Feed additive and application thereof in improvement of aquatic animal protein anabolism

Abstract

The invention belongs to the technical field of feed additive production, and particularly relates to a feed additive and application thereof in improving aquatic animal protein anabolism. The invention provides a feed additive which comprises bacillus lentus powder, an amino acid compound, a plant extract and multivitamins, can be widely applied to the field of aquaculture, and can effectively promote the growth of bred animals, remarkably improve the oxidation resistance and immunity of the bred animals and reduce the occurrence of diseases. The feed additive can directly act on a muscle mTOR signal channel, protein synthesis is promoted, the meat structure is improved, the protein content is increased, and the application prospect is wide.

Description

Technical Field

[0001] The invention belongs to the technical field of feed additive production, and in particular relates to a feed additive and application thereof in improving protein anabolism of aquatic animals. Background Art

[0002] Studies have shown that fish demand for feed protein significantly higher than terrestrial animals, 2 to 4 times that of terrestrial animals. Protein content and feed amino acid levels are the main factors affecting fish growth, reproduction, health and feed composition. When there is insufficient protein in the feed, fish will inevitably consume protein from non-essential tissues to maintain the normal functions of more important tissues, resulting in growth stunting and reduced body weight. Amino acids are not only the basic building blocks of protein, but can also act as signal molecules to play a specific role in regulating key metabolic pathways. It has been confirmed in many fish that some functional amino acids play a key role in promoting growth and enhancing metabolic function.

[0003] At present, the main source of animal protein for aquatic feed is fish meal, but the fluctuation and decline of fish meal resources has led to an increase in its price, which has seriously restricted the development of the aquatic feed industry. At present, fish meal is mainly replaced by animal and plant proteins that are widely available, low in price and rich in protein. However, plant proteins such as soybean meal, fermented soybean meal and corn gluten meal have disadvantages such as poor palatability and many anti-nutritional factors; animal protein (such as poultry by-product meal and chicken meal) is expensive and the quality is difficult to guarantee. Excessive replacement of fish meal with animal and plant proteins will cause lipid metabolism and protein metabolism disorders. Therefore, improving the anabolism of aquatic animals' own proteins is a research hotspot in this field. Summary of the invention

[0004] The purpose of the present invention is to provide a feed additive and its application in improving protein anabolism of aquatic animals.

[0005] In order to achieve the above object, the present invention adopts the following technical solution:

[0006] In one aspect, the present invention provides a feed additive, comprising Bacillus lentus powder, an amino acid complex, a plant extract and a complex vitamin.

[0007] The Bacillus lentus is Bacillus lentus SK001, which was deposited in the China Type Culture Collection of Wuhan University, Wuhan, China on September 2, 2024, and its deposit number is CCTCC NO: M20241882.

[0008] The live bacteria quantity of the Bacillus lentus powder is not less than 1.0×10 10 CFU / g.

[0009] The components and their weight proportions in the feed additive are: 30-65 parts of Bacillus lentus powder, 30-50 parts of amino acid complex, 20-30 parts of plant extract and 10-15 parts of complex vitamins.

[0010] The components in the feed additive and their weight proportions are: 50 parts of Bacillus lentus powder, 30 parts of amino acid complex, 30 parts of plant extract and 15 parts of complex vitamins.

[0011] Further, the components of the amino acid complex are L-methionine, L-lysine, L-glutamine, L-threonine, L-arginine and L-phenylalanine, and the weight ratios thereof are 10:8-9:3-5:3-5:4-5:3-4, respectively;

[0012] The components of the plant extract are astragalus extract, yam extract and rehmannia extract, and the weight ratios thereof are 5:4:3 respectively;

[0013] The components of the vitamin complex are vitamin A, vitamin B1, vitamin B2, vitamin B5, vitamin B6, vitamin B 12 , vitamin D3, vitamin E, vitamin K3, biotin, folic acid, D-pantothenic acid, niacin, and their weight ratios are 15:15:7-10:7-10:6-8:4-8:4-8:7:6:3-5:4-5:4-5:3-4 respectively;

[0014] The invention also provides application of the feed additive in aquatic feed production.

[0015] The feed additive is added in an amount of 10-30 kg / ton in aquatic feed.

