A kind of Litopenaeus vannamei feed and preparation method thereof

Abstract

The invention discloses a Penaeus vannamei feed and a preparation method thereof, and belongs to the technical field of aquatic animal feed. The Penaeus vannamei feed comprises the following raw materials: basic feed, seabuckthorn flavonoids and cassia seed polysaccharide. The invention also discloses a preparation method of the Penaeus vannamei feed. The Penaeus vannamei feed prepared by the invention can improve the metabolic capacity, antioxidant capacity and immune capacity of Penaeus vannamei, reduce its oxidative stress, improve its digestive enzyme activity, effectively control and prevent the outbreak of shrimp diseases, and has important practical significance for the development of green and environmentally friendly shrimp farming industry.

Description

Technical Field

[0001] The invention belongs to the technical field of aquatic animal feed, and in particular relates to a Litopenaeus vannamei feed and a preparation method thereof. Background Art

[0002] Litopenaeus vanmamei, also known as whiteleg shrimp, is the largest species among the three major farmed shrimps in the world and is also the main farmed shrimp species in my country. In order to ensure the supply of shrimp, intensive farming is often used in the shrimp farming process. Intensive farming can further increase the farming density and ensure that the supply of shrimp meets market demand.

[0003] However, due to the high stocking density of this farming method, the water environment is easily deteriorated, and the content of ammonia, nitrogen, salt and toxic substances exceeds the standard for a long time. In addition, the protein and fat content in some shrimp feeds during the breeding process is high. Long-term feeding will cause fat to deposit in the liver, causing the shrimp hepatopancreas to be in an overloaded state for a long time, causing liver cell damage and ultimately leading to the death of the shrimp.

[0004] At the same time, the intensive farming model has increased the outbreak rate of shrimp diseases, especially hepatitis and white spot syndrome. Some farmers often use antibiotics and other drugs to deal with such diseases, but these drugs will remain in the water, further damaging the water quality and increasing the detoxification and detoxification pressure of the shrimp hepatopancreas, and cannot play a good preventive and therapeutic role. At the same time, the occurrence of shrimp diseases is related to many factors such as pathogens, shrimp physique and environmental factors, and it is difficult to provide comprehensive protection with a single method. This series of problems has brought economic losses to the shrimp farming industry and seriously affected the development of the shrimp farming industry. How to use green and safe methods to improve the immunity and disease resistance of shrimp and fundamentally reduce the use of external drugs is one of the problems that need to be solved in the current shrimp farming process. Summary of the invention

[0005] The purpose of the present invention is to provide a feed for Penaeus vannamei and a preparation method thereof, so as to improve the immunity, antioxidant capacity and disease resistance of Penaeus vannamei and effectively control and prevent the outbreak of shrimp diseases during intensive breeding.

[0006] The technical solution adopted to achieve the above purpose is to provide a vannamei shrimp feed, comprising the following raw materials in parts by weight: 99.84-99.98 parts of basic feed, 0.01-0.08 parts of seabuckthorn flavonoids and 0.01-0.08 parts of cassia seed polysaccharide.

[0007] The beneficial effects of the above technical scheme of the present invention are as follows: the feed of Penaeus vannamei of the present invention contains seabuckthorn flavonoids and cassia seed polysaccharides. Seabuckthorn flavonoids, as an antioxidant in the body, can effectively reduce the damage of oxidative stress to the body of Penaeus vannamei, and have a certain positive effect on the digestive function of the stomach and intestines, and can enhance the digestive ability of the shrimp by promoting the secretion of digestive enzymes or improving the intestinal absorption efficiency. Cassia seed polysaccharides have a significant regulatory effect, can enhance the immune function of the body, and at the same time have a certain protective effect on the hepatopancreas of the shrimp, and maintain its normal function by reducing the burden of the hepatopancreas or promoting the repair of hepatopancreas cells. Compared with the addition of seabuckthorn flavonoids or cassia seed polysaccharides alone, the combination of the two can produce a synergistic effect, significantly improve the immune enzyme activity of Penaeus vannamei, thereby improving immunity, synergistically expanding the protective effect and antioxidant capacity of the hepatopancreas, and enhancing its digestive ability. The mixed use of the two can amplify their respective biological activities and bring better overall effects. At the same time, the side effects caused by excessive dosage or single mechanism of action caused by the addition of seabuckthorn flavonoids or cassia seed polysaccharides alone can be avoided. The Penaeus vannamei feed of the present invention can effectively reduce the use of antibiotics in the Penaeus vannamei breeding process.

[0008] Preferably, the feed for Penaeus vannamei comprises the following raw materials in parts by weight: 99.85-99.95 parts of basic feed, 0.025-0.075 parts of seabuckthorn flavonoids and 0.025-0.075 parts of cassia seed polysaccharide.

[0009] More preferably, the basic feed includes the following raw materials in parts by weight: 15-20 parts of fish meal, 20-25 parts of soybean meal, 10-15 parts of peanut bran, 8-10 parts of chicken meal, 5-7 parts of shrimp meal, 20-22 parts of flour, 1-3 parts of fish oil, 2-3 parts of soybean oil, 1-2 parts of phosphate and calcium nutritional additives, 0.1-0.3 parts of shrimp vitamins, 0.3-0.7 parts of shrimp minerals, 0.2-0.4 parts of amino acids, 0.1-0.3 parts of amino acid synthesis promoter, 0.2-0.4 parts of salt, and 1-1.5 parts of adhesive.

[0010] More preferably, the basic feed includes the following raw materials in parts by weight: 18 parts of fish meal, 24.5 parts of soybean meal, 11 parts of peanut bran, 10 parts of chicken meal, 6 parts of shrimp meal, 21.65-21.75 parts of flour, 2 parts of fish oil, 2.5 parts of soybean oil, 1.5 parts of phosphorus and calcium nutritional additives, 0.2 parts of shrimp vitamins, 0.5 parts of shrimp minerals, 0.3 parts of amino acids, 0.2 parts of amino acid synthesis promoter, 0.3 parts of salt, and 1.2 parts of adhesive.

