Composition containing astragalus polysaccharide and fructus forsythiae extract and application of composition in larimichthys crocea juvenile fish feed

Abstract

The invention relates to the technical field of breeding. The invention provides a composition containing astragalus polysaccharide and a fructus forsythiae extract, the mass ratio of the astragalus polysaccharide to the fructus forsythiae extract is 10: (2-15), and the composition can be used for preparing feed with the effects of resisting oxidation, relieving inflammation or improving growth rate and survival rate. The invention also provides a larimichthys crocea juvenile fish feed containing 0.1-0.2% of the composition, and the growth rate, the survival rate and the stress resilience of the larimichthys crocea juvenile fish are obviously improved by using the feed combined with the astragalus polysaccharide and the fructus forsythiae extract; the expression level of inflammatory factors is effectively reduced, and inflammation is relieved; the related digestive enzyme and villus length are also obviously enhanced, and the intestinal health is ensured; and an Nrf2 pathway is activated, and the expression level of related antioxidant enzymes is improved, so that the anti-oxidation defense capability is enhanced.

Description

Technical Field

[0001] The invention relates to the technical field of breeding, and in particular to a composition containing astragalus polysaccharide and forsythia suspensa extract and application of the composition in feed for juvenile large yellow croaker. Background Art

[0002] In recent years, the aquaculture industry has faced major challenges, especially viral and bacterial infections, which have caused significant damage to the industry. These infections are generally treated with highly toxic chemicals and antibiotics, which inadvertently also led to the emergence of drug-resistant pathogens, triggering restrictions or outright bans in various countries. In addition, Chinese herbal ingredients in aquaculture feeds have gained attention for their ability to improve the disease resistance of aquatic animals. Chinese herbal medicines play a key role in strengthening the immune defense of aquatic species such as fish, shrimp and crabs, effectively enhancing their antiviral, antibacterial and antiparasitic immune responses. Chinese herbal medicines have a wide range of bioactive components, including glycosides, polyphenols, organic acids, polysaccharides, alkaloids and quinones. These important active substances are intricately interconnected with various physiological functions of aquatic organisms, especially enhancing their immune defense, metabolic efficiency, anti-stress properties and growth potential. The main advantages of Chinese herbal medicines as additives include low adverse reactions, less residue accumulation, low environmental impact, and no drug resistance or related health problems. In addition, their natural origin ensures high acceptance and tolerance in aquatic animals. As a result, Chinese herbal medicines have gradually replaced chemical synthetics and hormones as feed additives, thereby improving the safety of aquaculture products for human consumption.

[0003] Large yellow croaker (Larimichthys crocea) is the main economic marine fish species in Chinese aquaculture, accounting for the largest share of marine fish production, with an annual production of 257,700 tons in 2022. However, the continued growth of aquaculture activities, as well as increasing stocking densities, continued aquatic environmental degradation and recurring extreme weather, have driven a sharp increase in disease outbreaks. The critical early life stages of large yellow croaker, namely larvae and juveniles, are particularly prone to high mortality, which is a key bottleneck that needs to be overcome in large yellow croaker farming. These juveniles are susceptible to oxidative stress and intestinal damage caused by environmental stressors.

[0004] In summary, the development of reliable and effective immunomodulators is very necessary, which has the potential to enhance the nonspecific immune defense of fish by enhancing antioxidant capacity and immune response, thereby reducing the incidence of diseases. Summary of the invention

[0005] The purpose of the present invention is to provide a composition containing astragalus polysaccharide and forsythia extract and its application in large yellow croaker fry feed. Through the use of the composition containing astragalus polysaccharide and forsythia extract, the growth rate, survival rate and stress resilience of large yellow croaker fry are significantly improved; the expression level of inflammatory factors is effectively reduced, and inflammation is relieved; the relevant digestive enzymes and villus length are significantly enhanced, ensuring intestinal health; the Nrf2 pathway is activated, the expression level of relevant antioxidant enzymes is increased, and the antioxidant defense ability is enhanced.

[0006] In order to achieve the above-mentioned object of the invention, the present invention provides the following technical solutions:

[0007] The invention provides a composition containing astragalus polysaccharide and forsythia suspensa extract, wherein the mass ratio of the astragalus polysaccharide to the forsythia suspensa extract is 10:(2-15).

