CN115812871A

CN115812871A - Portunus trituberculatus feed and preparation method thereof

Info

Publication number: CN115812871A
Application number: CN202211327403.8A
Authority: CN
Inventors: 谢适超; 周歧存; 金敏; 焦乐飞; 孙蓬
Original assignee: Ningbo University
Current assignee: Ningbo University
Priority date: 2022-10-27
Filing date: 2022-10-27
Publication date: 2023-03-21

Abstract

The invention provides a portunus trituberculatus feed, which contains 9-10% of low gossypol cottonseed protein concentrate by mass percent, and specifically comprises the following raw materials by mass percent: 9-10% of low gossypol cottonseed protein concentrate, 17-22% of soybean meal, 20-25% of flour, 0.5-1% of arginine, 0.1-0.2% of lysine, 0.05-0.1% of methionine, 1-3% of calcium dihydrogen phosphate, 0.2-0.4% of choline chloride, 0.8-1.2% of vitamin premix, 1.5-2.5% of mineral premix, 2.5-3.5% of fish oil, 1.5-2.5% of soybean lecithin, 1.5-2.5% of sodium alginate, 3-4% of carboxymethyl cellulose and the balance of fish meal.

Description

Portunus trituberculatus feed and preparation method thereof

Technical Field

The invention belongs to the technical field of cultivation, and particularly relates to a portunus trituberculatus feed and a preparation method thereof.

Background

Portunus trituberculatus, an important economic cultured crab in coastal areas of the east of China, is deeply favored due to delicious taste. In 2021, the yield of the swimming crabs in mariculture reaches 105283t, and is mainly concentrated in Fujian, jiangsu, zhejiang, shandong, guangdong and other provinces. At present, both fishes and shrimps have mature commercial feeds to meet the nutritional requirements of high-density culture. In comparison, the feeding of the seawater crabs is mainly performed on low-value fishes, shrimps and shellfish, the quantity and the quality are unstable, pathogens are easily brought in, the water quality is spoiled, and the immunity of cultured animals is reduced. The prawn feed is directly used for feeding, the effect is not high, and the price is high.

Fish meal is a high-quality protein source which is most widely applied in aquaculture, has balanced nutrition and is a main protein source of feed for a plurality of carnivorous and marine fishes. The fish meal is mainly prepared by processing some low-economic value fished fishes through means of steaming, crushing and the like, the fishing amount of fishery in the world is reduced, but the aquaculture industry increases year by year for aquatic products supplied by the food industry, the fish meal is not in demand, and the price is increased year by year. In order to meet the theme of sustainable resource utilization and environmental protection, the aquaculture industry must develop alternatives to fish meal to mitigate its dependence.

Cottonseed, the third family of soya seeds in the world (second only to soybean and rapeseed). In 2019 to 2020, the global cottonseed yield has reached 4484 ten thousand tons. India, china and the united states are the three major cotton producing countries. By 2020, cotton yields in these three countries will account for around 60% of the world's yield. Cottonseed meal is a by-product of cottonseed oil, contains 38% -45% of protein, and is less expensive than beans and fish meal. However, the main reason why the use of cottonseed meal in aquatic products is limited is the anti-nutritional factors such as gossypol. Cottonseed meal typically contains about 400-800mg of free gossypol per kg and is toxic to animal organs and tissues. Free gossypol is easily combined with lysine, so that the activity of the lysine is reduced, and the palatability and the protein conversion rate of the feed are reduced.

Therefore, an economic and good-effect commercial feed is urgently needed to be found to meet the blue crab breeding industry.

Disclosure of Invention

The invention aims to provide a portunus trituberculatus feed, which adopts low gossypol cottonseed concentrated protein to partially replace fish meal, thereby reducing the using amount of the fish meal, not only reducing the sample cost of the portunus trituberculatus, but also promoting the growth of the portunus trituberculatus, the anabolism of protein and the expression of key genes of Toll and IMD immune pathways.

The purpose of the invention can be realized by the following technical scheme: the portunus trituberculatus feed contains 9-10% of low gossypol cottonseed protein concentrate by mass percent.

The TOR pathway is a main nutrient sensitive pathway, receives and integrates amino acid, growth factor, energy state and stress signal in cells, regulates the growth and metabolism of the cells, and has been proved by a great deal of researches in fish and crustacean that the protein anabolism of the TOR pathway is closely related, the immune system of shrimps and crabs is lack of T lymphocytes and B lymphocytes compared with that of higher vertebrates, cannot rely on complement and antibody systems to induce acquired immune response reaction, but can rely on the innate immune system to survive and evolve in the environment filled with pathogenic microorganisms. The main immunization modes of crustaceans are divided into cellular immunity and humoral immunity. Cellular immunity causes the body to generate immune responses such as apoptosis, phagocytosis, nodule, package and the like through different types of blood cells; humoral immunity is mainly performed by active ingredients of body fluids such as a phenol oxidase system, antimicrobial peptides, and lectins. With the continuous development of molecular technology, the genome of shrimps and crabs is continuously perfected, and pathways and key genes related to immunity are successively discovered. The Toll and IMD pathways are key pathways for shrimps and crabs to recognize pathogenic microorganisms and induce the synthesis and secretion of antibacterial peptide and the like, and are similar to TLR4 and TNRF-1 pathways of higher animals.