[0011] More preferably, the phosphate-calcium nutritional additive is calcium dihydrogen phosphate; the binder is sodium alginate; the amino acid is methionine; and the amino acid synthesis promoter is choline chloride.

[0012] More preferably, the vitamins per kilogram of shrimp include: vitamin A 4000000 IU, vitamin D3 2000000 IU, vitamin E 30g, vitamin K3 10g, vitamin B15g, vitamin B2 15g, vitamin B6 8g, vitamin B12 0.02g, niacin 40g, calcium pantothenate 25g, folic acid 2.5g, inositol 150g and biotin 0.08g.

[0013] More preferably, the minerals per kilogram of shrimp include: 12g magnesium sulfate monohydrate, 90g sodium chloride, 3g metallic copper, 1g ferrous sulfate monohydrate, 0.06g calcium iodate, 0.16g cobalt methionine, 10g zinc sulfate monohydrate and 0.0036g sodium selenite.

[0014] The present invention also provides a method for preparing a Litopenaeus vannamei feed, comprising the following steps:

[0015] The raw materials are crushed and sieved, mixed and then granulated, matured and dried in the shade to obtain the vannamei shrimp feed.

[0016] Preferably, the particle size of the granules obtained by granulation is 1.3 to 1.7 mm.

[0017] More preferably, the particle size of the granules obtained by granulation is 1.5 mm.

[0018] Preferably, the aging temperature is 55-65° C. and the time is 2.5-3.5 h.

[0019] More preferably, the aging temperature is 60°C and the time is 3 hours.

[0020] The present invention has the following beneficial effects:

[0021] (1) The shrimp feed prepared by the present invention is nutritionally comprehensive and can meet the shrimp's needs for essential fatty acids, proteins, amino acids and other nutrients.

[0022] (2) The vannamei shrimp feed of the present invention can improve the metabolic capacity, antioxidant capacity and immune capacity of vannamei shrimp, reduce its oxidative stress, and improve its digestive enzyme activity; it can effectively control and prevent the outbreak of shrimp diseases, and has important practical significance for the development of green and environmentally friendly shrimp farming industry. BRIEF DESCRIPTION OF THE DRAWINGS

[0023] Figure 1 Figure 1 is a graph of growth indexes of Penaeus vannamei; (a) is a graph of weight gain rate changes; (b) is a graph of specific growth rate changes;

[0024] Figure 2The diagrams are the serum enzyme activity diagrams of Penaeus vannamei; wherein, (a) is the activity diagram of alanine aminotransferase; (b) is the activity diagram of aspartate aminotransferase; (c) is the activity diagram of alkaline phosphatase; (d) is the activity diagram of acid phosphatase;

[0025] Figure 3 The diagrams are of oxidative stress in the hepatopancreas of Penaeus vannamei; (a) is a diagram of superoxide dismutase activity; (b) is a diagram of catalase activity; (c) is a diagram of active oxygen content; and (d) is a diagram of malondialdehyde content.

[0026] Figure 4 These are the digestive enzyme activity diagrams of the intestinal tract of Penaeus vannamei; among them, (a) is the protease activity diagram; (b) is the amylase activity diagram; (c) is the lipase activity diagram;

[0027] Figure 5 This is a graph showing the number of differentially expressed genes in Litopenaeus vannamei;

[0028] Figure 6 The top 20 GO terms with significant enrichment of differentially expressed genes in the S1 group fed with comparative example 2 are shown;

[0029] Figure 7 The top 20 GO terms with significant enrichment of differentially expressed genes in the J1 group fed with Comparative Example 4 are shown;

[0030] Figure 8 The top 20 GO terms with significant enrichment of differentially expressed genes in the SJ1 group fed in Example 1 are shown;

[0031] Fig. 9 The KEGG pathway diagram of the top 20 significantly enriched differentially expressed genes in the S1 group fed with comparative example 2;

[0032] Fig.10 The KEGG pathway diagram of the top 20 significantly enriched differentially expressed genes in the J1 group fed with comparative example 4;

[0033] Fig.11 The KEGG pathway diagram of the top 20 significantly enriched differentially expressed genes in the SJ1 group fed in Example 1;

[0034] Fig.12 This is a graph of the number of differential metabolites in Litopenaeus vannamei;

[0035] Fig.13 The top 20 KEGG pathway maps of significantly enriched differential metabolites in the S1 group fed with comparative example 2;

[0036] Fig.14 The top 20 KEGG pathway maps of significantly enriched differential metabolites in the J1 group fed with comparative example 4;

[0037] Fig.15The top 20 KEGG pathway maps of significantly enriched differential metabolites in the SJ1 group fed with Example 1 are shown. DETAILED DESCRIPTION

[0038] The features and properties of the present invention are further described in detail below in conjunction with the examples: Seabuckthorn flavonoids, with a content of 40%, were provided by Shaanxi Wanzhiyuan Biotechnology Co., Ltd.; Cassia seed polysaccharide, with a content of 80%, was provided by Peptide Love Peptide Manufacturer.

