Hybrid grouper low fish meal feed containing black soldier fly larvae powder

Abstract

The application discloses a hybrid grouper low-fish meal feed containing black soldier fly larvae powder, relates to the technical field of aquatic feed, and comprises the following steps: defatting and enzymolysis treatment of the black soldier fly larvae powder; mixing taurine, chenodeoxycholic acid and a nucleotide mixture to form a regulator; mixing fish meal, enzymolysis products and auxiliary materials and adding the regulator, and then performing conditioning, extrusion puffing, drying and post-spraying to obtain a feed product. Through the synergistic effect of the defatting and enzymolysis pretreatment and the bile acid metabolism regulator, the liver metabolism state and the feed utilization efficiency of the hybrid grouper can be improved under the condition that the black soldier fly larvae powder is used to replace fish meal at a high proportion.

Description

Technical Field

[0001] This invention belongs to the field of aquatic feed technology, and in particular relates to a low-fish meal feed for hybrid grouper containing black soldier fly larvae powder. Background Technology

[0002] Hybrid grouper (Epinephelus lanceolatus ♂×E. fuscoguttatus ♀) is an important marine aquaculture fish species in the coastal areas of southern my country, widely favored by aquaculture enterprises due to its rapid growth, delicious meat, and stable market price. In recent years, with the promotion and popularization of recirculating aquaculture systems (RAS) technology, the farming model of hybrid grouper is gradually shifting from traditional offshore cage culture to intensive land-based culture. RAS, through water treatment units such as mechanical filtration, biological purification, and ultraviolet disinfection, achieves the recycling of aquaculture water, offering advantages such as water conservation, land conservation, and environmental control, and has become an important development direction for high-end marine fish aquaculture. However, under this system, the stocking density is usually maintained at a high level (25-35 kg / m³), and the fish are confined to a limited space for extended periods, resulting in a corresponding increase in their physiological metabolic load and more stringent requirements for feed nutritional quality.

[0003] Fishmeal is the most important high-quality protein source in aquatic feed, possessing advantages such as balanced amino acid composition, good palatability, and low levels of anti-nutritional factors. For a long time, the proportion of fishmeal added to hybrid grouper compound feed has generally been high, typically between 35% and 45%. Due to the increasing scarcity of global fishery resources, fishmeal production has been declining year by year while prices have continued to rise. Finding sustainable protein sources to replace fishmeal has become a pressing technical problem for the aquatic feed industry. Black soldier fly larvae meal (BSFL) is an insect-derived protein raw material that has received widespread attention in recent years. Its crude protein content can reach 35%-45%, and it is rich in medium-chain fatty acids such as lauric acid. It features high feed conversion rate, short rearing cycle, and can be produced using organic waste such as kitchen waste, making it one of the potential raw materials for replacing fishmeal.

[0004] Currently, some studies have applied black soldier fly larvae meal to aquatic feed, confirming that it can partially replace fishmeal within a certain range without significantly negatively impacting fish growth performance. However, under recirculating aquaculture systems with high density, when the proportion of black soldier fly larvae meal replacing fishmeal exceeds a certain threshold, hybrid grouper exhibit decreased feed intake, reduced feed conversion efficiency, and pathological changes in liver tissue, significantly hindering aquaculture performance. This negative effect is closely related to the compositional characteristics of black soldier fly larvae meal: firstly, the larval exoskeleton contains a certain amount of chitin, and the fish digestive system has limited capacity to break down chitin; excessive intake may affect the digestion and absorption of nutrients in the intestines. Secondly, black soldier fly larvae meal has a high crude fat content (usually 25%-35%), and its fatty acid composition is predominantly saturated fatty acids, which differs significantly from the fatty acid composition of the fish's natural food organisms. These factors combined result in the fish liver facing an additional burden of fat metabolism when replaced at a high proportion.

[0005] This metabolic burden on the liver is particularly pronounced in high-density recirculating aquaculture systems (RAS). In RAS systems, the limited exchange rate of the water leads to the accumulation of fish excrement and metabolic products. Even after water treatment, water quality indicators fluctuate under high stocking densities, placing fish under continuous physiological stress. Under stress, the liver function and bile acid metabolism of fish are already under significant strain, affecting bile acid synthesis, secretion, and enterohepatic circulation efficiency. When a high proportion of black soldier fly larvae powder is introduced into the feed, its high saturated fatty acid composition increases the demand for bile acid emulsification during fat digestion and absorption. Furthermore, chitin may interfere with bile acid reabsorption in the intestines. The combined effect of these factors leads to an imbalance in the enterohepatic circulation of bile acids, consequently affecting the digestibility and utilization of fats and fat-soluble nutrients in the feed, further exacerbating the metabolic burden on the liver, ultimately resulting in liver tissue damage and decreased feed utilization.

[0006] To address the aforementioned issues, existing technologies primarily focus on reducing the proportion of black soldier fly larvae meal used, typically limiting its fishmeal replacement to below 20% to mitigate the impact on fish liver metabolism. However, this approach limits the potential of black soldier fly larvae meal to replace fishmeal in aquatic feed and fails to effectively alleviate the industry pressure of fishmeal resource shortages. Therefore, the following solution is proposed. Summary of the Invention

[0007] The purpose of this invention is to provide a low-fishmeal feed for hybrid grouper containing black soldier fly larvae powder. By pre-treating the black soldier fly larvae powder with defatting and enzymatic hydrolysis, and adding a bile acid metabolism regulator composition, the liver metabolism and feed utilization efficiency of hybrid grouper under recirculating aquaculture conditions can be improved. This solves the problem that when existing high proportions of black soldier fly larvae powder replace fishmeal, the physiological functions of fish are suppressed and the replacement ratio is difficult to increase.

[0008] To solve the above-mentioned technical problems, the present invention is achieved through the following technical solution:

[0009] This invention relates to a low-fishmeal feed for hybrid grouper containing black soldier fly larvae powder, comprising the following components:

[0010] Fish meal, black soldier fly larvae powder, soy protein concentrate, casein, shrimp meal, wheat flour, microcrystalline cellulose, fish oil, soybean oil, squid viscera powder, vitamin premix, mineral premix, choline chloride, calcium dihydrogen phosphate, and a composition of bile acid metabolism regulators.

[0011] The black soldier fly larvae powder is an enzymatically hydrolyzed black soldier fly larvae powder obtained after defatting pretreatment and enzymatic hydrolysis.

[0012] The bile acid metabolism regulator composition comprises taurine and chenodeoxycholic acid.

[0013] Furthermore, based on the total mass of the feed, the mass percentage of each component is as follows: fish meal 18-22%; enzymatically hydrolyzed black soldier fly larvae meal 13-20%; soybean protein concentrate 12-16%; casein 3-5%; shrimp meal 2-3%; wheat flour 8-12%; microcrystalline cellulose 1.5-2.5%; fish oil 5-9%; soybean oil 0.5-1.5%; squid viscera meal 1.2-1.8%; vitamin premix 0.8-1.2%; mineral premix 0.8-1.2%; choline chloride 0.4-0.6%; calcium dihydrogen phosphate 0.8-1.2%; and bile acid metabolism regulator composition 0.8-1.2%.