[0016] The beneficial effects of the present invention are as follows:

[0017] The feed additive provided by the present invention can be widely used in the field of aquaculture, can effectively promote the growth of farmed animals, significantly improve their antioxidant capacity and immunity, and reduce the occurrence of diseases. Compared with the control group, the fatness, intestine-body ratio, and visceral-body index of the grass carp in the experimental group fed with the feed additive described in Examples 6-8 were increased by 11.88%-20.63%, 11.81%-24.41%, and 16.14%-29.59%, respectively; the total protein, albumin, and urea nitrogen levels in the serum of the grass carp were increased by 24.29%-39.53%, 14.68%-22.22%, and 39.47%-107.61%, respectively, and the activities of aspartate aminotransferase and alanine aminotransferase were reduced by 14.89%-44.05% and 21.15%-42.01%, respectively, with significant effects.

[0018] The feed additive can significantly increase the protein content in grass carp muscle and reduce the fat content. Compared with the control group, the crude protein content of the grass carp back muscle in the experimental group fed with the feed additives described in Examples 6-8 increased by 8.86%-12.06%, and the crude fat content decreased by 15.01%-30.02%, among which the feed additive described in Example 6 had the best feeding effect; the triglyceride, total cholesterol, low-density lipoprotein, and free fatty acid contents in the serum of the grass carp in the experimental group were respectively reduced by 16.00%-73.43%, 19.02%-26.65%, 35.38%-43.97%, and 9.87%-23.15% compared with the control group, and the high-density lipoprotein content was increased by 9.43%-49.88%, achieving unexpected technical effects.

[0019] The feed additive can significantly activate the mTOR signaling pathway and promote protein synthesis in grass carp. Compared with the control group, the expression levels of Akt, S6 and 4E-BP1, key factors of the TOR signaling pathway in the grass carp muscle of the experimental group fed with the feed additives described in Examples 6-8 increased by 19%-183%, 32%-225%, and 30%-165%, respectively, indicating that the mTOR signaling pathway in the grass carp muscle was enhanced and activated, and the phosphorylation level of related proteins in the mTOR signaling pathway was significantly increased (P < 0.05), among which the effect of the feed additive described in Example 2 was the most significant, achieving unexpected technical effects.

[0020] In summary, the feed additive provided by the present invention can be widely used in the field of aquaculture, can effectively promote the growth of farmed animals, significantly improve their antioxidant capacity and immunity, and reduce the occurrence of diseases. The feed additive can directly act on the muscle mTOR signaling pathway, promote protein synthesis, help improve meat structure, increase protein content, and has broad application prospects. BRIEF DESCRIPTION OF THE DRAWINGS

[0021] Figure 1 This is the colony morphology of Bacillus lentus SK001. DETAILED DESCRIPTION

[0022] The following will be combined with the drawings in the embodiments of the present invention to clearly and completely describe the technical solutions in the embodiments of the present invention. Obviously, the described embodiments are only part of the embodiments of the present invention, not all of the embodiments. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without creative work are within the scope of protection of the present invention.

[0023] The culture medium selected in the embodiments of the present invention is listed below, which is only for illustration and is not intended to limit the present invention.

[0024] Example 1 Screening, identification and functional analysis of Bacillus lentus SK001

[0025] A strain with the strongest protease production ability was screened from fresh squid viscera purchased from Manyuwan Farmers' Market in Licang District, Qingdao City. It was identified as Bacillus lentus and named Bacillus lentus SK001. It was deposited in the China Type Culture Collection of Wuhan University, Wuhan, China on September 2, 2024, and its preservation number is CCTCC NO: M20241882.

[0026] The colony morphology of Bacillus lentus SK001 on nutrient agar medium is as follows Figure 1 As shown, the colony is 1-2mm in diameter, round, with neat edges, opaque, grayish white on the front, convex in the middle, smooth, bright on the surface, moist in texture, and easy to pick up.

[0027] Bacillus lentus SK001 has good biosafety and is sensitive to common antibiotics such as ampicillin, furazolidone, chloramphenicol, tetracycline, penicillin G, and gentamicin.