[0008] Preferably, the Forsythia suspensa extract comprises forsythiaside A, forsythiaside and forsythiaside.

[0009] The invention provides application of the composition in preparing feed with antioxidant effect.

[0010] The invention provides application of the composition in preparing feed with the effect of alleviating inflammation.

[0011] The invention provides application of the composition in preparing feed with the efficacy of improving growth rate and survival rate.

[0012] The invention also provides a large yellow croaker fry feed, comprising the following components in percentage by mass: 38-42% of fish meal, 27-32% of soybean meal, 20-23% of flour, 2-3% of fish oil, 0.8-1.2% of soybean lecithin, 2-3% of brewer's yeast, 0.7-1.2% of complex vitamins, 0.8-1.2% of complex mineral salts and 0.1-0.2% of the composition.

[0013] The present invention also provides the use of the large yellow croaker fry feed in improving the growth rate and survival rate of large yellow croaker fry, alleviating inflammation of large yellow croaker fry, and enhancing the antioxidant capacity of large yellow croaker fry.

[0014] By adopting the above technical solution, the present invention has the following beneficial effects:

[0015] The technical solution of the present invention uses a composition containing astragalus polysaccharide and forsythia extract to significantly improve the growth rate, survival rate and stress resilience of yellow croaker fry; effectively reduce the expression level of inflammatory factors and relieve inflammation; also significantly enhance the relevant digestive enzymes and villus length to ensure intestinal health; also activate the Nrf2 pathway, increase the expression level of relevant antioxidant enzymes, and thus enhance the antioxidant defense ability. By comprehensively improving the immune function and environmental stress resistance of yellow croaker fry, the incidence of diseases can be reduced and the survival rate can be improved. BRIEF DESCRIPTION OF THE DRAWINGS

[0016] Figure 1 The midgut morphology of juvenile large yellow croaker in different treatment groups (n=4, scale bar is 100μm);

[0017] Figure 2 The length of mid-intestinal villi of juvenile large yellow croaker in different treatment groups (n=4);

[0018] Figure 3 The relative expression levels of inflammatory cytokines in the head kidney of juvenile large yellow croaker in different treatment groups;

[0019] Figure 4 The relative expression levels of antioxidant-related genes and apoptosis-related genes in the head kidney of large yellow croaker fry in different treatment groups. DETAILED DESCRIPTION

[0020] The present invention provides a composition containing Astragalus membranaceus polysaccharides (APS) and Forsythia suspensa extract (FSE), wherein the mass ratio of the Astragalus membranaceus polysaccharides to the Forsythia suspensa extract is preferably 10:(2-15), further preferably 10:(3-10), and further preferably 10:5.

[0021] In the present invention, the astragalus polysaccharide and the forsythia extract are purchased from Beijing Shengtaier Technology Co., Ltd., wherein the astragalus polysaccharide contains a net content of 60% polysaccharides, and the forsythia extract includes forsythiaside A, forsythiaside and forsythiaside. The forsythia extract of the present invention contains 35.6 mg / kg forsythiaside A, 37.2 mg / kg forsythiaside and 152.8 mg / kg forsythiaside.

[0022] The invention provides application of the composition in preparing feed with antioxidant effect.

[0023] The invention provides application of the composition in preparing feed with the effect of alleviating inflammation.

[0024] The invention provides application of the composition in preparing feed with the efficacy of improving growth rate and survival rate.

[0025] The invention also provides a large yellow croaker fry feed, comprising the following components: fish meal, soybean meal, flour, fish oil, soybean lecithin, brewer's yeast, complex vitamins, complex mineral salts and the above combination.

[0026] In the present invention, the large yellow croaker fry feed preferably includes the following components in percentage by weight: 38-42% fish meal, 27-32% soybean meal, 20-23% flour, 2-3% fish oil, 0.8-1.2% soybean lecithin, 2-3% brewer's yeast, 0.7-1.2% complex vitamins, 0.8-1.2% complex mineral salts and 0.1-0.2% of the above composition; further preferably includes 39-41% fish meal, 28-31% soybean meal, 21-22% flour , 2.3-2.8% fish oil, 0.9-1.1% soy lecithin, 2.2-2.8% brewer's yeast, 0.85-1.1% complex vitamins, 0.9-1.1% complex mineral salts and 0.12-0.17% of the above composition; further preferably including 40% fish meal, 30% soybean meal, 21.75% flour, 2.6% fish oil, 1% soy lecithin, 2.5% brewer's yeast, 1% complex vitamins, 1% complex mineral salts and 0.15% of the above composition.