[0014] Furthermore, the enzymatically hydrolyzed black soldier fly larvae powder is composed of enzymatically hydrolyzed black soldier fly larvae active peptide powder and enzymatically hydrolyzed black soldier fly larvae residue powder, wherein the mass ratio of the enzymatically hydrolyzed black soldier fly larvae active peptide powder to the enzymatically hydrolyzed black soldier fly larvae residue powder is (8-12):(5-8).

[0015] Furthermore, the bile acid metabolism regulator composition comprises, by weight, the following components:

[0016] Taurine 25-35 parts, chenodeoxycholic acid 8-12 parts, nucleotide mixture 5-8 parts, L-carnitine 4-7 parts, vitamin C polyphosphate 3-5 parts, vitamin E acetate 2-4 parts, zeolite powder 20-30 parts.

[0017] Furthermore, the nucleotide mixture consists of inosinic acid, guanosine acid, cytidine acid and uridine acid in a mass ratio of 1:1:1:1.

[0018] Furthermore, the particle size D90 of the bile acid metabolism regulator composition is ≤20 micrometers.

[0019] Furthermore, the enzymatically hydrolyzed black soldier fly larvae powder is prepared by the following method:

[0020] Black soldier fly larvae powder was pre-treated by defatting in a supercritical carbon dioxide extraction device. The extraction pressure was 30 MPa, the extraction temperature was 45℃, and the extraction time was 2.5 hours to obtain defatted black soldier fly larvae powder.

[0021] The defatted black soldier fly larvae powder was added to a phosphate buffer solution at pH 7.0 at a material-to-liquid ratio of 1:5. A compound enzyme preparation was then added for enzymatic hydrolysis. The compound enzyme preparation consisted of alkaline protease, papain, and chitinase in a mass ratio of 2:1:1. The enzymatic hydrolysis conditions were 55°C, pH 7.0, and 3 hours. After enzymatic hydrolysis, the powder was centrifuged, concentrated by ultrafiltration, and spray-dried to obtain the enzymatically hydrolyzed black soldier fly larvae powder.

[0022] Furthermore, before the supercritical carbon dioxide extraction, the black soldier fly larvae are dried to a moisture content of 8±0.5% and pulverized to a particle size D90≤45 micrometers; the amount of the compound enzyme preparation added is 0.3% of the dry weight of the defatted black soldier fly larvae powder.

[0023] Furthermore, the fishmeal feed is prepared by a method comprising the following steps:

[0024] Mix the dry powder ingredients, excluding fish oil and soybean oil, to obtain a basic dry powder mixture;

[0025] The bile acid metabolism regulator composition is added to the basic dry powder mixture and mixed further to ensure that the regulator composition is evenly distributed in the dry powder system;

[0026] Fish oil, soybean oil and distilled water were added for conditioning at a temperature of 85±2℃ to obtain a dough-like material.

[0027] The dough-like material is fed into a twin-screw extruder for extrusion molding. The extrusion temperature is controlled in three zones: zone 1 at 70°C, zone 2 at 110°C, and zone 3 at 130°C. The die pressure is 3.5-4.0 MPa. After extrusion, the material is dried to reduce the moisture content to below 10%.

[0028] The dried granules are vacuum-sprayed with fish oil and then cooled to obtain the feed.

[0029] Furthermore, 30-40% of the fishmeal protein in the feed is replaced by black soldier fly larvae meal protein.

[0030] The present invention has the following beneficial effects:

[0031] This invention reduces the interference of chitin and saturated fatty acids in insect-derived raw materials on the digestive physiology of fish through defatting pretreatment and enzymatic hydrolysis of black soldier fly larvae powder. Simultaneously, it optimizes the overall feed formulation using a bile acid metabolism regulator composition containing a mixture of taurine, chenodeoxycholic acid, and nucleotides. The synergistic effect of these treatments effectively regulates the enterohepatic bile acid cycle in hybrid grouper under conditions where a high proportion of black soldier fly larvae powder replaces fishmeal, maintaining normal liver tissue morphology and keeping feed conversion efficiency at a good level. This invention improves the physiological health and feed utilization efficiency of farmed fish while maintaining a high black soldier fly larvae powder substitution ratio, reducing dependence on fishmeal resources and providing a feasible sustainable feed solution for aquaculture.

[0032] Of course, any product implementing this invention does not necessarily need to achieve all of the advantages described above at the same time. Attached Figure Description

[0033] To more clearly illustrate the technical solutions of the embodiments of the present invention, the accompanying drawings used in the description of the embodiments will be briefly introduced below. Obviously, the drawings described below are only some embodiments of the present invention. For those skilled in the art, other drawings can be obtained based on these drawings without creative effort.

[0034] Figure 1 This is a schematic flowchart of a method for preparing a low-fishmeal feed for hybrid grouper containing black soldier fly larvae powder according to the present invention.

[0035] Figure 2 These are histopathological images of the liver tissue of hybrid grouper from each treatment group in this invention.

[0036] Figure 3 These are pathological images of the stomach tissue of hybrid grouper from various treatment groups in this invention. Detailed Implementation

[0037] The technical solutions of the embodiments of the present invention will be clearly and completely described below with reference to the accompanying drawings. Obviously, the described embodiments are only some embodiments of the present invention, and not all embodiments. Based on the embodiments of the present invention, all other embodiments obtained by those skilled in the art without creative effort are within the scope of protection of the present invention.

[0038] Please see Figure 1 As shown, this invention relates to a method for preparing a low-fishmeal feed for hybrid grouper containing black soldier fly larvae powder, comprising the following steps:

[0039] Step S1: Degreasing pretreatment of Black Soldier Fly Larvae (BSFL)

[0040] Fresh black soldier fly larvae were harvested when the rearing substrate temperature dropped to 25±2℃. After harvesting, the larvae were rinsed three times with clean water for 5 minutes each time to remove any residual rearing substrate adhering to their bodies. After rinsing, the larvae were laid flat on a stainless steel mesh screen and allowed to drain at room temperature for 30 minutes.

[0041] The drained larvae were placed in a vacuum drying oven for preliminary drying at a temperature of 55±2℃ and a vacuum degree of -0.08 MPa for 4 hours, reducing the moisture content of the larvae to below 15%. The dried larvae were then coarsely pulverized using a high-speed universal pulverizer at a speed of 12000 rpm for 30 seconds per pulverization, with a 1-minute cooling interval. This process was repeated three times to ensure all the material passed through a 60-mesh sieve, yielding coarsely pulverized BSFL powder.