[0039] Example 1

[0040] A feed for Penaeus vannamei, comprising the following raw materials in parts by weight: 18 parts of fish meal, 24.5 parts of soybean meal, 11 parts of peanut bran, 10 parts of chicken meal, 6 parts of shrimp meal, 21.75 parts of flour, 2 parts of fish oil, 2.5 parts of soybean oil, 1.5 parts of calcium dihydrogen phosphate, 0.2 parts of shrimp vitamins, 0.5 parts of shrimp minerals, 0.3 parts of methionine, 0.2 parts of choline chloride, 0.3 parts of salt, 1.2 parts of sodium alginate, 0.025 parts of seabuckthorn flavonoids and 0.025 parts of cassia seed polysaccharide;

[0041] Among them, the vitamins per kilogram of shrimp include: vitamin A 4000000IU, vitamin D3 2000000IU, vitamin E 30g, vitamin K3 10g, vitamin B15g, vitamin B2 15g, vitamin B6 8g, vitamin B12 0.02g, niacin 40g, calcium pantothenate 25g, folic acid 2.5g, inositol 150g and biotin 0.08g;

[0042] The minerals in each kilogram of shrimp include: 12g magnesium sulfate monohydrate, 90g sodium chloride, 3g metallic copper, 1g ferrous sulfate monohydrate, 0.06g calcium iodate, 0.16g cobalt methionine, 10g zinc sulfate monohydrate and 0.0036g sodium selenite.

[0043] The preparation method of the vannamei shrimp feed in this embodiment comprises the following steps:

[0044] The above raw materials were crushed and passed through a 60-mesh sieve, mixed and poured into a twin-screw extruder to form particles with a particle size of 1.5 mm. The particles were placed in a 60°C oven for aging for 3 hours, then taken out, dried in the shade to regain moisture, and thus the vannamei shrimp feed was obtained.

[0045] Example 2

[0046] A feed for Penaeus vannamei, comprising the following raw materials in parts by weight: 18 parts of fish meal, 24.5 parts of soybean meal, 11 parts of peanut bran, 10 parts of chicken meal, 6 parts of shrimp meal, 21.65 parts of flour, 2 parts of fish oil, 2.5 parts of soybean oil, 1.5 parts of calcium dihydrogen phosphate, 0.2 parts of shrimp vitamins, 0.5 parts of shrimp minerals, 0.3 parts of methionine, 0.2 parts of choline chloride, 0.3 parts of salt, 1.2 parts of sodium alginate, 0.075 parts of seabuckthorn flavonoids and 0.075 parts of cassia seed polysaccharide;

[0047] Among them, the vitamins per kilogram of shrimp include: vitamin A 4000000IU, vitamin D3 2000000IU, vitamin E 30g, vitamin K3 10g, vitamin B15g, vitamin B2 15g, vitamin B6 8g, vitamin B12 0.02g, niacin 40g, calcium pantothenate 25g, folic acid 2.5g, inositol 150g and biotin 0.08g;

[0048] The minerals in each kilogram of shrimp include: 12g magnesium sulfate monohydrate, 90g sodium chloride, 3g metallic copper, 1g ferrous sulfate monohydrate, 0.06g calcium iodate, 0.16g cobalt methionine, 10g zinc sulfate monohydrate and 0.0036g sodium selenite.

[0049] The preparation method of the vannamei shrimp feed in this embodiment comprises the following steps:

[0050] The above raw materials were crushed and passed through a 60-mesh sieve, mixed and poured into a twin-screw extruder to form particles with a particle size of 1.5 mm. The particles were placed in a 60°C oven for aging for 3 hours, then taken out, dried in the shade to regain moisture, and thus the vannamei shrimp feed was obtained.

[0051] Example 3

[0052] A feed for Penaeus vannamei, comprising the following raw materials in parts by weight: 20 parts of fish meal, 25 parts of soybean meal, 15 parts of peanut bran, 8 parts of chicken meal, 5 parts of shrimp meal, 20.94 parts of flour, 1 part of fish oil, 2 parts of soybean oil, 1 part of calcium dihydrogen phosphate, 0.1 part of shrimp vitamins, 0.3 part of shrimp minerals, 0.2 part of methionine, 0.1 part of choline chloride, 0.2 part of salt, 1 part of sodium alginate, 0.08 part of seabuckthorn flavonoids and 0.08 part of cassia seed polysaccharide;

[0053] Among them, the vitamins per kilogram of shrimp include: vitamin A 4000000IU, vitamin D3 2000000IU, vitamin E 30g, vitamin K3 10g, vitamin B15g, vitamin B2 15g, vitamin B6 8g, vitamin B12 0.02g, niacin 40g, calcium pantothenate 25g, folic acid 2.5g, inositol 150g and biotin 0.08g;

[0054] The minerals in each kilogram of shrimp include: 12g magnesium sulfate monohydrate, 90g sodium chloride, 3g metallic copper, 1g ferrous sulfate monohydrate, 0.06g calcium iodate, 0.16g cobalt methionine, 10g zinc sulfate monohydrate and 0.0036g sodium selenite.

[0055] The preparation method of the vannamei shrimp feed in this embodiment comprises the following steps:

[0056] The above raw materials were crushed and passed through a 60-mesh sieve, mixed and poured into a twin-screw extruder to form particles with a particle size of 1.5 mm. The particles were placed in a 60°C oven for aging for 3 hours, then taken out, dried in the shade to regain moisture, and thus the vannamei shrimp feed was obtained.

[0057] Example 4

[0058] A feed for Penaeus vannamei, comprising the following raw materials in parts by weight: 15 parts of fish meal, 20 parts of soybean meal, 15 parts of peanut bran, 10 parts of chicken meal, 7 parts of shrimp meal, 21.38 parts of flour, 3 parts of fish oil, 3 parts of soybean oil, 2 parts of calcium dihydrogen phosphate, 0.3 parts of shrimp vitamins, 0.7 parts of shrimp minerals, 0.4 parts of methionine, 0.3 parts of choline chloride, 0.4 parts of salt, 1.5 parts of sodium alginate, 0.01 parts of seabuckthorn flavonoids and 0.01 parts of cassia seed polysaccharide;

[0059] Among them, the vitamins per kilogram of shrimp include: vitamin A 4000000IU, vitamin D3 2000000IU, vitamin E 30g, vitamin K3 10g, vitamin B15g, vitamin B2 15g, vitamin B6 8g, vitamin B12 0.02g, niacin 40g, calcium pantothenate 25g, folic acid 2.5g, inositol 150g and biotin 0.08g;

[0060] The minerals in each kilogram of shrimp include: 12g magnesium sulfate monohydrate, 90g sodium chloride, 3g metallic copper, 1g ferrous sulfate monohydrate, 0.06g calcium iodate, 0.16g cobalt methionine, 10g zinc sulfate monohydrate and 0.0036g sodium selenite.

