preparing fish feed by reusing Chinese medicine waste includes the following steps:

[0072] (1) Immerse the Chinese medicine residue in a citric acid-chitosan composite solution for 6 h and then pulverize it;

[0073] (2) A complex strain composed of Bacillus subtilis, Aspergillus niger, Lactobacillus lactis, and Candida utilis is used to perform step-by-step fermentation on the medicinal residues treated in step (1).

[0074] (3) Fish meal, soybean meal, wheat bran, rapeseed oil, calcium phosphate, and seaweed powder are added to the fermentation product obtained in step (2) and mixed evenly to prepare a finished feed.

[0075] The preparation method of the citric acid-chitosan composite solution described in step (1) is as follows: A 2% w / v citric acid solution is dropped into a 2% w / v chitosan solution and stirred to dissolve; the mass ratio of the citric acid-chitosan composite solution to the traditional Chinese medicine residues is 1:7.

[0076] The fermentation process in step (2) includes the following steps:

[0077] (21) Bacillus subtilis, Aspergillus niger, Lactobacillus lactis, and Candida utilis are respectively activated and expanded to form seed solutions.

[0078] (22) After adding the fermentation medium to the fermenter and sterilizing it, the temperature is reduced to 35°C.

[0079] (23) The Bacillus subtilis seed solution and the Aspergillus niger seed solution are inoculated into the fermenter and initially fermented for 24 h.

[0080] (24) The Lactobacillus lactis seed solution and the Candida utilis seed solution are added for secondary fermentation for 36 h.

[0081] (25) Post-fermentation is carried out for 24 h on the basis of step (24).

[0082] The mass ratio of the freeze-dried fermentation broth, fish meal, soybean meal, wheat bran, rapeseed oil, calcium phosphate, and seaweed powder described in step (3) is 50:10:8:8:6:2:3.

[0083] The preparation of the Bacillus subtilis seed solution described in step (21) is as follows: A single colony of Bacillus subtilis is inoculated into 5 mL of LB medium and cultured in a shaker at 37°C and 220 rpm for 14 hours, and then expanded to LB medium at an inoculation amount of 1% until OD 600 reaches 1.1; the preparation of the Aspergillus niger seed solution is as follows: 0.3 mL of a spore suspension with a concentration of 10 6 spores / mL is added to 5 mL of Czapek-Dox medium, activated at 29°C for 60 h, and then added to Czapek-Dox medium at an inoculation amount of 5% and expanded and cultured at 29°C until OD 600 reaches 0.9; the preparation of the Lactobacillus lactis seed solution is as follows: A single colony of Lactobacillus lactis is inoculated into 5 mL of MRS medium, statically cultured at 37°C for 14 h for activation, and then expanded to MRS medium at an inoculation amount of 1% and statically cultured at 37°C until OD 600Reach 1.1; The preparation of Candida utilis seed liquid is as follows: Inoculate a single colony of Candida utilis into 5 mL of YPD medium, activate and culture it in a shaker at 30 °C and 220 rpm for 14 h, then add it to the YPD medium at an inoculation amount of 1%, and expand the culture in a shaker at 30 °C and 220 rpm until the OD of the bacterial liquid 600 Reach 1.1.

[0084] The fermentation medium described in step (22) includes the following components: 30% w / v Chinese medicine residue powder, 1% w / v yeast extract, 1% w / v glucose, 0.1% w / v sodium chloride, 0.05% w / v potassium chloride, 0.05% w / v potassium dihydrogen phosphate, and the balance is deionized water.

[0085] The inoculation amount of Bacillus subtilis described in step (23) is 2-4%, and the inoculation amount of Aspergillus niger is 1-3%; The primary fermentation conditions are: temperature 37 °C, pH 5.8, stirring speed 180 rpm, aeration rate: the ratio of air volume flow to liquid volume is 1.2 vvm.

[0086] The inoculation amount of Lactobacillus lactis described in step (24) is 4-6%, and the inoculation amount of Candida utilis is 4-6%; The secondary fermentation conditions are: temperature 35 °C, pH 4.8, stirring speed 140 rpm, aeration rate: the ratio of air volume flow to liquid volume is 0.8 vvm.

[0087] The later fermentation conditions described in step (25) are: temperature 31 °C, pH 4.2, stirring speed 160 rpm, aeration rate: the ratio of air volume flow to liquid volume is 1.8 vvm.

[0088] Comparative Example 1

[0089] On the basis of Example 1, step (1) was changed to use water immersion, and the rest was the same as in Example 1.