[0042] The coarsely pulverized BSFL powder was further pulverized in a low-temperature ultrafine pulverizer. The pulverization temperature was controlled below 10℃, the pulverization frequency was 45 Hz, and the pulverization time was 20 minutes, so that the powder particle size D90 ≤ 45 micrometers. The ultrafine pulverized BSFL powder was then transferred to a fluidized bed dryer and dried with hot air at 50℃ until the moisture content was 8±0.5%.

[0043] The dried ultrafine BSFL powder was loaded into the extraction vessel of a supercritical carbon dioxide extraction apparatus. The extraction pressure was 30 MPa, the extraction temperature was 45℃, the carbon dioxide flow rate was 20 kg / h, and the extraction time was 2.5 hours. The separation vessel pressure was 6 MPa, and the separation vessel temperature was 40℃, in order to remove some saturated fatty acids and free fatty acids from the BSFL. After extraction, the raffinate was collected, which is the defatted BSFL powder.

[0044] Step S2: Enzymatic hydrolysis of defatted BSFL powder

[0045] Take the defatted BSFL powder prepared in step S1, add it to pH 7.0 phosphate buffer at a material-to-liquid ratio of 1:5 (w / v), and disperse it for 15 minutes at 8000 rpm using a high-shear dispersing emulsifier to obtain a uniform BSFL suspension.

[0046] The suspension was placed in a constant-temperature water bath reaction vessel, heated to 55°C, and held at this temperature for 10 minutes to ensure uniform temperature. Then, a compound enzyme preparation was added at 0.3% of the dry weight of the defatted BSFL powder. This compound enzyme preparation consisted of alkaline protease (enzyme activity ≥200,000 U / g), papain (enzyme activity ≥100,000 U / g), and chitinase (enzyme activity ≥50,000 U / g) in a mass ratio of 2:1:1. The enzymatic hydrolysis reaction was carried out at 55°C and pH 7.0 for 3 hours, with continuous stirring at 100 rpm throughout the reaction. After the enzymatic hydrolysis reaction was completed, the temperature of the reaction vessel was raised to 90°C and held for 15 minutes to inactivate the enzyme.

[0047] The enzyme-inactivated hydrolysate was centrifuged at 10,000 rpm for 15 minutes using a disc centrifuge, and the supernatant and precipitate were collected separately. The supernatant was concentrated by ultrafiltration through an ultrafiltration membrane with a molecular weight cutoff of 10 kDa, and the retentate was collected. This retentate was rich in low molecular weight peptides and free amino acids. The ultrafiltration retentate was spray-dried at an inlet air temperature of 160℃ and an outlet air temperature of 80℃ to obtain enzymatically hydrolyzed BSFL active peptide powder. The precipitate was dried in an oven at 60℃ for 12 hours, pulverized, and passed through a 120-mesh sieve to obtain enzymatically hydrolyzed BSFL residue powder.

[0048] Step S3: Preparation of the bile acid metabolism regulator composition

[0049] Weigh the following raw materials according to the following mass proportions: 25-35 parts taurine, 8-12 parts chenodeoxycholic acid, 5-8 parts nucleotide mixture, 4-7 parts L-carnitine, 3-5 parts vitamin C polyphosphate, 2-4 parts vitamin E acetate, and 20-30 parts zeolite powder. The nucleotide mixture consists of inosinic acid (IMP), guanylic acid (GMP), cytidine monophosphate (CMP), and uridine monophosphate (UMP) in a mass ratio of 1:1:1:1.

[0050] The weighed components were sequentially added to a V-type mixer, with the mixer speed set to 25 rpm and the mixing time to 20 minutes. The uniformly mixed material was then subjected to micronization using an air jet mill at a grinding pressure of 0.6 MPa and a feed rate of 5 kg / h, resulting in a product particle size D90 ≤ 20 micrometers. The micronized mixture is the bile acid metabolism regulator composition.

[0051] Step S4: Preparation of the basic feed mixture

[0052] Weigh the following basic raw materials by weight percentage: fish meal 18-22%, enzymatically hydrolyzed BSFL active peptide powder prepared in step S2 8-12%, enzymatically hydrolyzed BSFL residue powder 5-8%, soybean protein concentrate 12-16%, casein 3-5%, shrimp meal 2-3%, wheat flour 8-12%, microcrystalline cellulose 1.5-2.5%, fish oil 3-6%, soybean oil 0.5-1.5%, squid viscera powder 1.2-1.8%, vitamin premix 0.8-1.2%, mineral premix 0.8-1.2%, choline chloride 0.4-0.6%, and calcium dihydrogen phosphate 0.8-1.2%.

[0053] All the dry powder ingredients, except for fish oil and soybean oil, from the weighed basic raw materials are put into a double cone high-efficiency mixer for premixing. The mixer speed is 15 rpm and the mixing time is 25 minutes to obtain the basic dry powder mixture.

[0054] Step S5: Preparation of compound feed containing regulator composition

[0055] Add the bile acid metabolism regulator composition prepared in step S3 to the basic dry powder mixture obtained in step S4 at a ratio of 0.8-1.2% of the total mass of the basic feed. Continue mixing in a three-dimensional motion mixer for 20 minutes, with the mixer speed set to 30 rpm, so that the regulator composition is evenly distributed in the dry powder system.

[0056] Preheat the weighed fish oil and soybean oil from step S4 in a 40°C water bath until completely melted. Then, add the oil mixture to the above-mentioned dry powder mixture in a thin stream while stirring continuously at a speed of 60 rpm. After the oil is added, add distilled water at 28-32% of the total mass of the dry powder. Use a twin-shaft paddle conditioner to condition the mixture at 85±2°C for 3 minutes to ensure thorough gelatinization and uniform mixing, forming a dough-like material with a certain degree of viscoelasticity.

[0057] Step S6: Extrusion puffing granulation

[0058] The dough-like material obtained in step S5 is fed into a twin-screw extruder via a screw conveyor for extrusion molding. The screw length-to-diameter ratio of the extruder is 20:1, and the screw speed is set to 280-320 rpm. The extrusion chamber temperature is controlled in three zones: zone 1 (feeding section) at 70℃, zone 2 (transition section) at 110℃, and zone 3 (discharge section) at 130℃. The die pressure is controlled at 3.5-4.0 MPa, and the die diameter is 3.0 mm. The residence time of the material in the extrusion chamber is 25-35 seconds.

[0059] The extruded strip material is cut by a rotary cutter, the cutter speed of which is adjusted in real time according to the extrusion speed to control the particle length at 4.0-5.5 mm. The freshly extruded feed particles have a high moisture content and are immediately fed into a multi-layer belt dryer for drying. The dryer's inlet air temperature is set to 105℃, the outlet air temperature to 65℃, and the drying time to 25 minutes, reducing the particle moisture content to below 10%.

