Production process of hermetia illucens-soybean meal fermented feed and application of hermetia illucens-soybean meal fermented feed in preparation of rana nigromaculata feed

Abstract

The invention discloses a production process of hermetia illucens-soybean meal fermented feed and application of the hermetia illucens-soybean meal fermented feed in preparation of rana nigromaculata feed, and belongs to the field of insect resource utilization. The preparation method comprises the following steps: preparing a bacterial liquid from a composite fermentation bacterial agent, compounding adult black soldier fly larvae into pulp, compounding the pulp with soybean meal to obtain a mixture, adding the bacterial liquid into the mixture, and carrying out microbial fermentation to prepare the black soldier fly-soybean meal fermented feed. Wherein the compound fermentation bacterial agent comprises bacillus subtilis, bacillus licheniformis, lactobacillus plantarum, saccharomyces cerevisiae and enterococcus faecalis. According to the black soldier fly-soybean meal fermented feed produced by the invention, the processing and storage cost of black soldier fly larvae is reduced to the greatest extent, and when the black soldier fly-soybean meal fermented feed is applied to a compound feed of rana nigromaculata, the use amount of fish meal in an aquatic feed can be reduced, and the black soldier fly-soybean meal fermented feed can effectively promote growth, improve intestinal health, improve internal immunity and disease resistance and effectively prevent and control diseases; and fishery medicine application is reduced.

Description

Technical Field

[0001] The invention belongs to the field of insect resource utilization, and particularly relates to a black soldier fly-soybean meal fermented feed production process and application thereof in black spot frog feed preparation. Background Art

[0002] Using black soldier fly larvae to biologically treat organic waste such as kitchen waste and livestock and poultry manure can convert the nutrients in them into high-value-added insect protein, which is a green, low-carbon production technology for high-quality protein raw materials. Regarding the technology of using black soldier fly larvae as feed, there are mainly fresh worms (or frozen worms), dried worms and insect powder, as well as processed products such as insect paste or insect paste. Among them, frozen worms retain the active substances of black soldier flies, but have high requirements for storage and transportation conditions; dried worms are obtained by high-temperature or low-temperature heating and drying or microwave drying. Although they are easy to store and transport, the processing energy consumption is high and the active substances of the insect body are reduced; insect paste or insect paste products are obtained by treating fresh worms through specific processes such as enzymolysis, concentration, fermentation or enzyme-bacteria combination, which retains the active substances of the larvae. The use of solid-state fermentation technology to treat black soldier fly larvae is not only conducive to the storage and transportation of raw materials, but also conducive to the function of active substances such as antimicrobial peptides in the black soldier fly raw materials after fermentation treatment.

[0003] The combined fermentation of Bacillus, yeast, lactic acid bacteria and other strains can rapidly degrade the protein of black soldier fly larvae, and produce organic acids, bacteriocins, hydrogen peroxide and other substances during the fermentation process, which have a wide range of antibacterial effects. The production of organic acids can also effectively extend the shelf life of fermented feed and improve its taste. Based on these characteristics, the addition of composite microbial strains to ferment black soldier fly larvae feed can produce flavor substances such as lactic acid, higher alcohols and esters during the fermentation process, improve the flavor of the feed, and make it have good appetizing properties. Active substances such as antimicrobial peptides in the fermented feed still have high antibacterial activity. It can greatly improve its problems such as short shelf life and poor flavor. The application of black soldier fly larvae-soybean meal fermented feed in aquaculture feed is conducive to the production and application of aquatic feed, improves the growth rate and cultivation specifications of aquatic animals, and reduces the occurrence of diseases. Summary of the invention

[0004] The purpose of the present invention is to provide a black soldier fly-soybean meal fermented feed production process which can maximize the reduction of the processing and storage costs of black soldier fly larvae, retain as many active substances in the larvae as possible, and facilitate transportation and storage, so as to form a standardized raw material product for application in aquaculture, specifically a black soldier fly-soybean meal fermented feed production process and its application in the preparation of black-spotted frog feed.

[0005] The object of the present invention is achieved in this way: a black soldier fly-soybean meal fermentation feed production process comprises the following steps:

[0006] (1) preparing a fermentation solution by mixing a composite fermentation agent containing Bacillus subtilis, Bacillus licheniformis, Lactobacillus plantarum, Saccharomyces cerevisiae and Enterococcus faecalis with water in a ratio of 1:19;

[0007] (2) The black soldier fly larvae are ground into insect pulp using a colloid refiner, and the soybean meal is ultrafinely ground and passed through an 80-mesh sieve.