[0061] The preparation method of the vannamei shrimp feed in this embodiment comprises the following steps:

[0062] The above raw materials were crushed and passed through a 60-mesh sieve, mixed and poured into a twin-screw extruder to form particles with a particle size of 1.5 mm. The particles were placed in a 60°C oven for aging for 3 hours, then taken out, dried in the shade to regain moisture, and thus the vannamei shrimp feed was obtained.

[0063] Comparative Example 1

[0064] A feed for Penaeus vannamei, comprising the following raw materials in parts by weight: 18 parts of fish meal, 24.5 parts of soybean meal, 11 parts of peanut bran, 10 parts of chicken meal, 6 parts of shrimp meal, 21.8 parts of flour, 2 parts of fish oil, 2.5 parts of soybean oil, 1.5 parts of calcium dihydrogen phosphate, 0.2 parts of shrimp vitamins, 0.5 parts of shrimp minerals, 0.3 parts of methionine, 0.2 parts of choline chloride, 0.3 parts of salt and 1.2 parts of sodium alginate;

[0065] Among them, the vitamins per kilogram of shrimp include: vitamin A 4000000IU, vitamin D3 2000000IU, vitamin E 30g, vitamin K3 10g, vitamin B15g, vitamin B2 15g, vitamin B6 8g, vitamin B12 0.02g, niacin 40g, calcium pantothenate 25g, folic acid 2.5g, inositol 150g and biotin 0.08g;

[0066] The minerals in each kilogram of shrimp include: 12g magnesium sulfate monohydrate, 90g sodium chloride, 3g metallic copper, 1g ferrous sulfate monohydrate, 0.06g calcium iodate, 0.16g cobalt methionine, 10g zinc sulfate monohydrate and 0.0036g sodium selenite.

[0067] The preparation method of the vannamei shrimp feed in this embodiment comprises the following steps:

[0068] The above raw materials were crushed and passed through a 60-mesh sieve, mixed and poured into a twin-screw extruder to form particles with a particle size of 1.5 mm. The particles were placed in a 60°C oven for aging for 3 hours, then taken out, dried in the shade to regain moisture, and thus the vannamei shrimp feed was obtained.

[0069] Comparative Example 2

[0070] A feed for Penaeus vannamei, comprising the following raw materials in parts by weight: 18 parts of fish meal, 24.5 parts of soybean meal, 11 parts of peanut bran, 10 parts of chicken meal, 6 parts of shrimp meal, 21.75 parts of flour, 2 parts of fish oil, 2.5 parts of soybean oil, 1.5 parts of calcium dihydrogen phosphate, 0.2 parts of shrimp vitamins, 0.5 parts of shrimp minerals, 0.3 parts of methionine, 0.2 parts of choline chloride, 0.3 parts of salt, 1.2 parts of sodium alginate and 0.05 parts of seabuckthorn flavonoids;

[0071] Among them, the vitamins per kilogram of shrimp include: vitamin A 4000000IU, vitamin D3 2000000IU, vitamin E 30g, vitamin K3 10g, vitamin B15g, vitamin B2 15g, vitamin B6 8g, vitamin B12 0.02g, niacin 40g, calcium pantothenate 25g, folic acid 2.5g, inositol 150g and biotin 0.08g;

[0072] The minerals in each kilogram of shrimp include: 12g magnesium sulfate monohydrate, 90g sodium chloride, 3g metallic copper, 1g ferrous sulfate monohydrate, 0.06g calcium iodate, 0.16g cobalt methionine, 10g zinc sulfate monohydrate and 0.0036g sodium selenite.

[0073] The preparation method of the vannamei shrimp feed in this embodiment comprises the following steps:

[0074] The above raw materials were crushed and passed through a 60-mesh sieve, mixed and poured into a twin-screw extruder to form particles with a particle size of 1.5 mm. The particles were placed in a 60°C oven for aging for 3 hours, then taken out, dried in the shade to regain moisture, and thus the vannamei shrimp feed was obtained.

[0075] Comparative Example 3

[0076] A feed for Penaeus vannamei, comprising the following raw materials in parts by weight: 18 parts of fish meal, 24.5 parts of soybean meal, 11 parts of peanut bran, 10 parts of chicken meal, 6 parts of shrimp meal, 21.65 parts of flour, 2 parts of fish oil, 2.5 parts of soybean oil, 1.5 parts of calcium dihydrogen phosphate, 0.2 parts of shrimp vitamins, 0.5 parts of shrimp minerals, 0.3 parts of methionine, 0.2 parts of choline chloride, 0.3 parts of salt, 1.2 parts of sodium alginate and 0.15 parts of seabuckthorn flavonoids;

[0077] Among them, the vitamins per kilogram of shrimp include: vitamin A 4000000IU, vitamin D3 2000000IU, vitamin E 30g, vitamin K3 10g, vitamin B15g, vitamin B2 15g, vitamin B6 8g, vitamin B12 0.02g, niacin 40g, calcium pantothenate 25g, folic acid 2.5g, inositol 150g and biotin 0.08g;

[0078] The minerals in each kilogram of shrimp include: 12g magnesium sulfate monohydrate, 90g sodium chloride, 3g metallic copper, 1g ferrous sulfate monohydrate, 0.06g calcium iodate, 0.16g cobalt methionine, 10g zinc sulfate monohydrate and 0.0036g sodium selenite.

[0079] The preparation method of the vannamei shrimp feed in this embodiment comprises the following steps:

[0080] The above raw materials were crushed and passed through a 60-mesh sieve, mixed and poured into a twin-screw extruder to form particles with a particle size of 1.5 mm. The particles were placed in a 60°C oven for aging for 3 hours, then taken out, dried in the shade to regain moisture, and thus the vannamei shrimp feed was obtained.