[0060] Step S7, Post-coating treatment

[0061] The dried feed pellets were naturally cooled to room temperature in a cooling tower for 10 minutes. The cooled feed pellets were then placed in a vacuum sprayer, and a vacuum of -0.06 MPa was applied. Fish oil was sprayed at 2-3% of the feed pellet weight for 8 minutes, with the vacuum sprayer speed set to 20 rpm. After spraying, the vacuum was slowly released to atmospheric pressure, and mixing continued for 5 minutes to ensure the oil evenly coats the pellet surface.

[0062] The oil-coated pellets are transferred to a fluidized bed cooling dryer and cooled and dried with 25°C cold air at a velocity of 1.5 m / s for 10 minutes to solidify the oil on the pellet surface. Finally, the finished feed pellets are screened through a vibrating grading screen to remove unqualified pellets with a diameter less than 2.8 mm and greater than 5.8 mm. The qualified pellets are collected, placed in aluminum foil composite bags, filled with nitrogen, and sealed to produce the final BSFL-containing hybrid grouper low-fishmeal feed.

[0063] The specific application of this embodiment is as follows:

[0064] This embodiment aims to verify the practical application effect of the hybrid grouper feed containing black soldier fly larvae powder (BSFL) prepared using steps S1 to S7 of the present invention under high-density recirculating aquaculture conditions. Three parallel treatment groups were set up for comparison to demonstrate the effectiveness of the present invention in improving bile acid metabolism, maintaining liver health, and increasing feed conversion efficiency.

[0065] 1. Preparation of experimental feed

[0066] 1.1 Preparation of the first control diet (hereinafter referred to as control diet A)

[0067] Weigh the following raw materials by weight percentage: fish meal 40.0%, soy protein concentrate 15.0%, casein 5.0%, shrimp meal 2.5%, wheat flour 16.4%, microcrystalline cellulose 2.0%, fish oil 10.1%, soybean oil 1.0%, squid viscera meal 1.5%, vitamin premix 1.0%, mineral premix 1.0%, choline chloride 0.5%, and calcium dihydrogen phosphate 1.0%. The fish meal contains 57.0% crude protein and 6.1% crude fat.

[0068] All the dry powder ingredients except fish oil and soybean oil were added to a double cone high-efficiency mixer and mixed for 25 minutes at 15 rpm. Then, fish oil and soybean oil were added while stirring at 60 rpm. After the oils were added, distilled water was added at 30% of the total dry powder mass, and the mixture was conditioned at 85°C for 3 minutes using a twin-shaft paddle conditioner to obtain a dough-like material. This material was then extruded into shape using a twin-screw extruder. The screw speed was set to 300 rpm, the temperatures in the extrusion chamber were 70°C in zone one, 110°C in zone two, and 130°C in zone three, the die pressure was controlled at 3.8 MPa, and the die diameter was 3.0 mm. The extruded strips were cut by a rotary cutter to control the particle length at 4.5 mm. The extruded particles were dried in a multi-layer belt dryer at an inlet air temperature of 105°C for 25 minutes to reduce the moisture content to below 10%. After drying, the granules are cooled to room temperature in a cooling tower and placed in a vacuum sprayer. Fish oil is sprayed at 2.5% of the granule mass. After spraying, the granules are cooled and dried in a fluidized bed and then screened by a vibrating grading screen to obtain granules with a particle size of 2.8-5.8 mm. The granules are then packaged to obtain control feed A.

[0069] 1.2 Preparation of the second control diet (hereinafter referred to as control diet B)

[0070] Weigh the following raw materials by weight percentage: fish meal 23.0%, untreated BSFL coarse powder 19.5%, soy protein concentrate 15.0%, casein 4.0%, shrimp meal 2.5%, wheat flour 10.5%, microcrystalline cellulose 2.0%, fish oil 4.5%, soybean oil 1.0%, squid viscera powder 1.5%, vitamin premix 1.0%, mineral premix 1.0%, choline chloride 0.5%, and calcium dihydrogen phosphate 1.0%. The untreated BSFL coarse powder has a crude protein content of 35.0%, a crude fat content of 32.0%, and a crude ash content of 14.8%. In this formula, BSFL-derived protein replaces 35% of the fish meal protein.

[0071] Weigh all the dry powder raw materials except for fish oil and soybean oil according to the stated mass percentages, and add them to a double cone high-efficiency mixer and mix at 15 rpm for 25 minutes. Then add fish oil and soybean oil, and stir at 60 rpm until the oil is evenly distributed. Add distilled water at 30% of the total dry powder mass, and condition at 85°C for 3 minutes using a twin-shaft paddle conditioner. The resulting material is processed using the same extrusion puffing, drying, cooling, post-spraying, and sieving process parameters as in step 1.1 to obtain control feed B.

[0072] 1.3 Preparation of the Invented Feed (hereinafter referred to as Invented Feed T)

[0073] Prepared according to steps S1 to S7 of the present invention.

[0074] Step S1: Degreasing pretreatment of black soldier fly larvae powder: Fresh black soldier fly larvae were taken, rinsed three times with clean water for 5 minutes each time, drained for 30 minutes, and then placed in a vacuum drying oven. They were dried at 55℃ and -0.08 MPa for 4 hours until the moisture content was reduced to below 15%. The dried larvae were then pulverized using a high-speed universal pulverizer at 12000 rpm for 30 seconds each time, with a 1-minute cooling interval, repeated three times until all the material passed through a 60-mesh sieve, yielding coarsely pulverized BSFL powder. The coarsely pulverized BSFL powder was transferred to a low-temperature ultrafine pulverizer and pulverized at below 10℃ and 45 Hz for 20 minutes until the particle size D90 ≤ 45 micrometers. The ultrafine pulverized powder was then dried in a fluidized bed dryer with hot air at 50℃ until the moisture content was 8±0.5%. The dried ultrafine BSFL powder was loaded into the extraction vessel of a supercritical carbon dioxide extraction device and extracted for 2.5 hours at an extraction pressure of 30 MPa, an extraction temperature of 45℃, and a carbon dioxide flow rate of 20 kg / h. The separation vessel pressure was 6 MPa and the separation vessel temperature was 40℃. The residue was collected to obtain defatted BSFL powder.

[0075] Step S2: Enzymatic hydrolysis of defatted BSFL powder: Take the above defatted BSFL powder and add phosphate buffer solution at pH 7.0 at a material-to-liquid ratio of 1:5 (w / v). Disperse the powder using a high-shear dispersing emulsifier at 8000 rpm for 15 minutes to obtain a BSFL suspension. Place the suspension in a constant temperature water bath reaction vessel and heat it to 55°C. Add a compound enzyme preparation (composed of alkaline protease, papain, and chitinase in a mass ratio of 2:1:1) at 0.3% of the dry weight of the defatted BSFL powder. Hydrolyze the powder at 55°C, pH 7.0, and 100 rpm for 3 hours. After hydrolysis, raise the temperature of the reaction vessel to 90°C and incubate for 15 minutes to inactivate the enzyme. Centrifuge the hydrolysate at 10000 rpm for 15 minutes using a disc centrifuge, and collect the supernatant and precipitate separately. The supernatant was concentrated by ultrafiltration through an ultrafiltration membrane with a molecular weight cutoff of 10 kDa, and then spray-dried at an inlet air temperature of 160°C and an outlet air temperature of 80°C to obtain enzymatically hydrolyzed BSFL active peptide powder. The precipitate was dried in an oven at 60°C for 12 hours, pulverized, and passed through a 120-mesh sieve to obtain enzymatically hydrolyzed BSFL residue powder.