[0008] (3) preparing insect pulp and soybean meal in a weight ratio of 2:(1-2), mixing them evenly to obtain a mixture with a water content of 40% to 50%;

[0009] (4) adding the fermentation bacteria liquid prepared in step (1) in an amount of 1% to 3% by weight of the mixture obtained in step (3), and mixing evenly to obtain a mixed material.

[0010] (5) The mixed material is put into a fermentation bag with an exhaust valve, packaged and sealed, and placed in a constant temperature room at 30° C. to 35° C. for fermentation for 5 to 7 days to obtain black soldier fly-soybean meal fermented feed.

[0011] The live bacteria content of the composite fermentation agent is: 1.0-3.0×10 11 CFU / g, Bacillus licheniformis 1.0~3.0×10 11 CFU / g, Lactobacillus plantarum 1.0~5.0×10 10 CFU / g, Saccharomyces cerevisiae 2.0~6.0×10 10 CFU / g and Enterococcus faecalis 1.0~5.0×10 10 CFU / g.

[0012] The moisture content of the black soldier fly larvae is 60% to 70%.

[0013] The soybean meal is soybean meal with skin, and the moisture content is 8% to 12%.

[0014] The weight ratio of the insect pulp to the soybean meal is 2:(1-2), and the preferred weight ratio is 1:1.

[0015] The black soldier fly-soybean meal fermented feed of the present invention is used in the black-spotted frog compound feed. The preparation method of the black-spotted frog compound feed comprises the following steps: preparing a base feed and the black soldier fly-soybean meal fermented feed in a weight ratio of (11-12): (8-9), mixing them evenly and then granulating them to obtain the black-spotted frog compound feed.

[0016] The basic feed comprises the following components in parts by weight: 15 parts of fish meal, 19 parts of flour, 6 parts of rapeseed meal, 10 parts of peanut meal, 5 parts of cottonseed meal, 1 part of soybean lecithin, 0.3 parts of 50% choline chloride, 1.5 parts of calcium dihydrogen phosphate, 0.1 parts of vitamin premix, and 0.25 parts of mineral premix.

[0017] The vitamin premix in the basic feed, calculated based on the total mass fraction of 100%, consists of the following components: 30.7% rice husk powder, 40.0% VC, 1.5% VA, 1.3% VB1, 2.0% VB2, 2.4% VB6, 12 0.8%, VD30.5%, VK3 3.5%, niacin 3.6%, folic acid 0.4%, D-biotin 0.8%, inositol 12.5%.

[0018] The mineral premix in the basic feed, calculated by total mass fraction of 100%, consists of the following components: 42.4% zeolite, 26.7% rice husk powder, 1.0% CuSO4·5H2O, 15.0% MgSO4·7H2O, 6.7% FeSO4·7H2O, 3.5% ZnSO4·7H2O, 2.4% KCl, 0.9% MnSO4·7H2O, 0.7% CaH4I2O6, 0.5% Na2SeO3, and 0.2% CoSO4·7H2O.

[0019] The invention utilizes microbial fermentation technology to make fermented feed from black soldier fly larvae with high water content through a solid-state fermentation production process. The preparation method is simple and quick, the processing cost of the black soldier fly larvae into feed is reduced, and the storage and transportation are convenient.

[0020] The present invention adopts soybean meal with husk as a compound auxiliary material. The soybean meal with husk has good water absorption and water retention properties. After being compounded with insect pulp, the porosity of the fermented material can be increased, the air permeability can be improved, and it is beneficial to the growth of aerobic microorganisms in the initial stage of material fermentation. Compared with the dehulled soybean meal, the soybean meal with husk has lower cost, and the soybean husk is processed by a microbial fermentation process to degrade cellulose and improve the nutritional value.

[0021] The black soldier fly-soybean meal fermented feed produced by the invention is applied in the black-spotted frog compound feed, which can reduce the amount of fish meal in the compound feed. As an insect, the black soldier fly is the food source of the black-spotted frog under natural conditions. The black soldier fly insect protein is used to replace part of the fish meal in the compound feed, which has good economy and strong feasibility.