Further, the feed additive comprises the following raw materials in percentage by mass: 9-10% of low gossypol cottonseed protein concentrate, 17-22% of soybean meal, 20-25% of flour, 0.5-1% of arginine, 0.1-0.2% of lysine, 0.05-0.1% of methionine, 1-3% of calcium dihydrogen phosphate, 0.2-0.4% of choline chloride, 0.8-1.2% of vitamin premix, 1.5-2.5% of mineral premix, 2.5-3.5% of fish oil, 1.5-2.5% of soybean phospholipid, 1.5-2.5% of sodium alginate, 3-4% of carboxymethyl cellulose and the balance of fish meal.

Further, the feed additive comprises the following raw materials in percentage by mass: 9.34 percent of low gossypol cottonseed protein concentrate, 20 percent of soybean meal, 22.11 percent of flour, 0.87 percent of arginine, 0.14 percent of lysine, 0.08 percent of methionine, 2 percent of monocalcium phosphate, 0.3 percent of choline chloride, 1 percent of vitamin premix, 2 percent of mineral premix, 2.79 percent of fish oil, 2 percent of soybean lecithin, 2 percent of sodium alginate, 3.37 percent of carboxymethyl cellulose and the balance of fish meal.

Further, the vitamin premix comprises the following components in percentage by mass: 0.1-0.2% of retinyl acetate, 0.1-0.2% of cholecalciferol, 1-3% of all-rac-alpha-tocopherol, 0.5-1% of menadione, 1-2% of thiamine, 0.7-0.8% of riboflavin, 2-4% of ascorbic acid, 1-1.5% of pyridoxine hydrochloride, 0.1-0.3% of cyanocobalamin, 3-5% of folic acid, 1.2-1.3% of biotin, 3.5-4.5% of nicotinic acid, 1.5-1.8% of D-calcium pantothenate, 20-21% of inositol and the balance of carboxymethyl cellulose.

Further, the vitamin premix comprises the following components in percentage by mass: 0.12% retinyl acetate, 0.12% cholecalciferol, 2% all-rac-alpha-tocopherol, 0.9% menadione, 1.1% thiamine, 0.75% riboflavin, 3% ascorbic acid, 1.2% pyridoxine hydrochloride, 0.2% cyanocobalamin, 4% folic acid, 1.25% biotin, 4.04% nicotinic acid, 1.6% d-calcium pantothenate, 20.4% inositol, the balance carboxymethylcellulose.

Further, the mineral premix comprises the following components in percentage by mass: 0.4-0.5% of ferric citrate, 0.9-1.0% of zinc sulfate heptahydrate, 0.4-0.5% of manganese sulfate monohydrate, 0.6-0.7% of copper sulfate pentahydrate, 23-24% of manganese sulfate heptahydrate, 23-24% of potassium dihydrogen phosphate, 12-14% of sodium dihydrogen phosphate, 3-4% of calcium lactate pentahydrate, 0.1-0.2% of cobalt chloride hexahydrate and the balance of zeolite powder.

Further, the mineral premix comprises the following components in percentage by mass: 0.457 percent of ferric citrate, 0.943 percent of zinc sulfate heptahydrate, 0.414 percent of manganese sulfate monohydrate, 0.661 percent of copper sulfate pentahydrate, 23.9 percent of manganese sulfate heptahydrate, 23.3 percent of monopotassium phosphate, 13.7 percent of sodium dihydrogen phosphate, 3.4 percent of calcium lactate pentahydrate, 0.136 percent of cobalt chloride hexahydrate and the balance of zeolite powder.

Further, the extraction method of the low gossypol cottonseed protein concentrate comprises the following steps:

s1, husking delinted cottonseed to obtain cottonseed kernels, and then fermenting and finely dividing the obtained cottonseed kernels;

s2, softening and flaking the hulled cotton seeds obtained in the step S1 to obtain embryo pieces, and then carrying out ventilation treatment;

s3, preparing a mixed solvent, namely feeding the embryo slices obtained in the step S2 into a continuous sprayer, spraying the mixed solvent for 40-45min to soak the surfaces of the embryo slices with the mixed solvent, then putting the embryo slices into the mixed solvent again to be soaked for 20-30min to complete the dephenolization process, removing the mixed solvent after soaking is finished, and then extracting and drying to obtain the low gossypol cottonseed concentrated protein.