[0028] The Bacillus lentus SK001 provided by the present invention has a strong tolerance to the digestive tract environment. After being treated in artificial gastric juice and artificial intestinal juice for 3 hours, its survival rate is still as high as over 85%. It can enter the intestine through the stomach better, exert its probiotic effect, and achieve unexpected technical effects.

[0029] The Bacillus lentus SK001 can produce a high amount of protease, and the protease activity in the fermentation supernatant reaches 389.1 U / mL, which is significantly higher than that of the commercially available Bacillus lentus in the control group, achieving an unexpected technical effect.

[0030] Example 2 Analysis of antibacterial properties of Bacillus lentus SK001

[0031] Centrifuge the 18-20h Bacillus lentus SK001 nutrient broth culture at 12000r / min for 5min, take the supernatant, and place it in a 4℃ refrigerator for later use. 6 100 μL of indicator bacteria solution with CFU / mL was evenly spread on the LB solid plate, and an Oxford cup was placed. 200 μL of fermentation supernatant of Bacillus lentus SK001 was added, and cultured at 37°C for 24 hours to observe whether an inhibition zone was produced and measure the size of the inhibition zone. At the same time, commercially available Bacillus lentus was used as a control.

[0032] Table 1 Antibacterial activity of Bacillus lentus SK001

[0033]

[0034] From the data in Table 1, it can be seen that the Bacillus lentus SK001 provided by the present invention can effectively inhibit various pathogens such as Staphylococcus aureus, Escherichia coli, Bacillus cereus, Vibrio parahaemolyticus, Aeromonas hydrophila, Pseudomonas fluorescens, Escherichia coli, etc., especially the strongest antibacterial effect on Staphylococcus aureus and Escherichia coli, with the diameter of the inhibition zone exceeding 22 mm, which is significantly better than the commercially available Bacillus lentus.

[0035] Example 3 Analysis of the self-aggregation and hydrophobicity of Bacillus lentus

[0036] The self-aggregation and surface hydrophobicity of probiotics are important factors that reflect the strength of their adhesion ability and whether they can colonize in the intestines and play a probiotic role. The greater the hydrophobicity of the cell surface, the stronger its adhesion. The self-aggregation of probiotics refers to the phenomenon that the bacteria aggregate into clusters due to large-scale and rapid reproduction during the cultivation process. The stronger the surface hydrophobicity and self-aggregation, the stronger its intestinal adhesion.

[0037] Inoculate Bacillus in LB liquid culture medium and culture at 37℃ for 18h. Centrifuge the cultured bacterial solution at 4℃ and 12000r / min for 5min to collect the bacteria, and wash twice with sterile saline to remove the residual liquid of the culture medium. Adjust the OD600 of the bacterial suspension to 0.5 (A0). Take 2mL of the bacterial suspension and let it stand at room temperature for 3h, take 200μL of the upper bacterial solution, and measure the OD value at 600nm (A1); take another 2mL of the bacterial suspension and mix it with an equal volume of xylene for 2min, let it stand for 30min, and take the aqueous phase to measure the OD at 600nm (A2). Use saline as a blank.

[0038] The calculation formula of self-aggregation and hydrophobicity is:

[0039] Self-aggregation (%) = (1-A1 / A0) x 100%.

[0040] Hydrophobicity (%) = [(A0-A2) / A0] x 100%.

[0041] Where: A0 is the OD value of the initial bacterial solution at 600nm; A1 is the OD value of the supernatant at 600nm after standing for 3 hours; A2 is the OD value of the aqueous phase at 600nm after mixing with xylene.

[0042] Table 2 Self-aggregation and hydrophobicity of Bacillus lentus SK001

[0043]

[0044] As can be seen from Table 2, the self-aggregation and hydrophobicity of Bacillus lentus SK001 are as high as 43.1% and 56.1%, respectively, which are higher than those of commercially available strains, thus facilitating its better colonization in the intestinal tract of aquatic organisms.

[0045] Example 4 Application of Bacillus lentus SK001 in the cultivation of Litopenaeus vannamei

[0046] 1. Preparation of bacterial powder

[0047] The Bacillus lentus SK001 was activated, expanded, and fermented in liquid form, centrifuged, concentrated, and freeze-dried to produce a culture medium with a live bacterial content of about 10 10 CFU / g of bacterial powder.