[0081] Comparative Example 4

[0082] A feed for Penaeus vannamei, comprising the following raw materials in parts by weight: 18 parts of fish meal, 24.5 parts of soybean meal, 11 parts of peanut bran, 10 parts of chicken meal, 6 parts of shrimp meal, 21.75 parts of flour, 2 parts of fish oil, 2.5 parts of soybean oil, 1.5 parts of calcium dihydrogen phosphate, 0.2 parts of shrimp vitamins, 0.5 parts of shrimp minerals, 0.3 parts of methionine, 0.2 parts of choline chloride, 0.3 parts of salt, 1.2 parts of sodium alginate and 0.05 parts of cassia seed polysaccharide;

[0083] Among them, the vitamins per kilogram of shrimp include: vitamin A 4000000IU, vitamin D3 2000000IU, vitamin E 30g, vitamin K3 10g, vitamin B15g, vitamin B2 15g, vitamin B6 8g, vitamin B12 0.02g, niacin 40g, calcium pantothenate 25g, folic acid 2.5g, inositol 150g and biotin 0.08g;

[0084] The minerals in each kilogram of shrimp include: 12g magnesium sulfate monohydrate, 90g sodium chloride, 3g metallic copper, 1g ferrous sulfate monohydrate, 0.06g calcium iodate, 0.16g cobalt methionine, 10g zinc sulfate monohydrate and 0.0036g sodium selenite.

[0085] The preparation method of the vannamei shrimp feed in this embodiment comprises the following steps:

[0086] The above raw materials were crushed and passed through a 60-mesh sieve, mixed and poured into a twin-screw extruder to form particles with a particle size of 1.5 mm. The particles were placed in a 60°C oven for aging for 3 hours, then taken out, dried in the shade to regain moisture, and thus the vannamei shrimp feed was obtained.

[0087] Comparative Example 5

[0088] A feed for Penaeus vannamei, comprising the following raw materials in parts by weight: 18 parts of fish meal, 24.5 parts of soybean meal, 11 parts of peanut bran, 10 parts of chicken meal, 6 parts of shrimp meal, 21.65 parts of flour, 2 parts of fish oil, 2.5 parts of soybean oil, 1.5 parts of calcium dihydrogen phosphate, 0.2 parts of shrimp vitamins, 0.5 parts of shrimp minerals, 0.3 parts of methionine, 0.2 parts of choline chloride, 0.3 parts of salt, 1.2 parts of sodium alginate and 0.15 parts of cassia seed polysaccharide;

[0089] Among them, the vitamins per kilogram of shrimp include: vitamin A 4000000IU, vitamin D3 2000000IU, vitamin E 30g, vitamin K3 10g, vitamin B15g, vitamin B2 15g, vitamin B6 8g, vitamin B12 0.02g, niacin 40g, calcium pantothenate 25g, folic acid 2.5g, inositol 150g and biotin 0.08g;

[0090] The minerals in each kilogram of shrimp include: 12g magnesium sulfate monohydrate, 90g sodium chloride, 3g metallic copper, 1g ferrous sulfate monohydrate, 0.06g calcium iodate, 0.16g cobalt methionine, 10g zinc sulfate monohydrate and 0.0036g sodium selenite.

[0091] The preparation method of the vannamei shrimp feed in this embodiment comprises the following steps:

[0092] The above raw materials were crushed and passed through a 60-mesh sieve, mixed and poured into a twin-screw extruder to form particles with a particle size of 1.5 mm. The particles were placed in a 60°C oven for aging for 3 hours, then taken out, dried in the shade to regain moisture, and thus the vannamei shrimp feed was obtained.

[0093] Experimental example

[0094] 1. Nutritional level of Penaeus vannamei feed

[0095] The ingredients of the Penaeus vannamei feeds prepared in Examples 1 to 2 and Comparative Examples 1 to 5 were tested, and the test results are shown in Table 1.

[0096] Table 1 Nutritional level of Penaeus vannamei feed (dry matter basis %)

[0097]

[0098] 2. Determination of growth indexes of Penaeus vannamei

[0099] 2.1 Farming of Penaeus vannamei

[0100] Vannamei shrimp were purchased from Panyu District, Guangzhou City, Guangdong Province. After 7 days of temporary cultivation, healthy and uniform shrimps (1.2g±0.15g) were selected and randomly divided into 7 groups, each with 40 shrimps, 3 replicates per group, and cultured in water with a water temperature of 20-30°C, a salinity of 5‰, a pH of 7.8-8.0, and a dissolved oxygen content of 5-6 mg / L. Vannamei shrimp feeds of Examples 1-2 and Comparative Examples 1-5 were fed at 8:00, 14:00, and 20:00 every day, and the feed feeding situation was recorded after the last meal. The sewage was discharged once in the morning and evening every day, and the oxygen supply machine was kept to continuously supply oxygen, and the culture was carried out for 8 weeks.

[0101] The shrimps fed with the vannamei feed of comparative example 1 were recorded as group C; the shrimps fed with the vannamei feed of comparative example 2 were recorded as group S1; the shrimps fed with the vannamei feed of comparative example 3 were recorded as group S2; the shrimps fed with the vannamei feed of comparative example 4 were recorded as group J1; the shrimps fed with the vannamei feed of comparative example 5 were recorded as group J2; the shrimps fed with the vannamei feed of example 1 were recorded as group SJ1; and the shrimps fed with the vannamei feed of example 2 were recorded as group SJ2.