In order to reduce the toxicity of the gossypol, the invention adopts measures such as softening, embryo rolling, solvent extraction and the like to obtain the low gossypol cottonseed protein concentrate, wherein a large amount of plant anti-nutritional factors and crude fibers which influence the mouthfeel can be removed and eliminated to the greatest extent through fermentation and fine separation treatment processes, and the protein denaturation can be greatly reduced by ventilation treatment, so that the obtained low gossypol cottonseed protein concentrate can replace fish meal to reduce the using amount of fish meal, reduce the feeding cost of the portunus trituberculatus, and promote the growth of the portunus trituberculatus, the protein anabolism and the key gene expression of Toll and IMD immune channels.

Preferably, in the step S2, the thickness of the obtained blank is less than 5mm, and the environmental temperature during ventilation treatment is 55-60 ℃;

preferably, in the step S3, the mixed solvent is prepared from 90% methanol and 97% n-hexane in a volume ratio of 1:1, the extraction temperature is 60 ℃, and the drying temperature is 85 ℃.

Further, the nutritional ingredients in the portunus trituberculatus feed comprise the following components in percentage by mass: 45-48% of crude protein, 8-9% of crude fat, 14-15% of carbohydrate, 10-12% of crude fiber, 10-11% of ash, 9-10% of water and 0.0065-0.007% of free gossypol. When the feed is designed, the requirements of each breeding animal on protein fat are not the same as much, and the nutritional ingredients of the feed for the portunus trituberculatus are designed according to the invention, so that the nutritional ingredient requirements in the feeding of the portunus trituberculatus can be ensured.

The invention also aims to provide a preparation method of the portunus trituberculatus feed, which specifically comprises the following steps:

s1, dissolving soybean lecithin in fish oil in a formula ratio in advance, and uniformly mixing;

s2, crushing other raw materials except the soybean lecithin and the fish oil in advance, sieving the crushed raw materials with a 60-mesh sieve, accurately weighing the raw materials according to a formula, uniformly mixing various feed components in a step-by-step manner from small to large according to the specific gravity, adding a mixture of the soybean lecithin and the fish oil, fully mixing the mixture, adding water accounting for 30% of the mass of the raw materials, and fully mixing the mixture;

and S3, putting the uniformly mixed feed raw materials obtained in the step S2 into an automatic granulator to prepare feed particles with the particle size of 2-4mm, and naturally drying to obtain the portunus trituberculatus feed.

Compared with the prior art, the invention has the following advantages:

the feed formula disclosed by the invention is balanced in nutrition, the use amount of the fish meal is reduced by partially replacing the fish meal with the low gossypol cottonseed concentrated protein, the feeding cost of the portunus trituberculatus is reduced, and the feed has a promoting effect on the growth of the portunus trituberculatus, the protein anabolism and the expression of key genes of Toll and IMD immune pathways.

Secondly, the economic portunus trituberculatus feed provided by the invention has sufficient raw material sources and can be produced in a large scale. The cottonseed meal is used as a non-grain protein source, the detoxification means of the cottonseed meal is mature, and the cottonseed meal is widely applied in factories.

Thirdly, the invention provides a widely sustainable economic novel feed formula for the blue crab in the growing period, and reduces the environmental pollution caused by feeding fresh and alive baits.

Drawings

FIG. 1 shows the effect of different feeds with low gossypol cottonseed protein concentrate content on the expression of the genes of the hepatopancreas (tor, s6k1, s6 and akt) of Portunus trituberculatus;

FIG. 2 shows the effect of different feeds with low gossypol cottonseed protein concentrate content on the expression of the genes of the hepatopancreas of Portunus trituberculatus 4ebp1, eif4e1a, eif4e2 and eif4e 3;

FIG. 3 shows the effect of feed with different low gossypol cottonseed protein concentrates on the hepatopancreas Toll and IMD pathway key gene expression of blue crab;

FIG. 4 shows the effect of feed with different gossypol cottonseed protein concentrates on the diameter of the vacuole of the hepatopancreas tissue of portunus trituberculatus

FIG. 5 shows the effect of different low gossypol cottonseed protein concentrate contents on the height of fat droplets in the liver pancreatic tissue R cells of Portunus trituberculatus.

Detailed Description

In order to make the aforementioned objects, features and advantages of the present invention comprehensible, embodiments accompanied with figures are described in detail below.