[0022] The black soldier fly-soybean meal fermented feed produced by the invention has a certain feeding effect on insectivorous animals such as frogs, and improves the appetite and immunity of black-spotted frogs.

[0023] Instruction Manual

[0024] Figure 1 The effect of different feeds on the intestinal tissue morphology of black-spotted frog. DETAILED DESCRIPTION

[0025] The present invention will be further described below in conjunction with specific embodiments.

[0026] Embodiment 1:

[0027] Preparation of black soldier fly-soybean meal fermented feed:

[0028] (1) The composite fermentation agent and water were mixed in a ratio of 1:19 to prepare a fermentation solution; the live bacteria content of the composite fermentation agent was: Bacillus subtilis 1.0×10 11 CFU / g, Bacillus licheniformis 1.0×10 11 CFU / g, Lactobacillus plantarum 1.0×10 10 CFU / g, Saccharomyces cerevisiae 2.0×10 10 CFU / g and Enterococcus faecalis 1.0×10 10 CFU / g.

[0029] (2) The black soldier fly larvae are ground into insect pulp using a colloid refiner, and the soybean meal with husk is ultra-finely ground and passed through an 80-mesh sieve.

[0030] (3) preparing insect pulp and soybean meal in a weight ratio of 1:1, mixing them evenly to obtain a mixture with a water content of 40%;

[0031] (4) adding the fermentation bacteria liquid prepared in step (1) in an amount of 3% by weight of the mixture, and mixing evenly to obtain a mixed material.

[0032] (5) The mixed material is put into a fermentation bag with an exhaust valve, packaged and sealed, and placed in a constant temperature room at 35° C. for fermentation for 5 days to obtain black soldier fly-soybean meal fermented feed.

[0033] The compound fermentation agent was purchased from Henan Xinyang Bioenzyme Preparation Co., Ltd.

[0034] The nutritional components of black soldier fly larvae, soybean meal with skin, and black soldier fly-soybean meal materials before and after fermentation are shown in Table 1. After the black soldier fly-soybean meal materials were fermented, the content of nutrients such as crude protein and crude fat increased, and the crude fiber content decreased.

[0035] Table 1 Nutritional components of black soldier fly larvae, soybean meal with skin, and black soldier fly-soybean meal materials before and after fermentation %

[0036]

[0037]

[0038] Embodiment 2:

[0039] In order to study the application effect of black soldier fly-soybean meal fermented feed in black-spotted frog compound feed, the present invention designs 3 treatments in the experiment: a compound feed without adding black soldier fly raw materials as comparative example 1, a compound feed with black soldier fly fresh worm paste replacing 40% of fish meal as comparative example 2, and a compound feed with black soldier fly-soybean meal fermented feed and equal protein replacing 40% of fish meal as embodiment 2, and 3 kinds of equal nitrogen and equal fat compound feeds are prepared, and the specific feed formula is shown in Table 2. The feed is granulated after preparation and air-dried for standby use.

[0040] Table 2 Comparative Examples and Examples Feed Raw Material Composition and Nutritional Components (Air-dried Sample Basis) %

[0041]

[0042]

[0043] Note: 1) The vitamin premix, calculated by total mass fraction as 100%, comprises the following components: 30.7% rice bran powder, 40.0% VC, 1.5% VA, 1.3% VB1, 2.0% VB2, 2.4% VB6, 12 0.8%, VD3 0.5%, VK3 3.5%, niacin 3.6%, folic acid 0.4%, D-biotin 0.8%, inositol 12.5%. 2) The mineral premix, calculated based on a total mass fraction of 100%, consists of the following components: 42.4% of zeolite, 26.7% of rice husk powder, 1.0% of CuSO4·5H2O, 15.0% of MgSO4·7H2O, 6.7% of FeSO4·7H2O, 3.5% of ZnSO4·7H2O, 2.4% of KCl, 0.9% of MnSO4·7H2O, 60.7% of CaH4I2O, 30.5% of Na2SeO, and 0.2% of CoSO4·7H2O.