[0048] 2. Experimental design

[0049] The experiment set up a control group and a probiotic group. The control group was fed with basic feed. The probiotic group added the above-mentioned Bacillus lentus SK001 powder to the basic feed at a weight ratio of 3‰, mixed evenly and fed. Three parallels were set up for each group, with 50 shrimps in each parallel. The breeding experiment lasted for 8 weeks, and the daily feeding amount was 3% of the total weight of the shrimp. The shrimps were fed twice a day, and the shrimps were weighed every 2 weeks to adjust the daily feeding amount. The dry matter weight of the feed fed to the control group and the probiotic group was consistent. The breeding water was replaced once a week, and the water change amount was 1 / 4 of the total water volume. During the experiment, the dissolved oxygen was ≥8.3mg / l, the temperature was 25-29℃, the salinity was 6-7‰, and the pH was 7.5-7.9. After the breeding experiment, the shrimps were counted and weighed, and fasted for one day before weighing. Its survival rate, specific growth rate, weight gain rate, etc. were calculated. Hepatopancreatic T-AOC, superoxide dismutase, and lysozyme were detected using the Nanjing Jiancheng Bioengineering Institute kit. The specific results are shown in Tables 3 and 4.

[0050] Table 3 Effects of Bacillus lentus SK001 on growth and survival of Litopenaeus vannamei

[0051] Control group Probiotics group Initial body weight / g 5.41±0.12 5.37±0.23 Final body weight / g 14.42±0.56 16.21±0.27 Survival rate / % <![CDATA[74.34±3.33 a ]]> <![CDATA[93.12±3.54 b ]]> Weight gain rate / % <![CDATA[166.54±20.25 a ]]> <![CDATA[201.86±15.92 b ]]> Specific growth rate / % <![CDATA[3.50±0.11 a ]]> <![CDATA[3.95±0.14 b <!-- 4 -->]]> Feed coefficient <![CDATA[1.73±0.21 a ]]> <![CDATA[1.41±0.35 b ]]>

[0052] Note: Different letters indicate significant differences (P<0.05).

[0053] Table 4 Effects of Bacillus lentus SK001 on immunity of Litopenaeus vannamei

[0054] Project (U / mg prot) Control group Probiotics group T-AOC 2.12±0.13 2.47±0.15 Superoxide dismutase 43.52±2.10 43.46±2.43 Lysozyme activity <![CDATA[101.30±7.67 b ]]> <![CDATA[143.82±5.32 a ]]>

[0055] Note: Different letters indicate significant differences (P<0.05).

[0056] From the results of Table 3 and Table 4, it can be seen that compared with the control group, the survival rate, weight gain rate, and specific growth rate of the probiotic group of shrimp fed with Bacillus lentus SK001 powder increased by 25.3%, 21.2%, and 12.9%, respectively, and the feed coefficient decreased by 18.5%; and the hepatopancreatic lysozyme activity of the probiotic group increased by 42.0%. This shows that the Bacillus lentus SK001 provided by the present invention can better induce the production of immune factors, actively participate in the body's immune defense, thereby improving the body's immune defense, and achieving unexpected technical effects.

[0057] Example 5 Application of Bacillus lentus SK001 in freshwater grouper farming

[0058] Freshwater grouper with an average weight of about 60g was selected, and a control group and a probiotic group were set. The control group was fed with basic bait, and the probiotic group was fed with the lentus SK001 powder described in Example 4 at a weight ratio of 5‰ on the basis of the basic bait, and the mixture was mixed and fed. Three parallels were set for each group, and each parallel had 40 freshwater groupers, and the water temperature was controlled to be 22-25°C. The feeding time was 9:00 and 17:00 respectively. The feeding time at 9:00 should be appropriate. After 0.5h, the feeding situation of the fish was observed and recorded. The feeding amount was adjusted appropriately according to the feeding situation, and the remaining bait was removed after 0.5h of feeding. The bottom of the breeding barrel was sucked every morning and evening, and 1 / 3 of the breeding water was replaced. The replaced water was disinfected, aerated, and tap water with the same temperature. Feeding and breeding for 28d.