Example 1:

the portunus trituberculatus feed consists of the following raw materials in percentage by mass: 9% of low gossypol cottonseed protein concentrate, 17% of soybean meal, 20% of flour, 0.5% of arginine, 0.1% of lysine, 0.05% of methionine, 1% of monocalcium phosphate, 0.2% of choline chloride, 0.8% of vitamin premix, 1.5% of mineral premix, 2.5% of fish oil, 1.5% of soybean lecithin, 1.5% of sodium alginate, 3% of carboxymethyl cellulose and the balance of fish meal; the vitamin premix comprises the following components: 0.1% retinyl acetate, 0.1% cholecalciferol, 1% all-rac-alpha-tocopherol, 0.5% menadione, 1% thiamine, 0.7% riboflavin, 2% ascorbic acid, 1% pyridoxine hydrochloride, 0.1% cyanocobalamin, 3% folic acid, 1.2% biotin, 3.5% niacin, 1.5% d-calcium pantothenate, 20% inositol, the balance carboxymethylcellulose, wherein the mineral premix consists of: 0.4% of ferric citrate, 0.9% of zinc sulfate heptahydrate, 0.4% of manganese sulfate monohydrate, 0.6% of copper sulfate pentahydrate, 23% of manganese sulfate heptahydrate, 23% of monopotassium phosphate, 12% of sodium dihydrogen phosphate, 3% of calcium lactate pentahydrate, 0.1% of cobalt chloride hexahydrate and the balance of zeolite powder.

The extraction method of the low gossypol cottonseed protein concentrate in the raw materials comprises the following steps: s1, husking delinted cottonseed to obtain cottonseed kernels, and then fermenting and finely dividing the obtained cottonseed kernels;

s2, softening and flaking the hulled cottonseed obtained in the step S1 to obtain a blank sheet with the thickness of less than 5mm, and then ventilating at the ambient temperature of 55-60 ℃;

s3, preparing a mixed solvent prepared from 90% methanol and 97% n-hexane in a volume ratio of 1:1, feeding the embryo piece obtained in the step S2 into a continuous sprayer, spraying the mixed solvent for 40-45min to enable the surface of the embryo piece to be soaked in the mixed solvent, then placing the embryo piece into the mixed solvent again to be soaked for 20-30min to complete the dephenolization process, removing the mixed solvent after soaking is finished, and then extracting at 60 ℃ and drying at 85 ℃ to obtain the low-gossypol cottonseed concentrated protein.

In the embodiment, the nutritional ingredients in the feed for the portunus trituberculatus comprise the following components in percentage by mass: 45% of crude protein, 8% of crude fat, 14% of carbohydrate, 10% of crude fiber, 10% of ash, 9% of water and 0.0065% of free gossypol.

A preparation method of a portunus trituberculatus feed comprises the following steps: s1, dissolving soybean lecithin in fish oil in a formula ratio in advance, and uniformly mixing;

s2, crushing other raw materials except the soybean lecithin and the fish oil in advance, sieving the crushed raw materials with a 60-mesh sieve, accurately weighing the raw materials according to a formula, uniformly mixing various feed components in a step-by-step manner from small to large according to the specific gravity, adding a mixture of the soybean lecithin and the fish oil, fully mixing the mixture, adding water with the mass being 30% of the mass of the raw materials, and fully mixing the mixture;

and S3, putting the uniformly mixed feed raw materials obtained in the step S2 into an automatic granulator to prepare feed particles with the particle size of 2mm, and naturally drying the feed particles to obtain the portunus trituberculatus feed.

Example 2

The portunus trituberculatus feed consists of the following raw materials in percentage by mass: 9.2% of low gossypol cottonseed protein concentrate, 18% of soybean meal, 22% of flour, 0.65% of arginine, 0.12% of lysine, 0.06% of methionine, 1.5% of monocalcium phosphate, 0.25% of choline chloride, 0.9% of vitamin premix, 1.7% of mineral premix, 2.8% of fish oil, 1.8% of soybean lecithin, 1.8% of sodium alginate, 3.2% of carboxymethyl cellulose and the balance of fish meal, wherein the vitamin premix consists of the following components: 0.12% retinyl acetate, 0.12% cholecalciferol, 1.2% all-rac-alpha-tocopherol, 0.6% menadione, 1.2% thiamine, 0.72% riboflavin, 2.5% ascorbic acid, 1.1% pyridoxine hydrochloride, 0.15% cyanocobalamin, 3.5% folic acid, 1.22% biotin, 3.7% nicotinic acid, 1.6% d-calcium pantothenate, 20.2% inositol, balance carboxymethylcellulose, the mineral premix consisting of: 0.42 percent of ferric citrate, 0.92 percent of zinc sulfate heptahydrate, 0.42 percent of manganese sulfate monohydrate, 0.62 percent of copper sulfate pentahydrate, 23.2 percent of manganese sulfate heptahydrate, 23.2 percent of potassium dihydrogen phosphate, 12.5 percent of sodium dihydrogen phosphate, 3.2 percent of calcium lactate pentahydrate, 0.12 percent of cobalt chloride hexahydrate and the balance of zeolite powder.

In the embodiment, the nutritional ingredients in the portunus trituberculatus feed comprise the following components in percentage by mass: 46 percent of crude protein, 8.2 percent of crude fat, 14.2 percent of carbohydrate, 10.5 percent of crude fiber, 10.2 percent of ash, 9.2 percent of water and 0.0066 percent of free gossypol, and the requirements of each cultured animal on the protein fat are not the same when designing the feed.