[0044] When grouping the test, healthy, non-deformed, energetic, and similar-sized young frogs were selected for the test, with an average body weight of (16.24±0.04) g / frog, and randomly assigned to 9 water tanks (water tank diameter 1.2m, height 0.8m, water depth 0.1m, set with floating feeding platform), 18 frogs in each water tank, the 9 water tanks were randomly divided into 3 groups, 3 replicates in each group, and the corresponding test feed was fed respectively. Feeding was done twice a day (10:00, 16:00), and the feeding amount of the next day was adjusted according to the feeding situation of each group. If there was any leftover feed, it was collected and deducted. During the experiment, natural light was used, the water temperature was 24-33℃, and the pH was 7.46-8.42. The experimental breeding cycle was 42 days.

[0045] Animal experiments were conducted in accordance with the Regulations on the Administration of Experimental Animals and the regulations of the Animal Welfare and Ethics Committee of Jiangsu Academy of Agricultural Sciences. At the end of the breeding of black-spotted frogs, they were fasted for 24 hours, and each group of black-spotted frogs was caught and weighed one by one. The black-spotted frogs were killed and dissected using the double myelination method, and samples such as liver and intestinal contents were taken, placed in liquid nitrogen, and transferred to -80°C for storage.

[0046] Production performance index calculation formula:

[0047] Survival rate (SR,%) = 100 × N f / N i ;

[0048] Weight gain (WG,%) = 100 × (W f -W i ) / W i ;

[0049] Specific growth ratio (SGR, % / d) = 100 × ln (W f / W i ) / D;

[0050] Feed conversion ratio (FCR) = F / (W f -W i );

[0051] Protein efficiency ratio (PER) = (W f -W i ) / P;

[0052] Hepatosomatic index (HSI, %) = 100 × W h / W;

[0053] Visceral index (VSI, %) = 100 × W v / W;

[0054] Hind leg index (HLI, %) = 100 × W l / W.

[0055] Where N f is the number of frogs at the end of the experiment (g), N i is the number of frogs at the beginning of the experiment (g), W f is the frog weight at the end of the experiment (g), W iis the initial frog weight (g), D is the number of days of feeding, F is the dry weight of feed intake (g), P is the dry weight of protein intake (g), W h is the weight of frog liver (g), W v is the weight of frog viscera (g), W l is the weight of the frog’s hind legs (g), and W is the corresponding frog body weight (g).

[0056] The antioxidant indexes such as superoxide dismutase (SOD), malondialdehyde (MDA), glutathione peroxidase (GSH-Px), catalase (CAT) and total antioxidant capacity (T-AOC) of liver samples were determined by hydroxylamine method, thiobarbituric acid method, colorimetry, visible light method and iron ion reduction / antioxidant capacity method.

[0057] As shown in Table 3, the average weight, weight gain rate, specific growth rate, feed protein efficiency, and hind leg rate of black-spotted frog in Example 2 are significantly higher than those in Comparative Example 1 (P < 0.05), and there is no significant difference with Comparative Example 2 (P > 0.05), and the feed coefficient of Example 2 is significantly lower than that of Comparative Example 1 (P < 0.05). The results show that replacing 40% of fish meal with black soldier fly-soybean meal fermented feed in black-spotted frog compound feed can show better production performance and achieve the effect of using black soldier fly fresh insect paste, which further verifies that the black soldier fly-soybean meal fermented feed produced after the fermentation process improves the nutritional value of the material.

[0058] Table 3 Effects of different feeds on the growth performance of black-spotted frog

[0059]

[0060] Note: Data without letters in the same column indicate no significant difference (P>0.05), and different lowercase letters indicate significant difference (P<0.05).

[0061] As shown in Table 4, the superoxide dismutase, glutathione peroxidase, catalase and total antioxidant capacity of black-spotted frog in Example 2 are higher than those in Comparative Examples 1 and 2, and the catalase activity is significantly higher than that in Comparative Example 1 (P < 0.05), indicating that the black soldier fly larvae-soybean meal fermented feed can improve the antioxidant enzyme activity of black-spotted frog, especially the catalase activity. Malondialdehyde is a lipid peroxidation product in the hepatopancreas and is generally considered to be an indicator of oxidative damage. There is no significant difference in the malondialdehyde content between different groups (P > 0.05), indicating that the black soldier fly larvae-soybean meal fermented feed will not have a negative impact on the liver.