[0076] Step S3: Preparation of the bile acid metabolism regulator composition: Weigh out 30 parts by weight of taurine, 10 parts by weight of chenodeoxycholic acid, 6 parts by weight of a nucleotide mixture (inosine:guanosine:cytidine:uridine = 1:1:1:1), 5 parts by weight of L-carnitine, 4 parts by weight of vitamin C polyphosphate, 3 parts by weight of vitamin E acetate, and 25 parts by weight of zeolite powder. Add all the above components to a V-type mixer and mix at 25 rpm for 20 minutes. After uniform mixing, pass the mixture through an air jet mill at a pressure of 0.6 MPa and a feed rate of 5 kg / h for micronization treatment, so that the product particle size D90 ≤ 20 micrometers, thus obtaining the bile acid metabolism regulator composition.

[0077] Step S4, Preparation of the basic feed mixture: Weigh the following basic raw materials by mass percentage: fish meal 19.0%, enzymatically hydrolyzed BSFL active peptide powder prepared in step S2 10.0%, enzymatically hydrolyzed BSFL residue powder 6.5%, soybean protein concentrate 15.0%, casein 4.0%, shrimp meal 2.5%, wheat flour 10.5%, microcrystalline cellulose 2.0%, fish oil 4.5%, soybean oil 1.0%, squid viscera powder 1.5%, vitamin premix 1.0%, mineral premix 1.0%, choline chloride 0.5%, and calcium dihydrogen phosphate 1.0%. In this formula, the proportion of BSFL-derived protein (active peptide powder plus residue powder) replacing fish meal protein is also 35%. Add all the above dry powder raw materials except fish oil and soybean oil sequentially into a double cone high-efficiency mixer and mix at 15 rpm for 25 minutes to obtain the basic dry powder mixture.

[0078] Step S5: Preparation of the compound feed containing the regulator composition: Add the bile acid metabolism regulator composition prepared in Step S3 to the above-mentioned basic dry powder mixture at a ratio of 1.0% of the total mass of the basic feed, and continue mixing for 20 minutes at 30 rpm in a three-dimensional motion mixer. Preheat the fish oil and soybean oil weighed in Step S4 to a 40°C water bath until completely melted, then add them to the mixed dry powder in a thin stream while stirring at a speed of 60 rpm. After the oils are added, add distilled water at 30% of the total mass of the dry powder, and condition at 85°C for 3 minutes using a biaxial paddle conditioner to obtain a dough-like material.

[0079] Step S6, Extrusion and Pelletizing: The dough-like material obtained in step S5 is fed into a twin-screw extruder via a screw conveyor. The screw speed is set to 300 rpm, the temperature of zone 1 in the extrusion chamber is 70℃, zone 2 is 110℃, and zone 3 is 130℃, the die pressure is controlled at 3.8 MPa, and the die diameter is 3.0 mm. The extruded strip-shaped material is cut by a rotary cutter, and the particle length is controlled at 4.5 mm. The extruded granules are fed into a multi-layer belt dryer with an inlet air temperature of 105℃ and an outlet air temperature of 65℃ for 25 minutes to reduce the moisture content of the granules to below 10%.

[0080] Step S7, Post-coating treatment: The dried feed pellets are naturally cooled to room temperature in a cooling tower for 10 minutes. The cooled pellets are then placed in a vacuum sprayer, and a vacuum of -0.06 MPa is applied. Fish oil is sprayed at 2.5% of the pellet mass for 8 minutes at a speed of 20 rpm. After spraying, the vacuum is slowly released to atmospheric pressure, and mixing continues for 5 minutes. The oil-coated pellets are then transferred to a fluidized bed cooling dryer and cooled and dried with 25°C cold air (1.5 m / s) for 10 minutes. The finished pellets are screened by a vibrating grading sieve, and pellets with a diameter of 2.8-5.8 mm are collected, placed in aluminum foil composite bags, filled with nitrogen, and sealed to obtain the invented feed T.

[0081] The measured nutritional levels of the three experimental diets are shown in Table 1.

[0082] Table 1. Measured nutrient composition of the experimental diet (air-dried basis, %)

[0083] detection indicators Comparison Feed A Comparison Feed B Invented feed T crude protein 36.72 37.58 37.62 Crude fat 14.96 15.02 14.85 Coarse ash 10.54 12.37 11.96 crude fiber 7.86 9.12 7.45 Total energy (kJ / g) 17.21 17.38 17.29

[0084] 2. Experimental Subjects and Management

[0085] Juvenile hybrid grouper (Epinephelus lanceolatus ♂×E. fuscoguttatus ♀) from the same batch were obtained from a commercial hatchery in Hainan Province, with an average initial weight of 56.52 g ± 0.06 g. Before the experiment, the fish were transported to an indoor laboratory equipped with a recirculating aquaculture system (RAS), which included a protein skimmer, biofilter, UV sterilizer, and temperature control device. The rearing tanks were 500 L cylindrical fiberglass tanks, with 40 fish stocked per tank at an initial stocking density of 4.52 kg / m³. At the end of the experiment, the final stocking density of control feed A was approximately 8.7 kg / m³, while that of control feed B and the invented feed T was approximately 6.1–7.8 kg / m³. The stocking density fluctuations throughout the experimental period covered medium- to high-density rearing conditions.

[0086] The experiment consisted of three treatment groups: control feed group A, control feed group B, and the invented feed group T. Each treatment group had three replicates, for a total of nine culture units. Feeding was conducted twice daily (8:00 AM and 4:00 PM), with the amount of feed determined by whether the fish consumed within 30 minutes of feeding. Daily feed intake and the number of dead fish were recorded. The culture period was 56 days.

[0087] Water quality parameters were monitored twice daily (before feeding) during the aquaculture period using a portable multi-parameter instrument (HM-B200, Shandong Hengmei Electronic Technology Co., Ltd.). The controlled parameters were as follows: water temperature 30.0±0.5℃, dissolved oxygen 7.10±0.3 mg / L, salinity 29.5±0.5‰, pH 7.15±0.20, total ammonia nitrogen ≤0.3±0.2 mg / L, and nitrite nitrogen ≤0.05±0.02 mg / L. The water exchange rate of the circulating water system was 10% per day.