The portunus trituberculatus feed is prepared by the following steps: s1, dissolving soybean lecithin in fish oil in a formula ratio in advance, and uniformly mixing;

and S3, putting the uniformly mixed feed raw materials obtained in the step S2 into an automatic granulator to prepare feed particles with the particle size of 2mm, and naturally drying to obtain the portunus trituberculatus feed.

Example 3

The composite material consists of the following raw materials in percentage by mass: 9.34% of low gossypol cottonseed protein concentrate, 20% of soybean meal, 22.11% of flour, 0.87% of arginine, 0.14% of lysine, 0.08% of methionine, 2% of monocalcium phosphate, 0.3% of choline chloride, 1% of vitamin premix, 2% of mineral premix, 2.79% of fish oil, 2% of soybean lecithin, 2% of sodium alginate, 3.37% of carboxymethyl cellulose and the balance of fish meal; the vitamin premix comprises the following components: 0.12% retinyl acetate, 0.12% cholecalciferol, 2% all-rac-alpha-tocopherol, 0.9% menadione, 1.1% thiamine, 0.75% riboflavin, 3% ascorbic acid, 1.2% pyridoxine hydrochloride, 0.2% cyanocobalamin, 4% folic acid, 1.25% biotin, 4.04% nicotinic acid, 1.6% d-calcium pantothenate, 20.4% inositol, balance carboxymethylcellulose; the mineral premix comprises the following components: 0.457 percent of ferric citrate, 0.943 percent of zinc sulfate heptahydrate, 0.414 percent of manganese sulfate monohydrate, 0.661 percent of copper sulfate pentahydrate, 23.9 percent of manganese sulfate heptahydrate, 23.3 percent of monopotassium phosphate, 13.7 percent of sodium dihydrogen phosphate, 3.4 percent of calcium lactate pentahydrate, 0.136 percent of cobalt chloride hexahydrate and the balance of zeolite powder.

The preparation method of the portunus trituberculatus feed comprises the following steps: s1, dissolving soybean lecithin in fish oil in a formula ratio in advance, and uniformly mixing;

and S3, putting the uniformly mixed feed raw materials obtained in the step S2 into an automatic granulator to prepare feed particles, wherein one half of the feed particles is 2mm, and the other half of the feed particles is 4mm, and naturally drying the feed particles to obtain the portunus trituberculatus feed.

Example 4

The composite material consists of the following raw materials in percentage by mass: 9.8% of low gossypol cottonseed protein concentrate, 20% of soybean meal, 24% of flour, 0.8% of arginine, 0.18% of lysine, 0.08% of methionine, 2.5% of monocalcium phosphate, 0.35% of choline chloride, 1.1% of vitamin premix, 2.4% of mineral premix, 3.3% of fish oil, 2.4% of soybean lecithin, 2.2% of sodium alginate, 3.8% of carboxymethyl cellulose and the balance of fish meal; the vitamin premix comprises the following components: 0.18% retinyl acetate, 0.18% cholecalciferol, 2.5% all-rac-alpha-tocopherol, 0.8% menadione, 1.8% thiamine, 0.78% riboflavin, 3.5% ascorbic acid, 1.4% pyridoxine hydrochloride, 0.25% cyanocobalamin, 4.5% folic acid, 1.28% biotin, 4.2% nicotinic acid, 1.7% d-calcium pantothenate, 20.8% inositol, balance carboxymethylcellulose, wherein the mineral premix consists of: 0.48 percent of ferric citrate, 0.98 percent of zinc sulfate heptahydrate, 0.48 percent of manganese sulfate monohydrate, 0.68 percent of copper sulfate pentahydrate, 23.8 percent of manganese sulfate heptahydrate, 23.8 percent of potassium dihydrogen phosphate, 13.8 percent of sodium dihydrogen phosphate, 3.8 percent of calcium lactate pentahydrate, 0.18 percent of cobalt chloride hexahydrate and the balance of zeolite powder.

In the embodiment, the nutritional ingredients in the portunus trituberculatus feed comprise the following components in percentage by mass: 47% of crude protein, 8.8% of crude fat, 14.8% of carbohydrate, 10% of crude fiber, 10-11% of ash, 9% of water and 0.0065% of free gossypol, and when the feed is designed, the requirements of each breeding animal on the protein fat are not the same as each other.