[0027] In the present invention, the multivitamin preferably includes 245000-255000 IU / kg vitamin A, 720-780 mg / kg riboflavin, 380-420 mg / kg pyridoxine-hydrochloride, 0.8-1.2 mg / kg vitamin B12, 220-270 mg / kg ammonium sulfate, 220-280 mg / kg vitamin K, 100-150 mg / kg folic acid, 8-12 mg / kg biotin, 2-3 mg / kg vitamin E, 7800-8300 mg / kg inositol, 1200 -1300mg / kg calcium pantothenate, 1700-2200mg / kg niacin, 7500-8200mg / kg choline chloride, 44000-45500IU / kg vitamin D3, 6500-7500mg / kg vitamin C, further preferably including 248000-253000IU / kg vitamin A, 730-760mg / kg riboflavin, 390-410mg / kg pyridoxine-hydrochloride, 0.9-1.1mg / kg vitamin B12, 230-260mg / kg ammonium sulfate , 230-270 mg / kg vitamin K, 110-140 mg / kg folic acid, 9-11 mg / kg biotin, 2.2-2.8 mg / kg vitamin E, 7900-8200 mg / kg inositol, 1220-1280 mg / kg calcium pantothenate, 1800-2100 mg / kg niacin, 7800-8100 mg / kg choline chloride, 44500-45200 IU / kg vitamin D3, 6800-7200 mg / kg vitamin C, and more preferably 250000 IU / kg vitamin A, 750mg / kg riboflavin, 400mg / kg pyridoxine hydrochloride, 1mg / kg vitamin B12, 250mg / kg ammonium sulfate, 250mg / kg vitamin K, 125mg / kg folic acid, 10mg / kg biotin, 2.5mg / kg vitamin E, 8000mg / kg inositol, 1250mg / kg calcium pantothenate, 2000mg / kg niacin, 8000mg / kg choline chloride, 45000IU / kg vitamin D3, 7000mg / kg vitamin C.

[0028] In the present invention, the composite mineral salt preferably includes 0.03-0.05 g / kg zinc sulfate heptahydrate, 36-39 g / kg calcium carbonate, 4.5-5.8 g / kg potassium chloride, 0.03-0.05 g / kg potassium iodide, 2-3 g / kg sodium chloride, 0.015-0.025 g / kg copper sulfate pentahydrate, 0.015-0.025 g / kg cobalt sulfate heptahydrate, 0.6-1.2 g / kg iron sulfate heptahydrate, 0.02-0.04 g / kg manganese sulfate monohydrate, 3-4 g / kg magnesium sulfate heptahydrate, 9-11.5 g / kg calcium dihydrogen phosphate dihydrate, and further preferably includes 0.035-0.045 g / kg zinc sulfate heptahydrate, 37-38.3 g / kg calcium carbonate, 5-5.6 g / kg potassium chloride, 0.035-0.045 g / kg potassium iodide, 2.4 -2.8g / kg sodium chloride, 0.018-0.022g / kg copper sulfate pentahydrate, 0.018-0.022g / kg cobalt sulfate heptahydrate, 0.8-1.1g / kg ferric sulfate heptahydrate, 0.025-0.035g / kg manganese sulfate monohydrate, 3.2-3.8g / kg magnesium sulfate heptahydrate, 9.2-10.5g / kg calcium dihydrogen phosphate dihydrate, further preferably including 0.04g / kg zinc sulfate heptahydrate, 37.9g / kg calcium carbonate, 5.3g / kg potassium chloride, 0.04g / kg potassium iodide, 2.6g / kg sodium chloride, 0.02g / kg copper sulfate pentahydrate, 0.02g / kg cobalt sulfate heptahydrate, 0.9g / kg ferric sulfate heptahydrate, 0.03g / kg manganese sulfate monohydrate, 3.5g / kg magnesium sulfate heptahydrate, 9.8g / kg calcium dihydrogen phosphate dihydrate.