[0059] On the 28th day of the experiment, 3 freshwater groupers were selected from each barrel, weighed, and measured in length, and the weight gain rate, specific growth rate, survival rate, fatness, liver-body ratio, organ-body ratio, and feed coefficient were calculated. The fish were starved for 24 hours the day before blood collection, and blood was collected from the tail vein. The blood samples were centrifuged at 4°C and 4000r / min for 10 minutes, and the supernatant was taken to determine the antioxidant index, which was determined using the Nanjing Jiancheng Bioengineering Research Institute kit. The results are shown in Tables 5-7.

[0060] Table 5 Effects of Bacillus lentus SK001 on the growth performance of freshwater grouper

[0061] Group Control group Probiotics group Weight gain rate / % <![CDATA[57.61±0.15 a ]]> <![CDATA[98.42±0.74 b ]]> Specific growth rate / (% / d) <![CDATA[2.31±0.36 a ]]> <![CDATA[3.53±0.41 b ]]> Liver to body ratio / % 1.53±0.18 1.64±0.31 Viscera to body ratio / % 3.32±0.26 3.73±0.22 <![CDATA[Condition factor / (g / cm 3 )]]> 1.25±0.29 1.42±0.27 Feed coefficient 1.62±0.24 1.13±0.24 Survival rate / % 95 100

[0062] Table 6 Effects of Bacillus lentus SK001 on the immune capacity of freshwater grouper

[0063]

[0064] Table 7 Effects of Bacillus lentus SK001 on serum liver function indicators of freshwater grouper

[0065] index Control group Probiotics group Aspartate aminotransferase / (U / L) <![CDATA[21.32±1.47 a ]]> <![CDATA[15.73±0.54 b ]]> Alanine aminotransferase / (U / L) <![CDATA[21.07±1.22 a ]]> <![CDATA[15.62±1.42 a ]]>

[0066] From the data in Table 5, it can be seen that compared with the control group, the weight gain rate of the freshwater grouper in the probiotic group fed with Bacillus lentus SK001 powder increased by 70.8% (P<0.05), the specific growth rate increased by 52.8% (P<0.05), and the growth performance was significantly improved.

[0067] From the data in Table 6, it can be seen that compared with the control group, the activities of total superoxide dismutase and glutathione peroxidase in the serum of freshwater grouper in the probiotic group fed with Bacillus lentus SK001 powder increased by 24.2% and 13.5%, respectively, and the contents of catalase and malondialdehyde decreased by 126.7% and 9.5%, respectively.

[0068] As shown in Table 7, compared with the control group, the levels of aspartate aminotransferase and alanine aminotransferase in the serum of freshwater grouper in the probiotic group fed with Bacillus lentus SK001 powder decreased by 35.5% and 34.9%, respectively.

[0069] The above results show that the Bacillus lentus SK001 provided by the present invention can significantly improve the antioxidant capacity and immune level in the body of freshwater grouper, reduce liver damage in aquatic animals, and effectively promote the growth of farmed animals.

[0070] Example 6 A feed additive for improving protein anabolism in aquatic animals

[0071] A feed additive, the components and weight proportions of which are 50 parts of Bacillus lentus SK001 powder, 30 parts of amino acid complex, 30 parts of plant extract, and 15 parts of complex vitamins; wherein:

[0072] The components of the amino acid complex are L-methionine, L-lysine, L-glutamine, L-threonine, L-arginine and L-phenylalanine, and the weight ratios thereof are 10:8:3:3:5:4 respectively;

[0073] The components of the plant extract are astragalus extract, yam extract and rehmannia extract, and the weight ratios thereof are 5:4:3 respectively;

[0074] The components of the vitamin complex are vitamin A, vitamin B1, vitamin B2, vitamin B5, vitamin B6, vitamin B 12 , vitamin D3, vitamin E, vitamin K3, biotin, folic acid, D-pantothenic acid, and niacin, in weight ratios of 15:15:7:7:7:5:5:7:6:5:5:4:4 respectively.