Example 5

The embodiment provides a portunus trituberculatus which comprises the following raw materials in percentage by mass: 10% of low gossypol cottonseed protein concentrate, 22% of soybean meal, 25% of flour, 1% of arginine, 0.2% of lysine, 0.1% of methionine, 3% of monocalcium phosphate, 0.4% of choline chloride, 1.2% of vitamin premix, 2.5% of mineral premix, 3.5% of fish oil, 2.5% of soybean lecithin, 2.5% of sodium alginate, 4% of carboxymethyl cellulose and the balance of fish meal, wherein the vitamin premix comprises the following components: 0.2% retinyl acetate, 0.2% cholecalciferol, 3% all-rac-alpha-tocopherol, 1% menadione, 2% thiamine, 0.8% riboflavin, 4% ascorbic acid, 1.5% pyridoxine hydrochloride, 0.3% cyanocobalamin, 5% folic acid, 1.3% biotin, 4.5% nicotinic acid, 1.8% d-calcium pantothenate, 21% inositol, the balance carboxymethylcellulose, the mineral premix consisting of: 0.5 percent of ferric citrate, 1.0 percent of zinc sulfate heptahydrate, 0.5 percent of manganese sulfate monohydrate, 0.7 percent of copper sulfate pentahydrate, 24 percent of manganese sulfate heptahydrate, 24 percent of monopotassium phosphate, 14 percent of sodium dihydrogen phosphate, 4 percent of calcium lactate pentahydrate, 0.2 percent of cobalt chloride hexahydrate and the balance of zeolite powder.

s2, softening and flaking the hulled cottonseed obtained in the step S1 to obtain a blank piece with the thickness of less than 5mm, and then ventilating at the ambient temperature of 55-60 ℃;

s3, preparing a mixed solvent prepared from 90% methanol and 97% n-hexane in a volume ratio of 1:1, feeding the embryo piece obtained in the step S2 into a continuous sprayer, spraying the mixed solvent for 40-45min to enable the surface of the embryo piece to be soaked in the mixed solvent, then placing the embryo piece into the mixed solvent again to soak for 20-30min to complete the dephenolization process, removing the mixed solvent after soaking is finished, and then extracting at 60 ℃ and drying at 85 ℃ to obtain the low-gossypol cottonseed protein concentrate.

In the embodiment, the nutritional ingredients in the portunus trituberculatus feed comprise the following components in percentage by mass: 48% of crude protein, 9% of crude fat, 15% of carbohydrate, 12% of crude fiber, 11% of ash, 10% of water and 0.007% of free gossypol. When the feed is designed, the requirements of each breeding animal on protein fat are not the same as much, and the nutritional ingredients of the feed for the portunus trituberculatus are designed according to the invention, so that the nutritional ingredient requirements in the feeding of the portunus trituberculatus can be ensured.

and S3, putting the uniformly mixed feed raw materials obtained in the step S2 into an automatic granulator to prepare feed particles with the particle size of 4mm, and naturally drying the feed particles to obtain the portunus trituberculatus feed.

Example 6

The difference from the example 3 is that the raw material composition of the blue crab feed of the present embodiment does not contain the low gossypol cottonseed protein concentrate, and the rest is the same as the example 3, and the detailed description is omitted here.

Example 7

The difference from the example 3 is only that the content of the gossypol cottonseed protein concentrate in the raw material composition of the blue crab feed is 4.67%, and the rest is the same as the example 3, and the description is omitted here.

Example 8

The difference from the example 3 is that the content of the gossypol cottonseed protein concentrate in the raw material composition of the blue crab feed is 14.01%, and the rest is the same as the example 3, and the description is omitted.

Example 9

The difference from the example 3 is that the content of the gossypol cottonseed protein concentrate in the raw material composition of the blue crab feed is 18.68%, and the rest is the same as the example 3, and the description is omitted here.

Example 10

The difference from the example 3 is only that the content of the gossypol cottonseed protein concentrate in the raw material composition of the blue crab feed is 28.02%, and the rest is the same as the example 3, and the description is omitted here.

The composition of the nutritional components in the blue crab feed of example 3 and examples 6-10 is shown in table 1:

table 1: table of the composition of the nutritional ingredients in the blue crab feeds of example 3 and examples 6-10

Test part:

1. procedure of the test

The blue crabs were fed with the blue crabs fed with the feed prepared in example 3 and examples 6 to 10, and the contents of gossypol cottonseed protein concentrate in the raw materials were changed from low to high, as defined in example 6 as group 1, example 7 as group 2, example 3 as group 3, example 8 as group 4, example 9 as group 5, and example 10 as group 6. The feeding method comprises the following specific steps: selecting 180 blue crabs with the initial weight of 7.59 +/-0.06 g, randomly distributing the blue crabs into 180 plastic baskets (35 cm multiplied by 30cm multiplied by 35 cm) in a cement pond (6.8 m multiplied by 3.8m multiplied by 1.7 m), and feeding the blue crabs with the feed formulas in the groups 1 to 6. 1 feed component is divided into 3 parallel groups, and each parallel group contains 10 crabs. The breeding experiment lasts for 8 weeks, and the feeding is carried out once per day at 08 00 and 18. Counting the survival rate after the experiment is finished, measuring the weight of each crab, calculating the weight gain rate, the specific growth rate, the molting rate and the feed efficiency, measuring the key gene expression of the protein metabolism of the liver and the pancreas, and the key gene expression of Toll and IMD immune channels, manufacturing liver and pancreas tissue sections, and observing the health condition of the liver and the pancreas under an optical microscope.