[0088] 3. Sample collection and index determination

[0089] After the culture experiment concluded, fish were withheld from food for 24 hours. All surviving fish were randomly removed from each culture tank and rapidly anesthetized using MS-222 (50 mg / L, tricaine mesylate solution, Sigma, USA). Each fish was weighed and its body length measured, and the number of surviving fish was counted. Ten fish were randomly selected from each tank, and their livers and intra-abdominal fat tissues were dissected, weighed, and recorded separately for calculating the hepatic body index (HSI) and intra-abdominal fat percentage (IPR).

[0090] Five fish were taken from each bucket, and blood was collected from the tail vein using a disposable syringe. The blood samples were left to stand at 4°C for 2 hours, then centrifuged at 3500 rpm for 10 minutes. The separated serum was stored at -80°C for the determination of serum biochemical indicators.

[0091] Take about 1 g each of liver, pancreas and stomach tissue, add pre-cooled physiological saline at a ratio of 1:9 (w / v), homogenize in an ice bath, centrifuge at 3500 rpm for 10 minutes, collect the supernatant, aliquot and store at -80℃ for digestive enzyme activity determination.

[0092] Growth performance indicators are calculated using the following formula:

[0093] Weight gain rate (WG, %) = (final weight - initial weight) / initial weight × 100;

[0094] Specific growth rate (SGR, % / d) = (ln final body weight - ln initial body weight) / number of days of rearing × 100;

[0095] Feed conversion ratio (FCR) = Dry weight of feed ingested (g) / Fish body weight gain (g);

[0096] Protein efficiency (PER) = Fish body weight gain (g) / Protein intake (g);

[0097] Liver body index (HSI, %) = Liver wet weight (g) / Total body weight (g) × 100;

[0098] Intra-abdominal fat percentage (IPR, %) = Intra-abdominal fat wet weight (g) / Total body weight (g) × 100.

[0099] Serum biochemical indicators were measured using a fully automated biochemical analyzer, including alanine aminotransferase (ALT), aspartate aminotransferase (AST), alkaline phosphatase (ALP), total bile acids (TBA), blood urea nitrogen (BUN), and uric acid (UA). All reagent kits used were purchased from Nanjing Jiancheng Bioengineering Institute.

[0100] Digestive enzyme activity assays: The activities of pepsin, lipase, and trypsin were quantified using commercially available kits (Nanjing Jiancheng Bioengineering Institute), and the assay procedures were strictly performed according to the kit instructions. A unit of pepsin activity was defined as the amount of enzyme that, per milligram of tissue protein, breaks down casein to produce 1 μg of tyrosine per minute at 37°C and pH 2.0 (U / mg protein). A unit of lipase activity was defined as the amount of enzyme that, per gram of tissue protein, hydrolyzes fat to produce 1 μmol of fatty acid per minute at 37°C (U / g). A unit of trypsin activity was defined as the amount of enzyme that, per milligram of tissue protein, hydrolyzes BAEE to produce 1 μmol of p-nitroaniline per minute at 37°C and pH 8.0 (U / mg protein).

[0101] Histopathological observation: Fixed liver and stomach tissues were dehydrated with graded ethanol, cleared with xylene, embedded in paraffin, and sectioned at a thickness of 4 μm. Hematoxylin and eosin (H&E) staining was performed, and the sections were mounted with neutral resin and observed and photographed under an optical microscope. The degree of liver tissue damage was assessed using a semi-quantitative scoring method. The scoring criteria were: 0 points: normal hepatocyte structure, clear sinusoids, no inflammatory cell infiltration; 1 point: occasional mild dilation of sinusoids, scattered single inflammatory cells; 2 points: significant dilation of sinusoids, mild local congestion, focal inflammatory cell infiltration <3 cells / high-power field; 3 points: significant congestion of sinusoids and central vein, multifocal inflammatory cell infiltration (3-10 cells / high-power field), some hepatocytes showing ballooning degeneration; 4 points: severe diffuse congestion, abundant inflammatory cell infiltration (>10 cells / high-power field), widespread hepatocyte ballooning degeneration and lipid droplet vacuoles. Five high-power fields (400×) were randomly selected from each section for scoring, and the average value was taken.

[0102] 4. Test Results

[0103] 4.1 Growth performance and feed utilization

[0104] The growth performance and feed utilization indicators of hybrid grouper in each treatment group are shown in Table 2. The final body weight of the control group B (untreated high-proportion BSFL replacement, 35% replacement) was 88.64 g, the weight gain rate was 56.78%, the specific growth rate was 1.14% / d, and the protein efficiency was 1.65, all significantly lower than those of control group A (P<0.05). The feed conversion ratio was 1.62, significantly higher than that of control group A (P<0.05), indicating that the untreated high-proportion BSFL replacement significantly inhibited fish growth.

[0105] In comparison, the final body weight of the invented feed group T was 104.37 g, the weight gain rate was 84.72%, the specific growth rate was 1.48% / d, and the protein efficiency was 2.03, all of which were significantly higher than those of the control feed group B (P<0.05), and there was no statistically significant difference between the invented feed group T and the control feed group A (P>0.05). The feed conversion ratio of the invented feed group T was 1.31, which was 19.1% lower than that of the control feed group B (1.62), while the protein efficiency was 23.0% higher than that of the control feed group B.

[0106] Survival rates in all treatment groups remained above 96%, with no significant differences between groups (P>0.05). Regarding the liver body index, the HSI of control diet B group was 3.44%, significantly higher than that of control diet A group (2.81%) and the invented diet T group (2.92%) (P<0.05), indicating that the high proportion of BSFL in the untreated diet led to compensatory liver enlargement, while the treatment measures of this invention effectively alleviated this phenomenon. There were no significant differences in the proportion of intra-abdominal fat among the groups (P>0.05).

[0107] Table 2 Comparison of growth performance and feed utilization of hybrid grouper in different treatment groups (mean ± standard error, n=3)

[0108] detection indicators Comparison Feed A Comparison Feed B Invented feed T Initial body weight (g) 56.51±0.04 56.54±0.05 56.50±0.06 Final body weight (g) 108.70±0.53 a 88.64±0.48 b 104.37±0.55 a Weight gain rate (%) 92.35±0.95 a 56.78±0.87 b 84.72±0.96 a Specific growth rate (% / d) 1.56±0.03 a 1.14±0.03 b 1.48±0.04 a Feed conversion ratio (FCR) 1.51±0.03 b 1.62±0.04 a 1.31±0.03 c Protein efficiency (PER) 1.80±0.04 b 1.65±0.03 c 2.03±0.05 a Liver body index (HSI, %) 2.81±0.07 b 3.44±0.08 a 2.92±0.06 b Intra-abdominal fat percentage (IPR, %) 4.12±0.10 4.01±0.09 4.08±0.11 Survival rate (%) 97.50±1.44 96.67±1.67 98.33±0.83

[0109] Note: Different lowercase letters in the same row indicate significant differences (P<0.05), and no letters indicate no significant differences (P>0.05). The same applies below.