[0090] Comparative Example 2

[0091] On the basis of Example 1, in step (1), alkali treatment was used to remove heavy metals. Specifically, the Chinese medicine residue was impregnated in 0.1 M sodium hydroxide solution, and the rest was the same as in Example 1.

[0092] Comparative Example 3

[0093] On the basis of Example 1, during the fermentation process, Bacillus subtilis was not inoculated in the primary fermentation, and the inoculation amount of Aspergillus niger was changed to 5%, and the rest was the same as in Example 1.

[0094] Comparative Example 4

[0095] On the basis of Example 1, during the fermentation process, Aspergillus niger was not inoculated in the primary fermentation, and the inoculation amount of Bacillus subtilis was changed to 5%, and the rest was the same as in Example 1.

[0096] Comparative Example 5

[0097] On the basis of Example 1, during the fermentation process, Lactobacillus casei was not inoculated in the secondary fermentation, and the inoculation amount of Candida utilis was changed to 10%, and the rest was the same as in Example 1.

[0098] Comparative Example 6

[0099] On the basis of Example 1, during the fermentation process, Candida utilis was not inoculated in the secondary fermentation, and the inoculation amount of Lactobacillus casei was changed to 10%, and the rest was the same as in Example 1.

[0100] Performance test:

[0101] The fish feeds prepared in Examples 1-3 and Comparative Examples 1-6 were used as test experiments to feed perch;

[0102] Before the test, the juvenile Micropterus salmonides were pre-fed for 15 days to adapt them to the environment, and the basic feed was fed at 8:00 and 18:00 every day, and the feeding rate was 2%;

[0103] After the pre-feeding was completed, the juvenile Micropterus salmonides were randomly divided into 9 groups, with 100 tails in each group, and the mass of the Micropterus salmonides was weighed as the initial weight. Except for the different feeds fed, the rest was the same as the pre-feeding conditions, and the feeding was carried out for 56 days;

[0104] After the test was completed, the mass of the Micropterus salmonides was measured and recorded as the final weight, and the growth conditions and growth indexes of the perch were detected. The results are shown in Table 1

[0105] Survival rate (%) = number of surviving fish / number of initial fish × 100%

[0106] Weight gain rate (%) = (final weight - initial weight) × 100% / initial weight;

[0107] Condition factor (g / cm 3 ) = 100 × fish body weight / (fish body length) 3 ;

[0108] Randomly select 5 tails from Example 1 and Comparative Example 2 for detecting the heavy metal element content of the whole fish, and take the average value.

[0109] Table 1 Growth conditions of perch

[0110] Survival rate (%) Weight gain rate (%) <![CDATA[Condition factor (g / cm 3 )]]> Example 1 99 572 2.80 Example 2 98 563 2.68 Example 3 98 548 2.74 Comparative Example 1 80 425 2.29 Comparative Example 2 82 442 2.43 Comparative Example 3 95 465 2.39 Comparative Example 4 94 460 2.46 Comparative Example 5 96 483 2.48 Comparative Example 6 95 489 2.51

[0111] Table 2 Heavy metal content of whole fish (fresh weight, unit: mg / kg)

[0112] Fe Zn As Example 1 4.26 6.35 0.0093 Comparative Example 2 8.98 9.64 0.0134

[0113] As can be seen from the results of Examples 1-3 and Comparative Examples 1-2 in Table 1, impregnating the Chinese medicine residues with deionized water may not be able to remove heavy metals in the Chinese medicine residues, affecting the survival rate of juvenile fish, and the fermentation of the bacterial strains leads to a decrease in both the weight gain rate and the condition factor; pretreating the Chinese medicine residues with alkali also results in a decline in the growth of perch, probably because the alkali treatment dissolves and consumes some active ingredients and nutrients; from the heavy metal content data of Example 1 and Comparative Example 2 in Table 3, it can be seen that the citric acid-chitosan of the present invention has a more significant effect on removing heavy metals through chelation and adsorption than the alkali treatment of the prior art.

[0114] As can be seen from the perch growth results of Examples 1-3 and Comparative Examples 3-6 in Table 1, the present invention can degrade complex organic substances in Chinese medicine residues through Bacillus subtilis and Aspergillus niger, and then further ferment with Lactobacillus lactis and Candida utilis to generate beneficial metabolites, making more full use of the active ingredients and nutrients of Chinese medicine waste and promoting the growth of perch.