2. Test results

The test results of the above tests are shown in table 2-table 3 and fig. 1-fig. 4, where table 2 is a table of the results of statistics of the survival rate, the end weight and the weight gain rate of portunus trituberculatus after the portunus trituberculatus feed prepared by groups 1-6 is fed, and table 3 is a table of the results of statistics of the specific growth rate, the molting rate and the feed efficiency of portunus trituberculatus after the portunus trituberculatus feed prepared by groups 1-6 is fed.

Table 2: results table of the influence of different low gossypol cottonseed protein concentrate contents on the survival rate, the final weight and the weight gain rate of the portunus trituberculatus

Table 3: results table of the influence of different low gossypol cottonseed concentrated protein contents on specific growth rate, molting rate and feed efficiency of portunus trituberculatus

Group of	Specific growth Rate (%)	Molt rate (%)	Feed efficiency (%)
				Group 1	4.04±0.05 ^ab	2.72±0.13 ^ab	0.45±0.02 ^b
Group 2	4.08±0.00 ^ab	2.88±0.08 ^b	0.46±0.01 ^b
				Group 3	4.19±0.01 ^b	2.93±0.07 ^b	0.47±0.02 ^b
Group 4	4.09±0.07 ^ab	2.93±0.07 ^b	0.46±0.02 ^b
				Group 5	4.07±0.03 ^ab	2.70±0.13 ^ab	0.40±0.03 ^a
Group 6	3.90±0.07 ^a	2.42±0.05 ^a	0.37±0.04 ^a

The different letters in the same column in tables 2 and 3 represent significant differences (P < 0.05)

Tables 2 and 3 show that the survival rates of portunus trituberculatus, terminal weight, weight gain, specific growth rate, shelling rate and feed efficiency of groups 1-6 were significantly higher than those of group 6 (P < 0.05) and that of groups 3 and 4 reached 100%, terminal weight and weight gain of group 3 were significantly higher than those of group 6 (P < 0.05), specific growth rate of group 3 was significantly higher than that of group 6 (P < 0.05), shelling rate of group 2-4 was significantly higher than that of group 6 (P < 0.05), and feed efficiency of group 1-4 was significantly higher than those of groups 5 and 6 (P < 0.05).

Fig. 1 and fig. 2 show the influence of feeds with different low gossypol cottonseed protein concentrates on the expression of key genes of the hepatopancreas protein anabolism of the portunus trituberculatus, and the results show that the hepatopancreas of groups 4 to 6, s6k1, s6 and eif4e3 genes have significantly higher expression level than that of groups 1 to 3. The hepatopancreatic akt gene expression level of group 4 was significantly higher than that of group 6. The hepatopancreatic 4ebp1 gene expression level of the groups 3 to 6 is obviously higher than that of the groups 1 and 2. The hepatopancreatic eif4e1a gene expression level of group 4 and group 5 is significantly higher than that of group 1 to group 3. The hepatopancreatic eif4e2 gene expression levels of group 4 and group 5 were significantly higher than those of groups 1 to 3 and 6.

FIG. 3 shows the effect of different feeds with low gossypol cottonseed protein concentrate content on the expression of key genes of hepatopancreas Toll and IMD (in-mould decoration) pathways of Portunus trituberculatus. The results showed that the expression level of the hepatopancreatic tlr4 gene of group 5 and group 6 was significantly higher than that of group 1. The hepatopancreatic irak4 gene expression level of group 5 was significantly higher than that of groups 1 to 4. The hepatopancreata myd gene expression level of groups 4 to 6 is significantly higher than that of group 1. The hepatopancreatic traf6 gene expression levels of groups 4 to 6 were significantly higher than those of groups 1 to 3. The hepatopancreatoresh gene expression levels of groups 4 to 5 are significantly higher than those of groups 1 to 3.

Fig. 4-5 show the effect of different feeds with low gossypol cottonseed protein concentrate content on the liver pancreas histomorphology of blue crab. The results show that the vacuolar diameter of the hepatic tubules significantly increased and the lipid droplet content gradually decreased in the group 1 to group 3 treatment groups.

The results show that the feed formula of the invention has balanced nutrition, reduces the using amount of fish meal by partially replacing fish meal with the low gossypol cottonseed concentrated protein, can reduce the feeding cost of the portunus trituberculatus, and has promotion effects on the growth of the portunus trituberculatus, the protein anabolism and the expression of key genes of Toll and IMD immune pathways.

Although the present disclosure has been described above, the scope of the present disclosure is not limited thereto. Those skilled in the art can make various changes and modifications without departing from the spirit and scope of the present disclosure, and such changes and modifications will fall within the scope of the present invention.

Claims

1. The feed for the portunus trituberculatus is characterized by comprising 9 to 10 percent of low gossypol cottonseed protein concentrate by mass percent.