[0029] The present invention also provides the use of the large yellow croaker fry feed in improving the growth rate and survival rate of large yellow croaker fry, alleviating inflammation of large yellow croaker fry, and enhancing the antioxidant capacity of large yellow croaker fry.

[0030] The technical solutions provided by the present invention are described in detail below in conjunction with the embodiments, but they should not be construed as limiting the protection scope of the present invention.

[0031] The brewer's yeast described in the present invention was purchased from Fuzhou Haima Feed Co., Ltd.

[0032] Example 1

[0033] A composition containing astragalus polysaccharide and forsythia extract, wherein the mass ratio of astragalus polysaccharide to forsythia extract is 10:5, and the forsythia extract contains 35.6 mg / kg forsythiaside A, 37.2 mg / kg forsythiaside and 152.8 mg / kg forsythiaside.

[0034] Example 2

[0035] A composition containing astragalus polysaccharide and forsythia extract, wherein the mass ratio of the astragalus polysaccharide to the forsythia extract is 10:10, and the forsythia extract contains 35.6 mg / kg of forsythoside A, 37.2 mg / kg of forsythiaside and 152.8 mg / kg of forsythiaside.

[0036] Example 3

[0037] A composition containing astragalus polysaccharide and forsythia extract, wherein the mass ratio of astragalus polysaccharide to forsythia extract is 10:15, and the forsythia extract contains 35.6 mg / kg forsythoside A, 37.2 mg / kg forsythiaside and 152.8 mg / kg forsythiaside.

[0038] Example 4

[0039] A feed for juvenile yellow croaker, comprising the following components: 40% fish meal, 30% soybean meal, 21.75% flour, 2.6% fish oil, 1% soybean lecithin, 2.5% brewer's yeast, 1% complex vitamins, 1% complex mineral salts, and 0.15% of the composition described in Example 1;

[0040] The multivitamin comprises 250000 IU / kg vitamin A, 750 mg / kg riboflavin, 400 mg / kg pyridoxine-hydrochloride, 1 mg / kg vitamin B12, 250 mg / kg ammonium sulfate, 250 mg / kg vitamin K, 125 mg / kg folic acid, 10 mg / kg biotin, 2.5 mg / kg vitamin E, 8000 mg / kg inositol, 1250 mg / kg calcium pantothenate, 2000 mg / kg niacin, 8000 mg / kg choline chloride, 450 00IU / kg vitamin D3, 7000mg / kg vitamin C; the composite mineral salt is 0.04g / kg zinc sulfate heptahydrate, 37.9g / kg calcium carbonate, 5.3g / kg potassium chloride, 0.04g / kg potassium iodide, 2.6g / kg sodium chloride, 0.02g / kg copper sulfate pentahydrate, 0.02g / kg cobalt sulfate heptahydrate, 0.9g / kg iron sulfate heptahydrate, 0.03g / kg manganese sulfate monohydrate, 3.5g / kg magnesium sulfate heptahydrate, and 9.8g / kg calcium dihydrogen phosphate dihydrate.

[0041] Example 5

[0042] A feed for juvenile yellow croaker, comprising the following components: 38.09% fish meal, 31.2% soybean meal, 22% flour, 2.7% fish oil, 1.1% soybean lecithin, 2.5% brewer's yeast, 1.1% complex vitamins, 1.2% complex mineral salts, and 0.11% of the composition described in Example 1;

[0043] The multivitamin comprises 250000 IU / kg vitamin A, 750 mg / kg riboflavin, 400 mg / kg pyridoxine-hydrochloride, 1 mg / kg vitamin B12, 250 mg / kg ammonium sulfate, 250 mg / kg vitamin K, 125 mg / kg folic acid, 10 mg / kg biotin, 2.5 mg / kg vitamin E, 8000 mg / kg inositol, 1250 mg / kg calcium pantothenate, 2000 mg / kg niacin, 8000 mg / kg choline chloride, 450 00IU / kg vitamin D3, 7000mg / kg vitamin C; the composite mineral salt is 0.04g / kg zinc sulfate heptahydrate, 37.9g / kg calcium carbonate, 5.3g / kg potassium chloride, 0.04g / kg potassium iodide, 2.6g / kg sodium chloride, 0.02g / kg copper sulfate pentahydrate, 0.02g / kg cobalt sulfate heptahydrate, 0.9g / kg iron sulfate heptahydrate, 0.03g / kg manganese sulfate monohydrate, 3.5g / kg magnesium sulfate heptahydrate, and 9.8g / kg calcium dihydrogen phosphate dihydrate.