[0102] 2.2 Determination of growth indexes of Penaeus vannamei

[0103] The shrimps after 7 days of temporary culture were recorded as the initial shrimps, and the shrimps after 8 weeks of culture were recorded as the final shrimps. The quality of the initial shrimps and the final shrimps were recorded. The growth index was calculated according to the following formula (1) and formula (2):

[0104] Specific growth rate (SGR) = 100% × (lnW t -lnW 0 ) / t (1)

[0105] Weight gain rate (WGR) = 100% × [(W t -W 0 ) / W 0 ] (2)

[0106] In formula (1) and formula (2), W t and W 0 are the final shrimp mass and initial shrimp mass, in g; t is the number of days of culture experiment, in d. The results of growth index of Penaeus vannamei are shown in Figure 1 As shown in the figure, different small letters represent significant differences among the groups (p<0.05).

[0107] from Figure 1 It can be seen that the weight gain rate and specific growth rate of shrimp in group C, group S1, group S2, group J1, group J2, group SJ1 and group SJ2 have an upward trend, but only the weight gain rate and specific growth rate of group J1 and group SJ1 are significantly increased compared with group C (control) (p<0.05), and the weight gain rate and specific growth rate of group SJ1 are larger than those of group J1. The results show that Cassia seed polysaccharides and seabuckthorn flavonoids can synergistically improve the growth performance of Penaeus vannamei.

[0108] 3. Determination of serum, hepatopancreas and intestinal enzyme activities of Penaeus vannamei

[0109] 3.1 Processing of Penaeus vannamei samples

[0110] The culture of Penaeus vannamei was the same as that in 2.1. After 8 weeks of culture, serum, hepatopancreas and intestines were collected from Penaeus vannamei for later use. The specific operation was as follows: blood was collected from Penaeus vannamei with a 1.5 mL sterile syringe and centrifuged at 4 °C to obtain serum, which was then placed in a -80 °C refrigerator for later use. Twelve shrimps were collected for each replicate. The carapace of the shrimp was cut open with sterilized scissors, the hepatopancreas was taken out with tweezers and placed in a 2 mL enzyme-free tube, which was quickly placed in liquid nitrogen and transferred to a -80 °C refrigerator for later use. Three shrimps were collected for each replicate. The back of the shrimp was cut open with scissors, the muscles were peeled off, the intestines were picked out with tweezers, the intestines were cut off at both ends with scissors to remove the intestines, the intestines were placed in a 2 mL enzyme-free tube, which was quickly placed in liquid nitrogen and transferred to a -80 °C refrigerator for later use. Three shrimps were collected for each replicate.

[0111] 3.2 Serum, hepatopancreas, and intestinal enzyme activity assays

[0112] Serum stored at -80℃ was used to test the activities of acid phosphatase (ACP), alkaline phosphatase (ALP), aspartate aminotransferase (AST), and alanine aminotransferase (ALT); hepatopancreas samples stored at -80℃ were used to test the activities of reactive oxygen species (ROS), malondialdehyde (MDA), superoxide dismutase (SOD), and catalase (CAT); intestinal samples stored at -80℃ were used to test the activities of protease, amylase, and lipase. The specific steps were carried out according to the instructions of the kit, and the results were as follows: Figures 2 to 4 shown.

[0113] from Figure 2 It can be seen that, except for the J1 group, the activities of ALP and ACP in other experimental groups were significantly higher than those in the control group (p<0.05), indicating that seabuckthorn flavonoids and cassia seed polysaccharides can significantly improve the immune enzyme activity of Penaeus vannamei, thereby improving immunity; the activity of ALT in the experimental groups was significantly lower than that in the control group (p<0.05); except for the J2 group, the ACT activity of the other experimental groups was significantly lower than that of the C group, and the effects of the SJ1 and SJ2 groups in the experimental groups were better, indicating that seabuckthorn flavonoids and cassia seed polysaccharides have a significant effect on the protection of the hepatopancreas of shrimp, and the mixture will produce a certain synergistic effect, with better effects;

[0114] from Figure 3 It can be seen that the content of ROS in the experimental groups was significantly lower than that in the control group (p<0.05). Except for the S2 and J1 groups, the MDA content in other experimental groups was significantly lower than that in the control group (p<0.05), indicating that the addition of seabuckthorn flavonoids and cassia seed polysaccharides can significantly reduce the oxidative damage of Penaeus vannamei; except for the S1 and J2 groups, the activity of SOD in the experimental groups was significantly higher than that in the control group (p<0.05); the activity of CAT in all experimental groups was significantly higher than that in the control group (p<0.05), and the effect of SJ1 and SJ2 groups was better. It shows that feeding feeds supplemented with seabuckthorn flavonoids can significantly improve the antioxidant capacity of Penaeus vannamei;

[0115] from Figure 4 It can be seen that the protease activity of the experimental groups was significantly higher than that of the control group (p<0.05), and the activity of the SJ1 and SJ2 groups was higher. The amylase activity of all experimental groups was significantly higher than that of the control group (p<0.05), and the amylase activity of the SJ2 group was significantly higher than that of the other groups (p<0.05). Except for S1 and J2, the lipase activity of the other experimental groups was significantly higher than that of the control group (p<0.05), and the lipase activity of the two mixed groups of SJ1 and SJ2 was significantly higher than that of all other groups (p<0.05), indicating that seabuckthorn flavonoids and cassia seed polysaccharides can significantly improve the digestive enzyme activity of Penaeus vannamei, thereby enhancing its digestive ability, and the effect of mixed use is better.

[0116] 4. Transcriptomic Analysis

[0117] Perform transcriptomic analysis as follows:

[0118] (1) Total RNA was extracted from the hepatopancreas of shrimp in groups S1, C, J1, and SJ1 using the Trizol method. The RNA concentration was measured using a NanoDrop2000 spectrophotometer. The integrity of the RNA and the RNA integrity number (RIN) were then evaluated using 1% agarose gel electrophoresis and the Agilent2100 system. RNA with qualified quality was used to construct a transcriptome library and perform quality control. After completing the required identification and quantification steps, the library was sequenced on the Illumina HiSeq platform to generate pre-reads.