[0075] Example 7 A feed additive for improving protein anabolism in aquatic animals

[0076] A feed additive, the components and weight proportions of which are 30 parts of Bacillus lentus SK001 powder, 50 parts of amino acid complex, 20 parts of plant extract, and 10 parts of complex vitamins; wherein:

[0077] The components of the amino acid complex are L-methionine, L-lysine, L-glutamine, L-threonine, L-arginine and L-phenylalanine, and the weight ratios thereof are 10:8:5:5:4:4 respectively;

[0078] The components of the plant extract are astragalus extract, yam extract and rehmannia extract, and the weight ratios thereof are 5:4:3 respectively;

[0079] The components of the vitamin complex are vitamin A, vitamin B1, vitamin B2, vitamin B5, vitamin B6, vitamin B 12 , vitamin D3, vitamin E, vitamin K3, biotin, folic acid, D-pantothenic acid, and niacin, in weight ratios of 15:15:10:10:8:8:8:7:6:4:4:4:4 respectively.

[0080] Example 8 A feed additive for improving protein anabolism in aquatic animals

[0081] A feed additive, the components and weight proportions of which are 65 parts of Bacillus lentus SK001 powder, 35 parts of amino acid complex, 20 parts of plant extract, and 10 parts of complex vitamins; wherein:

[0082] The components of the amino acid complex are L-methionine, L-lysine, L-glutamine, L-threonine, L-arginine and L-phenylalanine, and the weight ratios thereof are 10:9:5:5:5:3 respectively;

[0083] The components of the plant extract are astragalus extract, yam extract and rehmannia extract, and the weight ratios thereof are 5:4:3 respectively;

[0084] The components of the vitamin complex are vitamin A, vitamin B1, vitamin B2, vitamin B5, vitamin B6, vitamin B 12 , vitamin D3, vitamin E, vitamin K3, biotin, folic acid, D-pantothenic acid, and niacin, in weight ratios of 15:15:8:8:6:4:4:7:6:3:5:5:3 respectively.

[0085] Example 9 Application of feed additives for improving protein anabolism in aquatic animals in grass carp farming

[0086] 9.1 Experimental Design

[0087] The experimental subjects are grass carp weighing about 600 grams. They are acclimated for 2 weeks before the formal breeding experiment to ensure that the farmed fish adapt to the breeding system. Before the experiment, the experimental fish are starved for 24 hours, and those with consistent specifications and strong physique are selected and randomly distributed in a breeding barrel (300L). The experiment is divided into a control group and an experimental group, with 3 replicates in each group and 30 fish in each replicate. The control group uses conventional aquatic feed, and the experimental group adds the feed additive of the present invention on the basis of conventional feed, and the addition amount is 2% of the mass of the aquatic basic feed. The experimental fish are fed at 6:00 and 16:00 every day to confirm that the experimental fish have reached a state of satiation. During the test period, the water temperature is maintained at 27-30°C, the pH value is 7.5-8.0, and the dissolved oxygen is greater than 7mg / L. The experimental period is 8 weeks. After the feeding is completed, fast for 24 hours, and the total weight of each barrel of fish is weighed. After eugenol anesthesia is performed on 3 fish randomly selected in each replicate, tail venous blood is collected, centrifuged at 3500r / min at 4°C for 10min, and the upper serum is collected and placed under -80°C for standby use. The muscle, liver, intestine and other tissues were separated and placed in a -20℃ refrigerator for later use. Three fish were randomly selected from each experimental group to measure their visceral weight, liver weight, body length, body width, intestinal weight and length, etc.

[0088] 9.2 Measurement indicators and methods

[0089] 9.2.1 Determination of growth performance and morphological indicators

[0090] Fatness (%) = final weight / body length 3×100%.

[0091] Hepatopancreas index (%) = hepatopancreas weight / final weight × 100%.

[0092] Spleen index (%) = hepatopancreas weight / final weight × 100%.

[0093] Intestine to body ratio = intestinal weight / end weight × 100%.

[0094] Visceral index = visceral mass weight / final weight × 100%.

[0095] Head-kidney index = head-kidney weight / end weight × 100%.

[0096] Table 8 Effects of feed additives on growth performance of grass carp

[0097]

[0098] From the data in Table 8, it can be seen that compared with the control group, the fatness, intestine-body ratio, and visceral-body index of the grass carp in the experimental group fed with the feed additives described in Examples 6-8 increased by 11.88%-20.63%, 11.81%-24.41%, and 16.14%-29.59%, respectively. This shows that the feed additive provided by the present invention can effectively promote the growth of farmed animals and significantly improve their various growth performances, with remarkable effects.