[0110] 4.2 Serum biochemical indicators

[0111] The serum biochemical parameters of each treatment group are shown in Table 3. The ALT activity of the control diet B group was 218.36 U / L, the AST activity was 278.45 U / L, and the ALP activity was 982.31 U / L, all of which were significantly higher than those of the control diet A group (P<0.05), indicating that the high proportion of BSFL in the untreated diet had caused hepatocyte damage and bile excretion disorders.

[0112] Regarding the total bile acid (TBA) content, the control diet B group had a TBA content of 19.87 μmol / L, significantly higher than that of control diet A (12.26 μmol / L) and the invented diet T group (13.54 μmol / L) (P<0.05), demonstrating that the fish in the control diet B group exhibited significant bile acid enterohepatic circulation disorder and bile stasis. The TBA content in the invented diet T group was 31.8% lower than that in the control diet B group, but there was no significant difference compared to the control diet A group (P>0.05), indicating that the bile acid metabolism regulator composition of the present invention effectively improves the balance of bile acid synthesis, secretion, and reabsorption.

[0113] There were no significant differences in blood urea nitrogen (BUN) and uric acid (UA) among the treatment groups (P>0.05), indicating that the treatment measures of the present invention did not have an adverse effect on renal function.

[0114] Table 3 Comparison of serum biochemical indicators of hybrid grouper in different treatment groups

[0115] detection indicators Comparison Feed A Comparison Feed B Invented feed T ALT (U / L) 182.91±2.59 b 218.36±3.12 a 176.52±2.87 b AST (U / L) 229.81±3.04 b 278.45±3.56 a 235.12±3.21 b ALP (U / L) 339.68±9.24 c 982.31±12.65 a 458.73±10.43 b Total bile acid TBA (μmol / L) 12.26±0.47 b 19.87±0.62 a 13.54±0.51 b Blood urea nitrogen (BUN) (mg / dL) 20.97±1.29 22.35±1.41 21.02±1.33 Uric acid (UA) (μmol / L) 307.34±3.71 332.18±4.02 315.67±3.89

[0116] 4.3 Digestive enzyme activity

[0117] The results of digestive enzyme activity assays in liver and stomach tissues of each treatment group are shown in Table 4.

[0118] In liver tissue, the lipase activity in control feed B group was 0.58 U / g, which was not significantly different from control feed A group (0.61 U / g) (P>0.05), but significantly lower than that in the invented feed T group (0.87 U / g) (P<0.05). The liver lipase activity in the invented feed T group was 50.0% higher than that in control feed B group and significantly higher than that in control feed A group (P<0.05), indicating that the defatting pretreatment and enzymatic hydrolysis steps of the present invention effectively removed residual chitin and other inhibitory factors in BSFL, while taurine in the bile acid metabolism regulator promoted the activation effect of bile acids on lipase.

[0119] Regarding hepatic pepsin activity, the activity in control feed B (0.14 U / mg protein) was significantly lower than that in control feed A (0.19 U / mg protein) and the invented feed T (0.25 U / mg protein) (P<0.05), while the activity in the invented feed T was significantly higher than that in control feed A (P<0.05), confirming that the bioactive peptides produced by enzymatic hydrolysis improved gastric digestion of protein. There was no significant difference in trypsin activity among the groups (P>0.05).

[0120] In the gastric tissue, the pepsin activity in control feed B (0.25 U / mg protein) was significantly lower than that in control feed A (0.61 U / mg protein) and the invented feed T (0.55 U / mg protein) (P<0.05), while there was no significant difference between the invented feed T and control feed A (P>0.05). Regarding gastric lipase activity, the invented feed T (1.48 U / g) was significantly higher than that in control feed A (1.18 U / g) and control feed B (1.05 U / g) (P<0.05). There was no significant difference in gastric trypsin activity among the groups (P>0.05).

[0121] Table 4 Comparison of digestive enzyme activities in hybrid grouper from different treatment groups

[0122] organize detection indicators Comparison Feed A Comparison Feed B Invented feed T liver Pepsin (U / mg protein) 0.19±0.02 b 0.14±0.02 c 0.25±0.03 a liver Lipase (U / g) 0.61±0.02 b 0.58±0.02 b 0.87±0.03 a liver Trypsin (U / mg protein) 37.26±4.09 41.35±4.22 39.82±4.15 Stomach Pepsin (U / mg protein) 0.61±0.02 a 0.25±0.02 c 0.55±0.03 a Stomach Lipase (U / g) 1.18±0.04 b 1.05±0.04 b 1.48±0.05 a Stomach Trypsin (U / mg protein) 95.97±3.87 89.64±4.01 97.23±3.95

[0123] 4.4 Liver Histopathology

[0124] Observe H&E stained sections under a light microscope (e.g.) Figure 2 (As shown). In contrast, the hepatocytes in group A of the control diet had good structure, clear cell boundaries and clearly visible nuclei, with no obvious pathological changes, and the damage score was 0.3±0.2.

[0125] Compared with the liver tissue of the feed group B, the liver tissue showed pathological damage: mild congestion of the hepatic sinusoids and hepatic veins, occasional focal infiltration of lymphocytes in local liver lobules, and significant swelling of hepatocytes, with a damage score of 3.2±0.4.

[0126] The hepatocytes in the T group of the invention feed had good structure, clear and distinguishable cell boundaries, no obvious dilation or congestion of the hepatic sinusoids, and neatly arranged hepatocyte nuclei. The overall histological appearance was close to that of the control group A, and the damage score was 0.6±0.2, which was significantly lower than that of the control group B (P<0.001).

[0127] Histopathological observation of the stomach tissue showed (e.g.) Figure 3 As shown in the diagram): In contrast, group B showed a decrease in the number of mucous glands in the upper part of the lamina propria and an increase in eosinophilic glands in the lower part, accompanied by local lymphocyte infiltration. Furthermore, a small number of eosinophils were observed in the muscularis mucosae, and local vascular dilation was observed in the submucosa, suggesting local inflammatory cell infiltration. In contrast, groups A and T showed well-developed gastric mucosal structures with orderly arranged epithelial cells and no obvious signs of shedding. The glands in the lamina propria were abundant and densely distributed, with normal morphology and structure, and no inflammatory cell infiltration was found.

[0128] 5. Summary of Results of This Example

[0129] The results of this embodiment show that:

[0130] Under high-density recirculating aquaculture conditions, untreated 35% BSFL replacing fishmeal feed (control feed B) significantly inhibited the growth of hybrid grouper, with a specific growth rate reduced by 26.9% and feed conversion ratio increased by 7.3% compared to control feed A. It also caused severe liver dysfunction (significantly elevated ALT, AST, and ALP, and a 62.1% increase in total bile acids) and histopathological damage.