[0044] Example 6

[0045] A feed for juvenile yellow croaker, comprising the following components: 41% fish meal, 29% soybean meal, 22% flour, 2.2% fish oil, 0.9% soybean lecithin, 2.7% brewer's yeast, 1.1% complex vitamins, 0.9% complex mineral salts and 0.2% of the composition described in Example 1;

[0046] The multivitamin comprises 250000 IU / kg vitamin A, 750 mg / kg riboflavin, 400 mg / kg pyridoxine-hydrochloride, 1 mg / kg vitamin B12, 250 mg / kg ammonium sulfate, 250 mg / kg vitamin K, 125 mg / kg folic acid, 10 mg / kg biotin, 2.5 mg / kg vitamin E, 8000 mg / kg inositol, 1250 mg / kg calcium pantothenate, 2000 mg / kg niacin, 8000 mg / kg choline chloride, 450 00IU / kg vitamin D3, 7000mg / kg vitamin C; the composite mineral salt is 0.04g / kg zinc sulfate heptahydrate, 37.9g / kg calcium carbonate, 5.3g / kg potassium chloride, 0.04g / kg potassium iodide, 2.6g / kg sodium chloride, 0.02g / kg copper sulfate pentahydrate, 0.02g / kg cobalt sulfate heptahydrate, 0.9g / kg iron sulfate heptahydrate, 0.03g / kg manganese sulfate monohydrate, 3.5g / kg magnesium sulfate heptahydrate, and 9.8g / kg calcium dihydrogen phosphate dihydrate.

[0047] Test Example 1

[0048] (I) Preparation of experimental feed

[0049] Five experimental diets with equal nitrogen and lipid content were set up, including the control group, APS diet group, 0.05% FSE group, 0.1% FSE group and 0.15% FSE group. The control group was a diet without APS and FSE; the APS diet group was a diet with 0.1% APS added; the 0.05% FSE group was a diet with 0.1% APS and 0.05% FSE added; the 0.1% FSE group was a diet with 0.1% APS and 0.1% FSE added; and the 0.15% FSE group was a diet with 0.1% APS and 0.15% FSE added. The specific composition of the experimental diets in each group is shown in Table 1.

[0050] Table 1 Composition of experimental diets in different groups

[0051]

[0052]

[0053] The multivitamins used in the feeds are 250000 IU / kg vitamin A, 750 mg / kg riboflavin, 400 mg / kg pyridoxine-hydrochloride, 1 mg / kg vitamin B12, 250 mg / kg ammonium sulfate, 250 mg / kg vitamin K, 125 mg / kg folic acid, 10 mg / kg biotin, 2.5 mg / kg vitamin E, 8000 mg / kg inositol, 1250 mg / kg calcium pantothenate, 2000 mg / kg niacin, 8000 mg / kg chloride. Choline, 45000IU / kg vitamin D3, 7000mg / kg vitamin C; complex mineral salts are 0.04g / kg zinc sulfate heptahydrate, 37.9g / kg calcium carbonate, 5.3g / kg potassium chloride, 0.04g / kg potassium iodide, 2.6g / kg sodium chloride, 0.02g / kg copper sulfate pentahydrate, 0.02g / kg cobalt sulfate heptahydrate, 0.9g / kg iron sulfate heptahydrate, 0.03g / kg manganese sulfate monohydrate, 3.5g / kg magnesium sulfate heptahydrate, and 9.8g / kg calcium dihydrogen phosphate dihydrate.