[0119] (2) Pre-reads were filtered by removing street sequences and low-quality sequences (Sanger base quality <20) and reads with unknown nucleotides greater than 10%, processed into clean-reads and compared with the reference genome for annotation; then the annotated genes were differentially analyzed to identify differentially expressed genes (DEGs), and the identification criteria were |log2fold change|≥1 and P-value<0.05;

[0120] (3) GO and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analysis was performed in the Gene Ontology (GO) database and the Kyoto Encyclopedia of Genes and Genomes (KEGG) database, with the standard of P-value < 0.05.

[0121] The results are as follows Figures 5 to 11 As shown, Figure 5As shown in the figure, a total of 863 (239 up-regulated, 624 down-regulated), 591 (322 up-regulated, 269 down-regulated) and 863 (239 up-regulated, 624 down-regulated) DEGs were identified in S1vsC, J1vsC and SJ1vsC. Among them, genes such as methylglutaryl-CoA hydratase, mitochondrial isozymes, C-type lectin, cuticle protein, anti-lipopolysaccharide factor isozymes, DNA translocase and multifunctional methyltransferase were up-regulated in the S1 group; genes such as cytochrome C reductase complex, α-(1,6)-fucosyltransferase, heat shock protein 70, juvenile hormone esterase-like carboxylesterase and heme binding protein were significantly down-regulated. In the J1 group, genes such as protease, sodium / glucose cotransporter, cytochrome P450, chitin, hemocyte homeostasis-related protein, neuropeptidase and trypsin were up-regulated; genes such as glucosamine-6-phosphate fructose aminotransferase, oxytocin receptor type I, apolipoprotein D, arylsulfatase B and metallothionein were down-regulated. This shows that Cassia seed polysaccharide may improve the ability of shrimp to utilize protein. In the SJ1 group, genes such as mitochondrial trifunctional enzymes, cytochrome P450, GTPase, chitinase, polyriboprotein, hemocyanin, glutathione transferase and NADPH oxidase were up-regulated; genes such as arylsulfatase, glycine receptor subunit, amylase were down-regulated. The results show that seabuckthorn flavonoids and Cassia seed polysaccharides may enhance the activity of energy metabolism and immune-related enzymes in shrimp.

[0122] At the same time, GO and KEGG enrichment analysis was performed on the identified DEGs, and the results are shown in Figures 6 to 11 ; DEGs in the S1 group were significantly enriched in GO terms such as carbohydrate phosphatase activity (GO:0015297), secondary active transmembrane transporter activity (GO:0015291), phosphatase activity (GO:0016791), and active transmembrane transporter activity (GO:0022804). Figure 6 ) and valine, leucine and isoleucine degradation (ko:00280), valine, leucine and isoleucine biosynthesis (ko:00290), metabolic pathways (ko:01100), pancreatic juice secretion (ko:04972), riboflavin metabolism (ko:00740), unsaturated fatty acid biosynthesis (ko:01040) and other pathways (e.g. Fig. 9 ), indicating that seabuckthorn flavonoids may affect the functions of amino acid synthesis and metabolism, fatty acid metabolism, etc. in Litopenaeus natans. DEGs in the J1 group were significantly enriched in GO items such as glycerophosphocholine phosphodiesterase activity (GO:0047389), cellular phosphate ion homeostasis (GO:0030643), regulation of extracellular matrix component secretion (GO:0003330), 1-phosphate glucose metabolism (GO:0019255), and phosphate ion homeostasis (GO:0055062). Figure 7) and pathways such as glycerophospholipid metabolism (ko00564), lysosome (ko04142), glycosaminoglycan degradation (ko00531), arachidonic acid metabolism (ko00590), protein digestion and absorption (ko04974), and pancreatic juice secretion (ko04972) (e.g. Fig.10 ). It shows that Cassia seed polysaccharide may affect the lipid metabolism, protein digestion and absorption of Penaeus vannamei. The DEGs in the SJ1 group were significantly enriched in steroid hormone receptor activity (GO:0003707), polymerase II transcription factor activity (GO:0004879), secondary active transmembrane transporter activity (GO:0015291), cotransporter activity (GO:0015293), active transmembrane transporter activity (GO:0022804), sodium ion transporter activity (GO:0015370), transmembrane transport and other GO items (such as Figure 8 ) and pathways such as sucrose and starch metabolism (ko00500), alanine, aspartate and glutamate metabolism (ko00250), antigen processing and delivery (ko04612), pancreatic juice secretion (ko04972), unsaturated fatty acid synthesis (ko01040), mineral absorption (ko04978), and fatty acid elongation (ko00062) (e.g. Fig.11 The results showed that seabuckthorn flavonoids and cassia seed polysaccharides may affect the transmembrane transport function, energy metabolism and lipid metabolism of Litopenaeus vannamei.

[0123] 5. Metabolomics analysis

[0124] Perform metabolomics analysis as follows:

[0125] (1) Metabolites were extracted from the hepatopancreas of shrimps in groups S1, C, J1, and SJ1. The extracted metabolites were then separated by hydrophilic interaction liquid chromatography (HILIC) using a Vanquish HPLC system, and the primary and secondary spectra of the samples were collected using an ABTriple TOF 6600 mass spectrometer.

[0126] (2) The collected data were tested for integrity, metabolites with more than 50% missing values ​​within the group were removed and excluded from subsequent analysis, extreme values ​​were deleted, and the data were normalized for total peak area to ensure parallel comparison between samples and metabolites;

[0127] (3) Metabolites were differentially analyzed and differential metabolites (DMs) were identified with VIP>1 and P-value<0.05 (Student's t test). KEGG pathway enrichment analysis was then performed on these differential metabolites with P-value<0.05.