[0131] The feed T prepared using steps S1 to S7 of this invention, through defatting pretreatment of BSFL, enzymatic hydrolysis treatment, and the addition of a bile acid metabolism regulator composition, restores the specific growth rate of hybrid grouper to a level that is not significantly different from that of control feed A at the same replacement ratio (35%). The feed conversion rate is reduced by 19.1% compared with control feed B, and the protein efficiency is increased by 23.0%.

[0132] This invention effectively solves the enterohepatic circulation disorder of bile acids caused by high-proportion BSFL replacement. The serum total bile acid content in the T group of the invention feed was reduced by 31.8% compared with the control feed B group, returning to the normal physiological range; the liver lipase activity was increased by 50.0%, and the liver tissue pathological damage score was reduced from 3.2 to 0.6.

[0133] In the description of this specification, references to terms such as "an embodiment," "example," "specific example," etc., indicate that a specific feature, structure, material, or characteristic described in connection with that embodiment or example is included in at least one embodiment or example of the invention. In this specification, illustrative expressions of the above terms do not necessarily refer to the same embodiment or example. Furthermore, the specific features, structures, materials, or characteristics described may be combined in any suitable manner in one or more embodiments or examples.

[0134] The preferred embodiments of the present invention disclosed above are merely illustrative of the invention. These preferred embodiments do not exhaustively describe all details, nor do they limit the invention to the specific implementations described. Clearly, many modifications and variations can be made based on the content of this specification. This specification selects and specifically describes these embodiments to better explain the principles and practical applications of the invention, thereby enabling those skilled in the art to better understand and utilize the invention. The invention is limited only by the claims and their full scope and equivalents.

Claims

1. A low-fishmeal feed for hybrid grouper containing black soldier fly larvae meal, characterized in that, The fishmeal feed contains the following components: Fish meal, black soldier fly larvae powder, soy protein concentrate, casein, shrimp meal, wheat flour, microcrystalline cellulose, fish oil, soybean oil, squid viscera powder, vitamin premix, mineral premix, choline chloride, calcium dihydrogen phosphate, and a composition of bile acid metabolism regulators. The black soldier fly larvae powder is an enzymatically hydrolyzed black soldier fly larvae powder obtained after defatting pretreatment and enzymatic hydrolysis. The bile acid metabolism regulator composition comprises taurine and chenodeoxycholic acid.

2. The hybrid grouper low-fishmeal feed containing black soldier fly larvae powder according to claim 1, characterized in that, The percentage content of each component by total feed weight is as follows: fish meal 18-22%; enzymatically hydrolyzed black soldier fly larvae meal 13-20%; soybean protein concentrate 12-16%; casein 3-5%; shrimp meal 2-3%; wheat flour 8-12%; microcrystalline cellulose 1.5-2.5%; fish oil 5-9%; soybean oil 0.5-1.5%; squid viscera meal 1.2-1.8%; vitamin premix 0.8-1.2%; mineral premix 0.8-1.2%; choline chloride 0.4-0.6%; calcium dihydrogen phosphate 0.8-1.2%; and bile acid metabolism regulator composition 0.8-1.2%.

3. The hybrid grouper low-fishmeal feed containing black soldier fly larvae powder according to claim 1, characterized in that, The enzymatically hydrolyzed black soldier fly larvae powder is composed of enzymatically hydrolyzed black soldier fly larvae active peptide powder and enzymatically hydrolyzed black soldier fly larvae residue powder, and the mass ratio of the enzymatically hydrolyzed black soldier fly larvae active peptide powder to the enzymatically hydrolyzed black soldier fly larvae residue powder is (8-12):(5-8).

4. The hybrid grouper low-fishmeal feed containing black soldier fly larvae powder according to claim 1, characterized in that, The bile acid metabolism regulator composition comprises the following components in parts by weight: Taurine 25-35 parts, chenodeoxycholic acid 8-12 parts, nucleotide mixture 5-8 parts, L-carnitine 4-7 parts, vitamin C polyphosphate 3-5 parts, vitamin E acetate 2-4 parts, zeolite powder 20-30 parts.

5. A low-fishmeal feed for hybrid grouper containing black soldier fly larvae powder according to claim 4, characterized in that, The nucleotide mixture consists of inosinic acid, guanosine, cytidine, and uridine in a mass ratio of 1:1:1:

1.

6. The hybrid grouper low-fishmeal feed containing black soldier fly larvae powder according to claim 1, characterized in that, The particle size D90 of the bile acid metabolism regulator composition is ≤20 micrometers.

7. A low-fishmeal feed for hybrid grouper containing black soldier fly larvae powder according to claim 1, characterized in that, The enzymatically hydrolyzed black soldier fly larvae powder was prepared by the following method: Black soldier fly larvae powder was pre-treated by defatting in a supercritical carbon dioxide extraction device. The extraction pressure was 30 MPa, the extraction temperature was 45℃, and the extraction time was 2.5 hours to obtain defatted black soldier fly larvae powder. The defatted black soldier fly larvae powder was added to a phosphate buffer solution at pH 7.0 at a material-to-liquid ratio of 1:

5. A compound enzyme preparation was then added for enzymatic hydrolysis. The compound enzyme preparation consisted of alkaline protease, papain, and chitinase in a mass ratio of 2:1:

1. The enzymatic hydrolysis conditions were 55°C, pH 7.0, and 3 hours. After enzymatic hydrolysis, the powder was centrifuged, concentrated by ultrafiltration, and spray-dried to obtain the enzymatically hydrolyzed black soldier fly larvae powder.

8. A low-fishmeal feed for hybrid grouper containing black soldier fly larvae powder according to claim 7, characterized in that, Before the supercritical carbon dioxide extraction, the black soldier fly larvae are dried to a moisture content of 8±0.5% and pulverized to a particle size D90≤45 micrometers; the amount of the compound enzyme preparation added is 0.3% of the dry weight of the defatted black soldier fly larvae powder.

9. A low-fishmeal feed for hybrid grouper containing black soldier fly larvae powder according to claim 1, characterized in that, The fishmeal feed is prepared by a method comprising the following steps: Mix the dry powder ingredients, excluding fish oil and soybean oil, to obtain a basic dry powder mixture; The bile acid metabolism regulator composition is added to the basic dry powder mixture and mixed further to ensure that the regulator composition is evenly distributed in the dry powder system; Fish oil, soybean oil and distilled water were added for conditioning at a temperature of 85±2℃ to obtain a dough-like material. The dough-like material is fed into a twin-screw extruder for extrusion molding. The extrusion temperature is controlled in three zones: zone 1 at 70°C, zone 2 at 110°C, and zone 3 at 130°C. The die pressure is 3.5-4.0 MPa. After extrusion, the material is dried to reduce the moisture content to below 10%. The dried granules are vacuum-sprayed with fish oil and then cooled to obtain the feed.

10. A low-fishmeal feed for hybrid grouper containing black soldier fly larvae powder according to claim 1, characterized in that, 30-40% of the fishmeal protein in the feed was replaced by black soldier fly larvae protein.

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