[0054] The above-mentioned dry ingredients were crushed and filtered with a 200 μm sieve, and the sieve was collected; then they were weighed and mixed according to the formula of each group to obtain a mixture; then fish oil, soybean lecithin and water were added to the mixture in turn, and stirred vigorously to mix evenly; then the obtained mixture was made into an expanded feed with a diameter of 2 mm, and dried in a ventilated oven at 40°C until the water content of the feed was reduced to less than 10%. Finally, the experimental feed was stored at -20°C. (two)

[0056] Large yellow croaker juveniles were obtained from a commercial seedling farm (Ningde Lingling Fishery Technology Co., Ltd.) in Ningde, Fujian Province, China, and placed in seawater floating cages with a size of 4.0 m × 3.0 m × 2.5 m for a two-week adaptation period in order to acclimate the fish to the experimental conditions. Before starting the feeding experiment, all fish were fasted for 24 h. Subsequently, 1200 large yellow croaker juveniles with an initial average body weight of 13.55 ± 0.56 g were selected and randomly distributed in 20 smaller floating cages with a size of 1 m × 1 m × 1.5 m, with 60 large yellow croaker juveniles in each cage. The experimental feed type of each group was then randomly assigned to four of the cages. Feeding was conducted twice a day at 7 am and 6 pm during the 8-week experiment. During this period, the water temperature was maintained within 27 °C to 29 °C, and the dissolved oxygen level was always maintained above 6.0 mg / L.

[0057] After the 8-week feeding experiment, the weight of the juvenile yellow croaker in different treatment groups was measured, and the weight gain ratio (WGR), specific growth rate (SGR), feed efficiency (FE) and survival rate (SR) were calculated. The specific results are shown in Table 2. The data are expressed as mean and standard deviation.

[0058] Weight gain rate (%) = (final weight - initial weight) / initial weight × 100;

[0059] Specific growth rate (%) = (ln final weight - ln initial weight) / experimental days × 100;

[0060] Feed efficiency = (final weight - initial weight) / feed intake;

[0061] Survival rate (%) = final number of fish / initial number of fish × 100.

[0062] Table 2 Growth performance of juvenile yellow croaker in different treatment groups

[0063]

[0064] According to Table 2, compared with the control group, the weight gain rate, specific growth rate and feed efficiency of the 0.05% FSE group and the 0.1% FSE group were significantly improved (P<0.05). Among them, although the values ​​of the above parameters in the APS diet group and the 0.15% FSE group were higher than those in the control group, these differences did not reach statistical significance (P>0.05). In addition, compared with the control group and the 0.15% FSE group, the survival rate of the APS diet group, the 0.05% FSE group and the 0.1% FSE group was significantly improved (P<0.05). (three)

[0066] After the 8-week feeding trial, the yellow croaker fry in each group was fasted for 24 h and then anesthetized with 100 mg / L eugenol solution. Subsequently, each yellow croaker fry in the cage was individually weighed and counted to measure their growth performance. Ten yellow croaker fry were randomly selected from each cage, and the liver and head kidney tissues were collected and immediately frozen in liquid nitrogen to further evaluate the antioxidant markers and mRNA expression levels. In addition, the intestines of these 10 yellow croaker fry were quickly dissected and frozen in liquid nitrogen for later analysis of digestive enzyme activity. Finally, three yellow croaker fry were randomly selected from each cage to extract midgut samples, which were preserved in 4% paraformaldehyde for intestinal structure histology and morphology analysis.

[0067] 1. Observation of midgut morphology

[0068] The midgut specimens of each group were fixed with 4% paraformaldehyde and then dehydrated in ethanol solutions with increasing concentrations (75% ethanol solution for 4 h, 85% ethanol solution for 2 h, 90% ethanol solution for 2 h, 95% ethanol solution for 1 h, and 100% ethanol solution for 1 h). After dehydration, they were embedded in paraffin and cut using a rotary microtome to obtain 5 μm midgut slices, which were then stained with hematoxylin and eosin. These prepared slices were then examined and photographed using an optical microscope (ECLIPSE Ci, Tokyo, Japan). The length of the villi depicted in the images was quantitatively determined using the ImagePro Plus6.0 software application.

[0069] The results are as follows Figure 1 and Figure 2 As shown, in all experimental groups, the intestinal tissue morphology remained histologically normal without any obvious pathological changes, indicating that the doses of APS and FSE had no toxic or harmful effects. In addition, the villi of the yellow croaker fry in the FSE group were significantly longer than those in the control group and the APS diet group (P<0.05), which also indicated that the combined use of APS and Forsythia extract enhanced intestinal function.