[0128] The results are as follows Figures 12 to 15 As shown, from Fig.12 It can be seen that 19 (2 up-regulated, 17 down-regulated), 7 (6 up-regulated, 1 down-regulated) and 14 (7 up-regulated, 7 down-regulated) DMs were identified in S1vsC, J1vsC and SJ1vsC. Among them, 18-carboxyleukotrienes and vanillylmandelic acid were up-regulated in the S1 group, and metabolites such as L-hydroxyarginine and deoxyguanosine were down-regulated. Metabolites such as prostaglandins and phosphatidylcholine were up-regulated in the J1 group, and leonurine was down-regulated. Metabolites such as 7-hydroxyflavone, oxidized glutathione and acetylcholine were up-regulated in the SJ1 group, and metabolites such as DL-leucine, dimethylbenzyl imidazole and linolenic acid were down-regulated. KEGG enrichment analysis of these DMs showed that biotin metabolism (ko00780), thyroid hormone synthesis (ko00981), α-linolenic acid metabolism (ko00592), glutathione metabolism (ko00480), amino sugar and nucleotide sugar metabolism (ko00520), metabolic pathway (ko01100) and other pathways were significantly enriched in the S1 group ( Fig.13 ), indicating that seabuckthorn flavonoids have an effect on the energy metabolism and lipid metabolism of Penaeus vannamei. DMs in the J1 group were significantly enriched in phosphoinositide metabolism and metabolic pathways ( Fig.14 ), indicating that Cassia seed polysaccharide may have an effect on the metabolism of Penaeus vannamei. DMs in the SJ1 group were significantly enriched in cholesterol metabolism (ko04979), riboflavin metabolism (ko00740), thyroid hormone synthesis (ko04918), α-linolenic acid metabolism (ko00592), unsaturated fatty acid biosynthesis (ko01040), glutathione metabolism (ko00480) and other pathways ( Fig.15 ). The results showed that seabuckthorn flavonoids and cassia seed polysaccharides may affect the hormone metabolism and fatty acid metabolism of Penaeus vannamei.

[0129] In summary, feeding the vannamei shrimp feed prepared in the present application can increase the activity of ALP and ACP and digestive enzyme activity of vannamei shrimp, reduce the content of cholesterol in serum and the activity of ACT and ALT, and increase the activity of antioxidant enzymes to reduce the content of ROS and MDA; the results of the transcriptome and metabolome show that seabuckthorn flavonoids and cassia seed polysaccharides have a beneficial effect on the lipid metabolism and hormone metabolism of vannamei shrimp. The vannamei shrimp feed of the present invention can effectively promote the health of vannamei shrimp and reduce the use of antibiotics in shrimp farming.

[0130] The present invention has been described according to the above embodiments. It should be understood that the above embodiments do not limit the present invention in any form. Any technical solutions obtained by equivalent replacement or equivalent transformation shall fall within the scope of the present invention.

Claims

1. A feed for Penaeus vannamei, characterized in that: The raw materials include the following parts by weight: 99.85-99.95 parts of basic feed, 0.025-0.075 parts of seabuckthorn flavonoids and 0.025-0.075 parts of cassia seed polysaccharide; The basic feed comprises the following raw materials in parts by weight: 15-20 parts of fish meal, 20-25 parts of soybean meal, 10-15 parts of peanut bran, 8-10 parts of chicken meal, 5-7 parts of shrimp meal, 20-22 parts of flour, 1-3 parts of fish oil, 2-3 parts of soybean oil, 1-2 parts of phosphorus and calcium nutritional additives, 0.1-0.3 parts of shrimp vitamins, 0.3-0.7 parts of shrimp minerals, 0.2-0.4 parts of amino acids, 0.1-0.3 parts of amino acid synthesis promoter, 0.2-0.4 parts of salt, and 1-1.5 parts of adhesive.

2. The vannamei shrimp feed according to claim 1, characterized in that The basic feed includes the following raw materials in parts by weight: 18 parts of fish meal, 24.5 parts of soybean meal, 11 parts of peanut bran, 10 parts of chicken meal, 6 parts of shrimp meal, 21.65-21.75 parts of flour, 2 parts of fish oil, 2.5 parts of soybean oil, 1.5 parts of phosphorus and calcium nutritional additives, 0.2 parts of shrimp vitamins, 0.5 parts of shrimp minerals, 0.3 parts of amino acids, 0.2 parts of amino acid synthesis promoter, 0.3 parts of salt, and 1.2 parts of adhesive.

3. The vannamei shrimp feed according to claim 2, characterized in that The phosphate calcium nutritional additive is calcium dihydrogen phosphate; the adhesive is sodium alginate; the amino acid is methionine; and the amino acid synthesis promoter is choline chloride.

4. The vannamei shrimp feed according to claim 2, characterized in that The vitamins per kilogram of shrimp include: vitamin A 4000000 IU, vitamin D 32000000 IU, vitamin E 30g, vitamin K3 10g, vitamin B15g, vitamin B2 15g, vitamin B6 8g, vitamin B12 0.02g, niacin 40g, calcium pantothenate 25g, folic acid 2.5g, inositol 150g and biotin 0.08g.

5. The vannamei shrimp feed according to claim 2, characterized in that The minerals per kilogram of shrimp include: 12g of magnesium sulfate monohydrate, 90g of sodium chloride, 3g of metallic copper, 1g of ferrous sulfate monohydrate, 0.06g of calcium iodate, 0.16g of cobalt methionine, 10g of zinc sulfate monohydrate and 0.0036g of sodium selenite.

6. The method for preparing the Litopenaeus vannamei feed according to any one of claims 1 to 5, characterized in that: The following steps are involved: The raw materials are crushed and sieved, mixed and then granulated, matured and dried in the shade to obtain the vannamei shrimp feed.

7. The method for preparing the Litopenaeus vannamei feed according to claim 6, characterized in that: The aging temperature is 55-65° C. and the aging time is 2.5-3.5 hours.

8. The method for preparing the vannamei shrimp feed according to claim 6, characterized in that: The aging temperature is 60° C. and the time is 3 hours.

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