[0099] 9.2.2 Determination of routine nutritional components of fish

[0100] The crude protein content was determined by the Kjeldahl method, with reference to GB5009.5; the crude fat content was determined by the Soxhlet extraction method, with reference to GB / T 5009.6; the ash content was determined by the muffle furnace ignition method, with reference to GB 5009.4.

[0101] Table 9 Effects of feed additives on grass carp muscle composition

[0102] Control group Example 6 Example 7 Example 8 Crude protein CP / % 83.83±0.21a 93.94±0.38c 91.47±0.42bc 91.26±0.11b Crude fat EE / % 4.53±0.27a 3.17±0.33b 3.72±0.42b 3.85±0.36b

[0103] It can be seen from the data in Table 9 that compared with the control group, the crude protein content of the grass carp back muscle in the experimental group fed with the feed additives described in Examples 6-8 increased by 8.86%-12.06%, and the crude fat content of the back muscle decreased by 15.01%-30.02%. Among them, the feed additive described in Example 6 had the best feeding effect, with the highest crude protein content and the lowest crude fat content in the grass carp back muscle.

[0104] 9.2.3 Determination of serum protein synthesis capacity

[0105] The activities of aspartate aminotransferase (AST) and alanine aminotransferase (ALT) and the levels of albumin (ALB), total protein (TP) and urea nitrogen (BUN) in serum were determined using kits produced by Nanjing Jiancheng Bioengineering Institute.

[0106] Table 10 Effects of feed additives on serum protein synthesis indices of grass carp

[0107]

[0108] It can be seen from the data in Table 10 that compared with the control group, the levels of total protein, albumin and urea nitrogen in the serum of grass carp in the experimental group fed with the feed additives described in Examples 6-8 increased by 24.29%-39.53%, 14.68%-22.22% and 39.47%-107.61%, respectively, and the activities of aspartate aminotransferase and alanine aminotransferase decreased by 14.89%-44.05% and 21.15%-42.01%, respectively, with significant effects.

[0109] 9.2.4 Determination of lipid metabolism indicators in serum

[0110] Serum triglyceride (TG), total cholesterol (T-CHO), high-density lipoprotein cholesterol (HDL-C), low-density lipoprotein cholesterol (LDL-C), and free fatty acids (FFA) were measured using kits produced by Nanjing Jiancheng Bioengineering Institute.

[0111] Table 11 Effects of feed additives on lipid metabolism indicators in serum

[0112]

[0113]

[0114] It can be seen from the data in Table 11 that compared with the control group, the triglyceride, total cholesterol, low-density lipoprotein and free fatty acid contents in the serum of the grass carp in the experimental group fed with the feed additives described in Examples 6-8 were reduced by 16.00%-73.43%, 19.02%-26.65%, 35.38%-43.97% and 9.87%-23.15%, respectively, and the high-density lipoprotein was increased by 9.43%-49.88%, and the effect was very significant.

[0115] 9.2.5 Serum antioxidant capacity and nonspecific immune index detection

[0116] Serum total antioxidant capacity (T-AOC), superoxide dismutase (SOD), catalase (CAT), and lysozyme (LYZ) activities were determined using kits from Nanjing Jiancheng Bioengineering Institute.

[0117] Table 12 Effects of feed additives on serum biochemical indicators

[0118]

[0119] From the data in Table 12, it can be seen that the total antioxidant capacity (T-AOC), superoxide dismutase (SOD), catalase (CAT), and lysozyme (LYZ) activities of the grass carp serum in the experimental group fed with the feed additives described in Examples 6-8 are higher than those in the control group, indicating that the additives of the present invention can improve the immune response and disease resistance of grass carp.

[0120] 9.2.6 Muscle Protein Synthesis Signaling Pathway

[0121] Total RNA from grass carp muscle was extracted according to the literature, and the content and integrity of RNA were detected by agarose gel electrophoresis and spectrophotometer. Reverse transcription was performed using a reverse transcription kit, and the CFX96 real-time quantitative PCR detection system was used.