[0070] 2. Detection of antioxidant capacity and digestive enzyme activity

[0071] The intestinal or liver tissue obtained above was taken, and then the intestinal or liver tissue was homogenized in an ice-cold physiological saline solution at a ratio of 10 volumes of physiological saline to 1 weight of tissue to obtain a mixture; the mixture was then rotated in a centrifuge at 4°C at 2000 times per minute for 30 minutes to obtain the intestinal tissue supernatant or liver tissue supernatant, respectively. These supernatants were then used to evaluate the activity of digestive enzymes and antioxidant parameters, respectively. The analysis of antioxidant capacity included the activity of peroxidase (CAT), superoxide dismutase (SOD) and glutathione peroxidase (GSH-PX), as well as the content of malondialdehyde (MDA) in liver samples; digestive enzyme activity analysis was also performed on intestinal samples, and the indicators included trypsin, lipase and α-amylase. The above determinations were all carried out according to the instructions of the kit, and each detection kit was purchased from Nanjing Jiancheng Bioengineering Institute. The determination results are shown in Tables 3 and 4.

[0072] Table 3 Digestive enzyme activities of juvenile yellow croaker in different treatment groups

[0073]

[0074] Table 4 Antioxidant capacity of juvenile yellow croaker in different treatment groups

[0075]

[0076] According to the results in Table 3, the activities of α-amylase and lipase in the APS, 0.05% FSE and 0.1% FSE diets were significantly higher than those in the control and 0.15% FSE diets (P<0.05). It is worth noting that the activities of α-amylase and lipase in juvenile yellow croaker fed with 0.05% FSE and 0.1% FSE diets were significantly enhanced compared with those in the APS diet (P<0.05). In addition, the trypsin activity of juvenile yellow croaker fed with 0.05% FSE and 0.1% FSE diets was significantly higher than that of fish fed with any other diet formula (P<0.05).

[0077] As can be seen from Table 4, compared with the large yellow croaker fry in the control group and the APS diet group, the SOD, CAT and GSH-PX activity levels of the large yellow croaker fry fed with the FSE feed were significantly increased, and the MDA content was significantly decreased (P<0.05).

[0078] 3. Transcription levels of genes related to inflammation, apoptosis and anti-oxidation in the head kidney

[0079] Head kidney samples of juvenile yellow croaker in each cage were taken and RNA was extracted using Trizol reagent (TaKaRa, Dalian, China) according to the instructions. Subsequently, the concentration and purity of the extracted RNA were quantified using Nanodrop 2000 spectrophotometer (Thermo, Wasserhaus, USA) and further confirmed by 1% agarose gel electrophoresis. Primescript TM Total RNA was reverse transcribed into cDNA using a RT assay kit (TaKaRa, Dalian, China) and prepared for qRT-PCR analysis. The qRT-PCR procedure was based on “Dietary Tribonema sp. supplementation increased growth performance, antioxidant capacity, immunity and improved hepatic health in golden pompano (Trachinotus ovatus)” (Zhao, Wei Fang, Hao-Hang Gao, Bao-Yan Dai, Chen-Min Liu, Zhen-Zhou Zhang, Cheng-Wu Niu, Jin. Dietary Tribonema sp. supplementation increased growth performance, antioxidant capacity, immunity and improved hepatic health in golden pompano (Trachinotus ovatus) [J]. Aquaculture, 2020, 529 (1).). The specific primer sequences used for qRT-PCR are shown in Table 5. 2 -ΔΔCt Methods The expression levels of target genes were calculated, and all results were normalized with the expression of the housekeeping gene β-actin to ensure consistency and accuracy.

[0080] Table 5 qRT-PCR specific primer sequences

[0081]

[0082]

[0083] Among the proinflammatory cytokines, the mRNA levels of interleukin-8 (IL-8) and toll-like receptor 1 (TLR1) in juvenile yellow croaker fed with APS, 0.05%, 0.1% FSE, and 0.15% FSE were significantly decreased compared with the control group (P<0.05). Similarly, the transcription levels of myeloid differentiation factor 88 (MYD88) and toll-like receptor 5A (TLR5A) in juvenile yellow croaker fed with FSE were significantly inhibited compared with the control group and APS group (P<0.05). Among them, the expression level of toll-like receptor 2A (TLR2α) in the APS, 0.05%, and 0.1% FSE groups was significantly downregulated compared with the control group and 0.15% FSE group (P<0.05). In contrast, there was no significant change in the mRNA level of tumor necrosis factor-α (TNF-α) among the different treatment groups (P>0.05) (e.g. Figure 3 shown).

[0084] according to Figure 3 Analysis of anti-inflammatory cytokines also showed that the expression level of interleukin 10 (IL-10) in the APS diet group and the FSE-supplemented diet was significantly increased compared with the control group (P<0.05). In addition, the expression of transforming growth factor-β (TGF-β) in the juvenile yellow croaker fed the 0.1% FSE diet was significantly increased compared with the fish fed other diets (P<0.05).

[0085] like Figure 4 As shown in the results, compared with the control group, the transcriptional levels of anti-apoptotic factors, especially B-cell lymphoma-2 (bcl2), were significantly upregulated in the juvenile yellow croaker fed with the APS diet and the FSE-supplemented diet group (P<0.05). In contrast, compared with the control group, the expression of pro-apoptotic factors, including Bcl2-associated X protein (bax), caspase 3, caspase 8, and caspase 9, were significantly downregulated in the juvenile yellow croaker fed with the APS diet and the FSE-supplemented diet group (P<0.05).

[0086] The expression of antioxidant-related genes can be analyzed (e.g. Figure 4 ), compared with the control group and APS diet group, the transcription levels of nuclear factor E2-related factor 2 (nrf2), sod and cat in large yellow croaker fry fed with FSE-supplemented diet were significantly upregulated (P<0.05).

[0087] In summary, it can be seen that when APS is used alone, it has a certain promoting effect on the growth of yellow croaker fry. When APS is combined with FSE, a synergistic effect is achieved, and the growth promotion effect is further improved. When APS is used in combination with 0.05% or 0.1% FSE, the intestinal health of yellow croaker fry is significantly improved and the activity of digestive enzymes is increased, making the growth rate and feed efficiency relatively the best. In addition, the synergistic application of APS and FSE also better activates the Nrf2 pathway and increases the level of antioxidant enzymes, further enhancing the antioxidant defense capacity of yellow croaker fry. APS and FSE, used alone or in combination, can achieve the effect of reducing inflammation-induced apoptosis and improving the health of yellow croaker fry. At the same time, they also show anti-inflammatory effects by inhibiting TLR / MyD88. However, considering the results of growth and physiological indicators, the combined use of 0.1% APS and 0.05% FSE added to the feed to cultivate yellow croaker fry has the best effect on improving their growth rate, survival rate and resilience to stress factors.

[0088] The above is only a preferred embodiment of the present invention. It should be pointed out that for ordinary technicians in this technical field, several improvements and modifications can be made without departing from the principle of the present invention. These improvements and modifications should also be regarded as the scope of protection of the present invention.

Claims

1. A composition containing astragalus polysaccharide and forsythia suspensa extract, characterized in that: The mass ratio of the astragalus polysaccharide to the forsythia extract is 10:(2-15).

2. The composition according to claim 1, characterized in that The Forsythia suspensa extract comprises forsythiaside A, forsythiaside and forsythiaside.

3. Use of the composition according to claim 1 or 2 in preparing feed with antioxidant effect.

4. Use of the composition according to claim 1 or 2 in preparing feed having the effect of alleviating inflammation.

5. Use of the composition according to claim 1 or 2 in preparing feed having the effect of improving growth rate and survival rate.

6. A feed for juvenile large yellow croaker, characterized in that: The invention comprises the following components in percentage by weight: 38-42% fish meal, 27-32% soybean meal, 20-23% flour, 2-3% fish oil, 0.8-1.2% soybean lecithin, 2-3% brewer's yeast, 0.7-1.2% complex vitamins, 0.8-1.2% complex mineral salts and 0.1-0.2% of the composition according to claim 1 or 2.

7. Use of the large yellow croaker fry feed according to claim 6 in improving the growth rate and survival rate of large yellow croaker fry, alleviating inflammation of large yellow croaker fry, and enhancing the antioxidant capacity of large yellow croaker fry.

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