[0115] The above are only the preferred embodiments of the present invention, and do not impose any form of limitation on the present invention. Although the present invention has been disclosed above with preferred embodiments, it is not intended to limit the present invention. Any person skilled in the art can make some modifications or equivalent changes and modifications within the scope of the technical solution of the present invention by using the disclosed technical content above. However, as long as it does not depart from the technical solution content of the present invention, any indirect modification, equivalent change and modification made to the above embodiments based on the technical essence of the present invention still fall within the scope of the technical solution of the present invention.

Claims

1. A method for preparing fish feed by reusing traditional Chinese medicine waste, characterized in that: It includes the following steps: (1) Impregnate the Chinese medicine residues with a citric acid-chitosan composite solution and then crush them; (2) Use a composite strain composed of Bacillus subtilis, Aspergillus niger, Lactobacillus lactis, and Candida utilis to perform step-by-step fermentation on the medicine residues treated in step (1); (3) Add fish meal, soybean meal, wheat bran, rapeseed oil, calcium phosphate, and seaweed powder to the fermentation product obtained in step (2) and mix evenly to make the finished feed.

2. The method for preparing fish feed by secondary utilization of traditional Chinese medicine waste according to claim 1, characterized in that: The preparation method of the citric acid-chitosan composite solution described in step (1) is: drop a 1-2% w / v citric acid solution into a 1-2% w / v chitosan solution and stir to dissolve; the mass ratio of the citric acid-chitosan composite solution to the Chinese medicine residues is 1:5-10.

3. A method for preparing fish feed by secondary utilization of traditional Chinese medicine waste according to claim 1, characterized in that: The fermentation process in step (2) includes the following steps: (21) Activate and expand Bacillus subtilis, Aspergillus niger, Lactobacillus lactis, and Candida utilis respectively to form seed solutions; (22) Add the fermentation medium to the fermenter, sterilize it, and then cool it to a predetermined temperature; (23) Inoculate the Bacillus subtilis seed solution and the Aspergillus niger seed solution into the fermenter and conduct primary fermentation for 20-28 h; (24) Add the Lactobacillus lactis seed solution and the Candida utilis seed solution and conduct secondary fermentation for 24-48 h; (25) Conduct post-fermentation for 20-28 h on the basis of step (24).

4. A method for preparing fish feed by secondary utilization of traditional Chinese medicine waste according to claim 1, characterized in that: The mass ratio of the freeze-dried powder of the fermentation broth, fish meal, soybean meal, wheat bran, rapeseed oil, calcium phosphate, and seaweed powder described in step (3) is 40-60:8-12:6-10:6-10:5-8:2-3:2-3.

5. A method for preparing fish feed by reusing traditional Chinese medicine waste according to claim 3, characterized in that: The OD of the Bacillus subtilis seed solution described in step (21) 600 is 1.0 - 1.2; the OD of the Aspergillus niger seed solution 600 is 0.8 - 1.0; the OD of the Lactobacillus lactis seed solution 600 is 1.0 - 1.2; OD of Candida utilis seed solution 600 is 1.0 - 1.

2.

6. The method for preparing fish feed by secondary utilization of traditional Chinese medicine waste according to claim 2, characterized in that: The fermentation medium described in step (22) includes the following components: Chinese medicine residue powder, yeast extract, glucose, sodium chloride, potassium chloride, potassium dihydrogen phosphate, and deionized water.

7. A method for preparing fish feed by secondary utilization of traditional Chinese medicine waste according to claim 2, characterized in that: The inoculation amount of Bacillus subtilis described in step (23) is 2-4%, and the inoculation amount of Aspergillus niger is 1-3%; the primary fermentation conditions are: temperature 33-37 °C, pH 5.5-6.0, stirring speed 150-200 rpm, and ventilation volume 1-1.5 vvm.

8. A method for preparing fish feed by reusing traditional Chinese medicine waste according to claim 2, characterized in that: The inoculation amount of Lactobacillus lactis described in step (24) is 4-6%, and the inoculation amount of Candida utilis is 4-6%; the secondary fermentation conditions are: temperature 30-35 °C, pH 4.5-5.0, stirring speed 120-150 rpm, and ventilation volume 0.5-1.0 vvm.

9. A method for preparing fish feed by secondary utilization of traditional Chinese medicine waste according to claim 2, characterized in that: The post-fermentation conditions described in step (25) are: temperature 30-32 °C, pH 4.0-4.5, stirring speed 150-180 rpm, and ventilation volume 1.5-2.0 vvm.