[0062] Table 4 Effects of different feeds on antioxidant enzyme activities in the liver of Rana nigromaculata

[0063]

[0064] The midgut tissues of each group were selected to prepare paraffin sections and stained with HE. The number of villi, villi height and cavity rate of the black-spotted frog intestinal tract were observed microscopically (see Table 5). The intestinal cavity rate of Example 2 was significantly higher than that of Comparative Example 1 and Comparative Example 2. The goblet cells in the intestinal epithelial cells secrete mucus proteins to form a mucus layer attached to the epithelial layer, thereby isolating and controlling the colonization and invasion of intestinal microorganisms, and play a key role in intestinal immunity. Compared with Comparative Example 1, more goblet cells were observed in Example 2 (see Table 5). Figure 1 ), indicating that the black soldier fly-soybean meal fermented feed has a certain stimulating effect on the intestine, promotes the secretion of mucus protein, and improves the intestinal immune function. In general, the intestinal wall of each group was thicker and the intestinal villi were not damaged, and different feeds had no negative impact on the intestinal tissue structure.

[0065] Table 5 Statistics of the number, height and cavity rate of intestinal villi of Rana nigromaculata in different diet groups

[0066]

[0067] Although the present invention has been described in detail above using general descriptions, specific implementation methods and experiments, it is obvious to those skilled in the art that some modifications or improvements can be made on the basis of the present invention. Therefore, these modifications or improvements made without departing from the spirit of the present invention are within the scope of protection claimed by the present invention.

Claims

1. A black soldier fly-soybean meal fermented feed production process, characterized in that: The production process is: First, the composite fermentation agent is prepared into a bacterial liquid, then the black soldier fly larvae are ground into insect pulp and compounded with soybean meal to obtain a mixture, and then the bacterial liquid is added to the mixture to perform microbial fermentation to prepare a black soldier fly-soybean meal fermented feed; The composite fermentation bacteria agent comprises: Bacillus subtilis, Bacillus licheniformis, Lactobacillus plantarum, Saccharomyces cerevisiae and Enterococcus faecalis.

2. The process for producing black soldier fly-soybean meal fermented feed according to claim 1, characterized in that: The bacterial liquid is prepared by mixing a composite fermentation bacterial agent with water in a weight ratio of 1:19, and the amount of bacterial liquid added is 1% to 3% by weight of the mixture obtained by compounding insect pulp and soybean meal.

3. The black soldier fly-soybean meal fermented feed production process according to claim 1, characterized in that: The black soldier fly larvae have a moisture content of 60%-70% and are ground into insect pulp by a colloid refiner.

4. The process for producing black soldier fly-soybean meal fermented feed according to claim 1, characterized in that: The soybean meal is soybean meal with husk, with a moisture content of 8% to 12%, and is crushed and passed through an 80-mesh sieve.

5. The process for producing black soldier fly-soybean meal fermented feed according to claim 1, characterized in that: The insect pulp and soybean meal are compounded in a weight ratio of 2: (1-2). The insect pulp and soybean meal are prepared in a weight ratio of 2: (1-2), mixed evenly, and a mixture with a water content of 40%-50% is obtained.

6. The black soldier fly-soybean meal fermented feed as claimed in claim 1 is used in the preparation of black-spotted frog feed, wherein the addition amount of the black soldier fly-soybean meal fermented feed is 40% to 45% based on the dry matter weight.

7. The use according to claim 6, characterized in that: The basic feed includes the following components by weight: 15 parts of fish meal, 19 parts of flour, 6 parts of rapeseed meal, 10 parts of peanut meal, 5 parts of cottonseed meal, 1 part of soybean lecithin, 0.3 parts of 50% choline chloride, 1.5 parts of monocalcium phosphate, 0.1 parts of vitamin premix, and 0.25 parts of mineral premix; Among them: Vitamin premix, calculated by total mass fraction 100%, consists of the following components: rice husk powder 30.7%, VC 40.0%, VA 1.5%, VB 11.3%, VB 22.0%, VB 6 2.4%, VB 12 0.8%, VD30.5%, VK3 3.5%, niacin 3.6%, folic acid 0.4%, D-biotin 0.8%, inositol 12.5%; The mineral premix, calculated based on the total mass fraction of 100%, consists of the following components: 42.4% zeolite, 26.7% rice husk powder, 1.0% CuSO4·5H2O, 15.0% MgSO4·7H2O, 6.7% FeSO4·7H2O, 3.5% ZnSO4·7H2O, 2.4% KCl, 0.9% MnSO4·7H2O, 0.7% CaH4I2O6, 0.5% Na2SeO3, and 0.2% CoSO4·7H2O.

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