[0122] Table 13 Effects of feed additives on the expression of key factors of TOR signaling pathway in grass carp muscle

[0123] TOR Akt S6 4E-BP1 Control group <![CDATA[1.00±0.05 a ]]> <![CDATA[1.00±0.63 a ]]> <![CDATA[1.00±0.31 a ]]> <![CDATA[1.00±0.21 a ]]> Example 6 <![CDATA[1.27±0.35 b ]]> <![CDATA[2.83±0.21 c ]]> <![CDATA[3.25±0.21 d ]]> <![CDATA[2.65±0.28 c ]]> Example 7 <![CDATA[1.15±0.28 b ]]> <![CDATA[1.28±0.13 c ]]> <![CDATA[1.46±0.32 c ]]> <![CDATA[1.39±0.19 d <!-- 10 -->]]> Example 8 <![CDATA[1.16±0.73 b ]]> <![CDATA[1.19±0.17 b ]]> <![CDATA[1.32±0.22 c ]]> <![CDATA[1.30±0.32 d ]]>

[0124] From the data in Table 13, it can be seen that compared with the control group, the expression levels of key factors Akt, S6 and 4E-BP1 in the TOR signaling pathway of grass carp muscle in the experimental group fed with the feed additives described in Examples 6-8 increased by 19%-183%, 32%-225%, and 30%-165%, respectively, indicating that the mTOR signaling pathway in grass carp muscle was enhanced and activated, and the phosphorylation level of related proteins in the mTOR signaling pathway was significantly increased (P < 0.05), among which the effect of the feed additive described in Example 2 was the most significant. This shows that the feed additive provided by the present invention can directly act on the mTOR signaling pathway, significantly promote protein synthesis in grass carp, and achieve unexpected technical effects.

[0125] In summary, the feed additive provided by the present invention can be widely used in the field of aquaculture, can effectively promote the growth of farmed animals, significantly improve their antioxidant capacity and immunity, and reduce the occurrence of diseases. The feed additive can directly act on the muscle mTOR signaling pathway, promote protein synthesis, help improve meat structure, increase protein content, and has broad application prospects.

Claims

1. A feed additive, characterized in that: The feed additive contains Bacillus lentus ( Bacillus lentus ) Bacterial powder, amino acid complex, plant extract, and multivitamin.

2. The feed additive according to claim 1, characterized in that The accession number of the Bacillus lentus contained in the Bacillus lentus powder is CCTCC NO: M20241882.

3. The feed additive according to claim 2, characterized in that The live bacteria quantity of the Bacillus lentus powder is not less than 1.0×10 10 CFU / g.

4. The feed additive according to claim 1, characterized in that The components in the feed additive and their weight proportions are: 30-65 parts of Bacillus lentus powder, 30-50 parts of amino acid complex, 20-30 parts of plant extract and 10-15 parts of complex vitamins.

5. The feed additive according to claim 4, characterized in that The components in the feed additive and their weight proportions are: 50 parts of Bacillus lentus powder, 30 parts of amino acid complex, 30 parts of plant extract and 15 parts of complex vitamins.

6. The feed additive according to claim 1, characterized in that The components of the amino acid complex are L-methionine, L-lysine, L-glutamine, L-threonine, L-arginine and L-phenylalanine, and the weight ratios thereof are 10:8-9:3-5:3-5:4-5:3-4 respectively.

7. The feed additive according to claim 1, characterized in that The components of the plant extract are astragalus extract, yam extract and rehmannia extract, and the weight ratios thereof are 5:4:3 respectively.

8. The feed additive according to claim 1, characterized in that The components of the complex vitamin are respectively vitamin A, vitamin B1, vitamin B2, vitamin B5, vitamin B6, vitamin B12, vitamin D3, vitamin E, vitamin K3, biotin, folic acid, D-pantothenic acid and niacin, and the weight ratios thereof are respectively 15:15:7-10:7-10:6-8:4-8:4-8:7:6:3-5:4-5:4-5:3-4.

9. Use of the feed additive according to any one of claims 1 to 8 in aquatic feed production.

10. The use according to claim 9, characterized in that The feed additive is added in an amount of 10-30 kg / ton in aquatic feed.

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