2. The portunus trituberculatus feed as claimed in claim 1, which is composed of the following raw materials by mass percentage: 9-10% of low gossypol cottonseed protein concentrate, 17-22% of soybean meal, 20-25% of flour, 0.5-1% of arginine, 0.1-0.2% of lysine, 0.05-0.1% of methionine, 1-3% of calcium dihydrogen phosphate, 0.2-0.4% of choline chloride, 0.8-1.2% of vitamin premix, 1.5-2.5% of mineral premix, 2.5-3.5% of fish oil, 1.5-2.5% of soybean phospholipid, 1.5-2.5% of sodium alginate, 3-4% of carboxymethyl cellulose and the balance of fish meal.

3. The portunus trituberculatus feed as claimed in claim 2, which is composed of the following raw materials by mass percent: 9.34 percent of low gossypol cottonseed protein concentrate, 20 percent of soybean meal, 22.11 percent of flour, 0.87 percent of arginine, 0.14 percent of lysine, 0.08 percent of methionine, 2 percent of monocalcium phosphate, 0.3 percent of choline chloride, 1 percent of vitamin premix, 2 percent of mineral premix, 2.79 percent of fish oil, 2 percent of soybean lecithin, 2 percent of sodium alginate, 3.37 percent of carboxymethyl cellulose and the balance of fish meal.

4. The blue crab feed according to claim 2, wherein the vitamin premix comprises the following components by mass percent: 0.1-0.2% of retinyl acetate, 0.1-0.2% of cholecalciferol, 1-3% of all-rac-alpha-tocopherol, 0.5-1% of menadione, 1-2% of thiamine, 0.7-0.8% of riboflavin, 2-4% of ascorbic acid, 1-1.5% of pyridoxine hydrochloride, 0.1-0.3% of cyanocobalamin, 3-5% of folic acid, 1.2-1.3% of biotin, 3.5-4.5% of nicotinic acid, 1.5-1.8% of D-calcium pantothenate, 20-21% of inositol and the balance of carboxymethyl cellulose.

5. The feed for blue crabs according to claim 4, wherein the vitamin premix comprises the following components by mass percent: 0.12% retinyl acetate, 0.12% cholecalciferol, 2% all-rac-alpha-tocopherol, 0.9% menadione, 1.1% thiamine, 0.75% riboflavin, 3% ascorbic acid, 1.2% pyridoxine hydrochloride, 0.2% cyanocobalamin, 4% folic acid, 1.25% biotin, 4.04% nicotinic acid, 1.6% D-calcium pantothenate, 20.4% inositol, the balance carboxymethyl cellulose.

6. The portunus trituberculatus feed as claimed in claim 2, wherein the mineral premix comprises the following components by mass percent: 0.4-0.5% of ferric citrate, 0.9-1.0% of zinc sulfate heptahydrate, 0.4-0.5% of manganese sulfate monohydrate, 0.6-0.7% of copper sulfate pentahydrate, 23-24% of manganese sulfate heptahydrate, 23-24% of potassium dihydrogen phosphate, 12-14% of sodium dihydrogen phosphate, 3-4% of calcium lactate pentahydrate, 0.1-0.2% of cobalt chloride hexahydrate and the balance of zeolite powder.

7. The feed for blue crabs according to claim 6, wherein the mineral premix comprises the following components by mass percent: 0.457 percent of ferric citrate, 0.943 percent of zinc sulfate heptahydrate, 0.414 percent of manganese sulfate monohydrate, 0.661 percent of copper sulfate pentahydrate, 23.9 percent of manganese sulfate heptahydrate, 23.3 percent of potassium dihydrogen phosphate, 13.7 percent of sodium dihydrogen phosphate, 3.4 percent of calcium lactate pentahydrate, 0.136 percent of cobalt chloride hexahydrate and the balance of zeolite powder.

8. The portunus trituberculatus feed as claimed in any of claims 1-6, wherein the extraction method of the low gossypol cottonseed protein concentrate comprises the following steps:

and S3, preparing a mixed solvent, namely firstly sending the embryo piece obtained in the step S2 into a continuous sprayer, spraying the mixed solvent for 40-45min to soak the surface of the embryo piece with the mixed solvent, then putting the embryo piece into the mixed solvent again to soak for 20-30min to complete the dephenolization process, removing the mixed solvent after soaking is finished, and then extracting and drying to obtain the low gossypol cottonseed concentrated protein.

9. The portunus trituberculatus feed as claimed in any of claims 1-6, wherein the nutritional ingredients in the portunus trituberculatus feed comprise the following components in percentage by mass: 45-48% of crude protein, 8-9% of crude fat, 14-15% of carbohydrate, 10-12% of crude fiber, 10-11% of ash, 9-10% of water and 0.0065-0.007% of free gossypol.

10. The preparation method of the portunus trituberculatus feed as claimed in any one of claims 1 to 6, which comprises the following steps: