Application of alkaline phosphatase in preparation of aquatic animal feed or medicine for preventing or reducing liver injury of aquatic animals

Abstract

The invention relates to application of alkaline phosphatase in preparation of aquatic animal feed or medicine for preventing or reducing liver injury of aquatic animals. In particular to application of alkaline phosphatase in preparation of aquatic animal feed or medicine for preventing or reducing liver injury of aquatic animals, the aquatic animal feed, a preparation method of the aquatic animal feed and a feeding method of the aquatic animals. According to the aquatic animal feed disclosed by the invention, the basic feed with relatively low fish meal is compounded with the alkaline phosphatase to prepare the aquatic animal feed, so that the oxidation resistance of livers of aquatic animals, especially largemouth bass, is effectively improved, the content of AST in serum of the aquatic animals is reduced, and the contents of ALP and ALB in the serum of the aquatic animals are improved, thereby improving the survival rate of the aquatic animals.

Description

Technical Field

[0001] The present disclosure relates to the field of feed technology, and in particular, to the use of alkaline phosphatase in preparing aquatic animal feed or medicine for preventing or reducing liver damage in aquatic animals, an aquatic animal feed, a method for preparing an aquatic animal feed, and a method for raising aquatic animals. Background Art

[0002] Fishmeal is the core raw material in the production of aquatic animal feed. It has high protein content, rich in essential amino acids, vitamins, minerals and balanced composition, high biological value, high effective energy value, good palatability and few anti-nutritional factors. However, with the intensive and large-scale development of aquatic animal farming, protein feed resources represented by fishmeal are becoming increasingly scarce worldwide. The imbalance between supply and demand has led to a continuous increase in fishmeal prices and breeding costs. Low-fishmeal feed has become a new development trend. However, fishmeal substitution can easily have a negative impact on the liver and intestinal health of aquatic animals, thereby reducing the survival rate of aquatic animals and affecting breeding benefits.

[0003] Largemouth bass ( Micropterus salmoides ), also known as California bass, is a freshwater fish that grows fast, has strong disease resistance, is highly adaptable to temperature, has delicious meat and high nutritional value, and is widely loved by breeders and consumers. With the promotion of intensive and large-scale aquaculture models in the aquaculture industry, largemouth bass grows slowly and suffers from frequent diseases during high-density aquaculture. In addition, there are restrictions on antibiotic additives in aquatic animal feed. Therefore, it is urgent to seek green, efficient and safe aquatic animal feed to protect the health of aquatic animals and improve their survival rate. Summary of the invention

[0004] In order to solve the above technical problems, the purpose of the present disclosure is to provide the use of alkaline phosphatase in the preparation of aquatic animal feed or medicine for preventing or reducing liver damage in aquatic animals, an aquatic animal feed, a method for preparing an aquatic animal feed, and a method for raising aquatic animals.

[0005] In order to achieve the above objectives, the present disclosure provides, in a first aspect, the use of alkaline phosphatase in the preparation of aquatic animal feed or medicine for preventing or reducing liver damage in aquatic animals.

[0006] Optionally, the aquatic animal feed or medicine improves the survival rate of aquatic animals.

[0007] Optionally, the aquatic animal feed or medicine reduces the content of AST in the serum of aquatic animals and increases the content of ALP and ALB in the serum of aquatic animals.

[0008] Optionally, the aquatic animal feed or medicine enhances the antioxidant capacity of the liver of aquatic animals; Preferably, the method of improving the antioxidant capacity of aquatic animal liver comprises: (1) Increase the activity of CAT and SOD in liver tissue of aquatic animals; (2) Reduce the MDA content in the liver tissue of aquatic animals.

[0009] Optionally, the aquatic animal is selected from one or more of fish, shrimp and crab; preferably, the fish is largemouth bass.

[0010] A second aspect of the present disclosure provides an aquatic animal feed, comprising a low-fish meal base feed and alkaline phosphatase.

[0011] Optionally, the amount of alkaline phosphatase added is 300-500 mg per 1 kg of the low fish meal base feed; Wherein, based on the total weight of the low fish meal basic feed, the low fish meal basic feed contains 25-35% by weight of fish meal.

[0012] A third aspect of the present disclosure provides a method for preparing aquatic animal feed, the method comprising the following steps: S1, performing a first mixing process on the solid raw materials in the low fish meal basic feed and water to obtain a first mixed feed; S2, subjecting the first mixed material to an expanded granulation process to obtain first granules; S3, performing a second mixing treatment on the oil phase raw material in the low fish meal base feed and alkaline phosphatase to obtain a second mixed feed; S4, spraying the second mixed material on the surface of the first particles and then drying them.

[0013] A fourth aspect of the present disclosure provides a method for raising aquatic animals, wherein the aquatic animals are raised in a breeding area and the aquatic animal feed described in the second aspect is fed to the aquatic animals; The feeding amount of the aquatic animal feed is 20-35 g / d relative to 1 kg of the aquatic animal; Among them, the breeding conditions in the breeding area include: water temperature of 27-32°C, pH value of 7.5-8.0, ammonia nitrogen content <0.2mg / L, nitrite content <0.01mg / L, and dissolved oxygen concentration ≥7.0mg / L.

[0014] Optionally, the content of alkaline phosphatase fed is 6.0-17.5 mg / d per 1 kg of the aquatic animal.

[0015] Through the above technical scheme, the present invention adopts a relatively low fish meal-based feed compounded with alkaline phosphatase to prepare aquatic animal feed, which effectively improves the antioxidant capacity of the liver of aquatic animals, especially largemouth black bass, reduces the AST content in the serum of aquatic animals, and increases the ALP and ALB contents in the serum of aquatic animals, thereby improving the survival rate of aquatic animals; at the same time, the use of low fish meal-based feed reduces the preparation cost of aquatic animal feed.

[0016] Other features and advantages of the present disclosure will be described in detail in the following detailed description. BRIEF DESCRIPTION OF THE DRAWINGS

[0017] The accompanying drawings are used to provide a further understanding of the present disclosure and constitute a part of the specification. Together with the following specific embodiments, they are used to explain the present disclosure but do not constitute a limitation of the present disclosure. In the accompanying drawings: Figure 1 1 is a staining image of liver tissue sections of largemouth bass from Group A, Group B and Group C of the present disclosure.

[0018] Figure 2 yes Figure 1 Enlarged view of the black box in the middle.

[0019] Figure 3 It is a staining picture of the liver tissue sections of the largemouth bass in Group D, Group E and Group F of the present disclosure.

[0020] Figure 4 yes Figure 3 Enlarged view of the black box in the middle. DETAILED DESCRIPTION

[0021] The specific implementation of the present disclosure is described in detail below in conjunction with the accompanying drawings. It should be understood that the specific implementation described herein is only used to illustrate and explain the present disclosure, and is not used to limit the present disclosure.

[0022] A first aspect of the present disclosure provides use of alkaline phosphatase in preparing aquatic animal feed or medicine for preventing or reducing liver damage in aquatic animals.

[0023] The inventors of the present disclosure have found that alkaline phosphatase can prevent or reduce liver damage in aquatic animals, thereby improving the survival rate of aquatic animals; in particular, it can prevent or reduce the occurrence of largemouth bass ( Micropterus salmoides Therefore, alkaline phosphatase has a good application prospect in the preparation of aquatic animal feed or medicine.

[0024] According to the present disclosure, the aquatic animal feed or medicine can improve the survival rate of aquatic animals.

[0025] In the present disclosure, aspartate aminotransferase (AST), alkaline phosphatase (ALP) and albumin (ALB) are important indicators for evaluating liver damage; when liver cells are damaged, AST will be released from liver cells into the blood, resulting in an increase in the AST content in serum; when the liver is damaged, the synthesis and secretion of ALP may be affected, resulting in a decrease in its content; ALB is mainly synthesized by the liver, and the ability of the liver to synthesize ALB decreases when the liver is damaged. In one embodiment of the present disclosure, aquatic animal feed or medicine can reduce the AST content in aquatic animal serum and increase the ALP and ALB content in aquatic animal serum. Thereby alleviating liver damage and protecting the liver.

[0026] In the present disclosure, the aquatic animal feed or medicine can enhance the antioxidant capacity of the liver of aquatic animals; specifically, the enhancement of the antioxidant capacity of the liver of aquatic animals includes: (1) enhancing the activity of CAT and SOD in the liver tissue of aquatic animals; (2) reducing the content of MDA in the liver tissue of aquatic animals.

[0027] Among them, increasing the activity of catalase (CAT) can eliminate hydrogen peroxide activity and protect cells from oxidative damage. Increasing the activity of superoxide dismutase (SOD) can enhance the antioxidant capacity of liver cells, reduce the damage of reactive oxygen to liver cells, promote the proliferation and regeneration of liver cells, and accelerate the repair of liver damage. Malondialdehyde (MDA) is a product of lipid peroxidation. Reducing the content of MDA can reduce lipid peroxidation, reduce cell membrane damage, and help repair and restore liver cells.

[0028] In one embodiment of the present disclosure, the aquatic animal can be selected from one or more of fish, shrimp and crab; preferably, the fish can be largemouth bass ( Micropterus salmoides ).

[0029] A second aspect of the present disclosure provides an aquatic animal feed, comprising a low-fish meal base feed and alkaline phosphatase.

[0030] In one embodiment of the present disclosure, the amount of alkaline phosphatase added is 300-500 mg per 1 kg of the low fish meal base feed; Wherein, based on the total weight of the low fish meal basic feed, the low fish meal basic feed contains 25-35% by weight of fish meal.

[0031] A third aspect of the present disclosure provides a method for preparing aquatic animal feed, the method comprising the following steps: S1, performing a first mixing process on the solid raw materials in the low fish meal basic feed and water to obtain a first mixed feed; S2, subjecting the first mixed material to an expanded granulation process to obtain first granules; S3, performing a second mixing treatment on the oil phase raw material in the low fish meal base feed and alkaline phosphatase to obtain a second mixed feed; S4, spraying the second mixed material on the surface of the first particles and then drying them.

[0032] In the present disclosure, the low fish meal basic feed includes: fish meal, chicken meal, soybean protein concentrate, soybean meal, gluten powder, pig blood cell powder, wheat flour, tapioca starch, fish oil, soybean oil, monocalcium phosphate, vitamin and mineral premix, choline chloride, VC phosphate, lysine (98.5%) and methionine.

[0033] Among them, vitamins can include: vitamin A; vitamin D3; vitamin E; vitamin K; thiamine; riboflavin; calcium pantothenate; pyridoxine hydrochloride; cyanocobalamin; biotin; folic acid; niacin; inositol; vitamin C.

[0034] Minerals may include: magnesium sulfate; ferrous sulfate; zinc sulfate; copper sulfate; manganese sulfate; carriers, with rice husks and zeolite as carriers.

[0035] Among them, fish oil and soybean oil are used as oil phase raw materials, and the rest are solid phase raw materials.

[0036] More specifically, in step S1, each solid raw material is subjected to ultrafine grinding and screening (80 mesh) in sequence, and then the ground solid raw material is subjected to a first mixing process with water to obtain a first mixed material. The first mixing process can be performed using a mixer conventionally used by those skilled in the art. The ultrafine grinding and screening processes are both performed using methods conventionally used by those skilled in the art, and will not be described in detail herein. Among them, the solid raw material and water can be subjected to a first mixing process at a mass ratio of 1:0.2.

[0037] Furthermore, in step S2, the expanded mass granulation process may adopt a dry extruder conventionally used by those skilled in the art.

[0038] Furthermore, in step S3, the second mixing treatment may be performed using a micro mixer to fully dissolve the alkaline phosphatase in the oil phase raw material.

[0039] Furthermore, in step S4, the spraying may be performed by using a grease spraying device. The drying process may be performed by natural air drying.

[0040] The fourth aspect of the present disclosure provides a method for raising aquatic animals, wherein the aquatic animals are raised in a breeding area and the aquatic animal feed described in the second aspect is fed to the aquatic animals; The feeding amount of the aquatic animal feed is 20-35 g / d relative to 1 kg of the aquatic animal; Among them, the breeding conditions in the breeding area include: water temperature of 27-32°C, pH value of 7.5-8.0, ammonia nitrogen content <0.2mg / L, nitrite content <0.01mg / L, and dissolved oxygen concentration ≥7.0mg / L.

[0041] In one embodiment of the present disclosure, the content of alkaline phosphatase fed is 6-17.5 mg / d per 1 kg of the aquatic animal.

[0042] The present disclosure is further described in detail below through examples, but the present disclosure is not limited thereby.

[0043] The materials and reagents used in the following examples can all be obtained through commercial sources.

[0044] Example 1 1. Black bass seedlings: Largemouth bass fingerlings were provided by the Freshwater Fisheries Research Center, Chinese Academy of Fishery Sciences.

[0045] A total of 360 largemouth bass juveniles of similar size and without external damage were selected, with an average initial body weight of 16.36±0.03g. They were fed twice a day at regular times (7:00 and 17:00) to show apparent satiation.

[0046] During the breeding experiment, the water quality was measured once a week to ensure the water quality standard. At the same time, the water temperature in the breeding area was controlled at 27-32°C, pH value 7.5-8.0, ammonia nitrogen content <0.2 mg / L, nitrite content <0.01 mg / L and dissolved oxygen concentration ≥7.0 mg / L. The light cycle was the natural light cycle.

[0047] 2. Feed composition: The compositions of low fish meal based feed formula 1 and formula 2 are shown in Table 1 below.

[0048] Table 1

[0049] According to the formula in Table 1, the preparation method of aquatic animal feed is as follows: S1. Superfinely grind and sieve each solid raw material in turn (80 meshes), and then mix the ground solid raw materials with water to obtain a first mixed material. Mix the solid raw materials in the low fish meal base feed with water at a mass ratio of 1:0.2 to obtain a first mixed material; S2, subjecting the first mixed material to an expanded granulation process to obtain first granules; S3, performing a second mixing treatment on the oil phase raw material in the low fish meal base feed and alkaline phosphatase to obtain a second mixed feed; S4, spraying the second mixed material on the surface of the first particles and then drying them naturally.

[0050] 3. Feeding method: 360 largemouth bass juveniles of similar size and without external damage were cultured in outdoor cages. They were randomly placed in 18 cages (1m×1m×1m), with 20 fish in each cage and 3 cages in each group. The culture period was 8 weeks. The experiment was divided into 6 groups, as follows: Group A: fed with low fish meal-based diet formula 20 g / kg·d (alkaline phosphatase 0 mg / kg·d); Group B: fed with aquatic animal feed formula 1, group 1, 20 g / kg·d (alkaline phosphatase 6 mg / kg·d), where aquatic animal feed formula 1, group 1, is low fish meal basic feed formula 1, with 300 mg alkaline phosphatase added per 1 kg; Group C: fed with aquatic animal feed formula 1 group 2 20g / kg·d (alkaline phosphatase 10mg / kg·d), where aquatic animal feed formula 1 group 2 is low fish meal basic feed formula 1 with 500mg alkaline phosphatase added per 1kg; Group A served as the control group for groups B and C.

[0051] Group D: fed with low fish meal based feed formula II 20 g / kg·d (alkaline phosphatase 0 mg / kg·d); Group E: fed with aquatic animal feed formula 2, group 1, 20 g / kg·d (alkaline phosphatase 6 mg / kg·d), where aquatic animal feed formula 2, group 1, is low fish meal-based feed formula 2 with 300 mg alkaline phosphatase added per 1 kg; Group F: fed with aquatic animal feed formula 2, group 2 20g / kg·d (alkaline phosphatase 10mg / kg·d), wherein the aquatic animal feed formula 2, group 2 is low fish meal based feed formula 2 with 500mg alkaline phosphatase added for every 1kg.

[0052] Group D served as the control group for groups E and F.

[0053] The alkaline phosphatase used in the experiment was provided by Beijing Auxit Biotechnology Co., Ltd. Its preparation method is to breed the production bacteria (Bacillus licheniformis) through seeds, and then carry out liquid deep fermentation with glucose, soybean meal and corn pulp as the main raw materials, and the obtained fermentation liquid is subjected to plate and frame filtration and ultrafiltration purification, concentrated, and added with corn starch and other carriers and spray-dried; the enzyme activity of alkaline phosphatase is 5000U / g.

[0054] 4. Data processing: SPSS 27.0 software was used for one-way ANOVA test for statistical analysis. The results were expressed as mean ± standard deviation, and P < 0.05 was considered significant.

[0055] Test Example 1 This test case was used to illustrate the effect of alkaline phosphatase on the survival and growth performance of largemouth bass.

[0056] Observe the fish feeding and mortality every day. If dead fish are found, remove them immediately, weigh them, and count them, and check the cause of death.

[0057] After 8 weeks of formal experimental feeding, the fish were stopped from feeding for 24 h, and the total weight of fish in each culture cage was weighed to calculate growth performance indicators such as weight gain rate, specific growth rate, survival rate and feed coefficient.

[0058] Weight gain rate (%) = (W 末 -W0) / W0×100%; Specific growth rate (%) = (InW 末 -InW0) / D×100%; Survival rate (%) = (N0-N 末 ) / N0×100%; Bait coefficient = M 料 / (W 末 -W0)×100%; Among them, W 末 : final body weight of the experiment, W0: initial body weight of the experiment, D: experimental time (d), N0: initial number of the experiment, N 末 : The number at the end of the experiment, M 料 : Total feed consumed during the experimental period.

[0059] The experimental results are shown in Table 2.

[0060] Table 2

[0061] The results in Table 2 show that the addition of alkaline phosphatase had no significant effect on the growth performance of largemouth bass, but the addition of 500 mg / kg alkaline phosphatase in groups C and F significantly increased the survival rate of largemouth bass ( P <0.05).

[0062] Test Example 2 This test example is used to illustrate the effect of alkaline phosphatase on serum biochemical parameters of largemouth bass.

[0063] At the end of the experiment, three fish were randomly selected from each treatment group for blood collection. The fish were quickly picked up and immediately put into MS-222 with a concentration of 200 mg / L for rapid deep anesthesia. The wall of the 2mL syringe was moistened with heparin, and blood was collected from the tail vein. The blood collection time for each treatment was completed within 2 hours, and the blood collection time was accurately recorded. After the blood sample was placed in a 4°C refrigerator for 1-2 hours, plasma was prepared at 4°C, 3000×g, and centrifuged for 10 minutes. The supernatant was transferred to -20°C for storage and prepared for the measurement of routine blood biochemical indicators of largemouth bass, such as glutamate aminotransferase (ALT), aspartate aminotransferase (AST), alkaline phosphatase (ALP), total protein (TP), albumin (ALB), and blood glucose (GLU).

[0064] The test method was determined using Mindray BS400 fully automatic blood biochemical analyzer, and the kit was purchased from Shanghai Zhicheng Biotechnology Co., Ltd.

[0065] The experimental results are shown in Table 3.

[0066] Table 3

[0067] As shown in Table 3, there was no significant difference in ALT and TP levels among the groups ( P >0.05). With the increase of alkaline phosphatase addition, AST levels showed a downward trend, with groups B, C, and F showing lower levels ( P <0.05); With the increase of the amount of alkaline phosphatase added, the ALP level showed an increasing trend, among which group B, group C, and group F showed higher levels ( P <0.05); the ALB content in group B and group C increased significantly ( P <0.05); GLU content in group C and group F was significantly reduced ( P <0.05).

[0068] Test Example 3 This test example was used to illustrate the effect of alkaline phosphatase on the antioxidant capacity of largemouth bass liver.

[0069] Three fish were randomly selected from each treatment group to collect liver tissue for the determination of liver tissue antioxidant indices, such as glutathione peroxidase (GSH-Px), catalase (CAT), malondialdehyde (MDA), superoxide dismutase (SOD), and glutathione (GSH).

[0070] The test method uses the test kit of Nanjing Jiancheng Bioengineering Institute for determination. The specific determination method refers to the kit instructions.

[0071] The results are shown in Table 4.

[0072] Table 4

[0073] As shown in Table 4, compared with groups A and D, the CAT activity of groups B and E was significantly increased ( P <0.05); SOD activity increased with the increase of alkaline phosphatase addition; with the increase of alkaline phosphatase addition, MDA content basically showed a decreasing trend, with the lowest levels in groups C and F ( P <0.05); there were no significant differences in GSH content and GSH-Px activity among the groups ( P >0.05).

[0074] Test Example 4 This test case was used to illustrate the effect of alkaline phosphatase on the histomorphology of liver of largemouth bass.

[0075] After the experiment, liver tissues of 2 fish were randomly selected for each treatment and immediately preserved in 4% paraformaldehyde solution. After fixation for 24 h, the liver tissues were dehydrated, embedded, sliced, stained and sealed. The slices were photographed and scanned, and the intestinal tissue morphology was analyzed using CaseViewer 2.4 software for pathological analysis of the intestinal tissue.

[0076] The results of liver tissue morphology of black sea bass in each group are as follows Figure 1 and 2 As shown, the yellow arrow indicates cytoplasmic vacuolation; the purple arrow indicates inflammatory cell infiltration; and the red arrow indicates vascular congestion.

[0077] The surface of liver tissue has a very thin serous membrane, which is composed of a single layer of squamous epithelium and very thin connective tissue. The liver cells are arranged in two rows in a plate-like shape. The liver plates are scattered around the central vein. The adjacent liver cell cords are irregular and interconnected into a network of hepatic sinusoids. The stem cell cytoplasm of each group was vacuolated; Figure 1 , Figure 2 It can be seen that small focal infiltration of lymphocytes around veins and in the parenchyma of the hepatocytes in group A, while no obvious infiltration of inflammatory cells was observed in groups B and C, indicating that the addition of 300 mg / kg or 500 mg / kg of fish meal to the feed in the low fish meal based feed formula 1 can alleviate liver damage.

[0078] Depend on Figure 3 , Figure 4 It can be seen that small focal infiltrations of lymphocytes around veins and in the parenchyma of the hepatocytes in group D, while no obvious infiltration of inflammatory cells was observed in groups E and F, indicating that the addition of 300 mg / kg or 500 mg / kg of fish meal to the low fish meal based feed formula 2 can alleviate liver damage.

[0079] The preferred embodiments of the present disclosure are described in detail above in conjunction with the accompanying drawings; however, the present disclosure is not limited to the specific details in the above embodiments. Within the technical concept of the present disclosure, a variety of simple modifications can be made to the technical solution of the present disclosure, and these simple modifications all fall within the protection scope of the present disclosure.

[0080] It should also be noted that the various specific technical features described in the above specific embodiments can be combined in any suitable manner without contradiction. In order to avoid unnecessary repetition, the present disclosure will not further describe various possible combinations.

[0081] In addition, various embodiments of the present disclosure may be arbitrarily combined, and as long as they do not violate the concept of the present disclosure, they should also be regarded as the contents disclosed by the present disclosure.

Claims

1. Application of alkaline phosphatase in the preparation of aquatic animal feed or medicine for preventing or reducing liver damage in aquatic animals.

2. The use according to claim 1, wherein: The aquatic animal feed or medicine improves the survival rate of aquatic animals.

3. The use according to claim 2, wherein: The aquatic animal feed or medicine reduces the content of AST in the serum of aquatic animals and increases the content of ALP and ALB in the serum of aquatic animals.

4. The use according to claim 2, wherein: The aquatic animal feed or medicine improves the antioxidant capacity of the liver of aquatic animals; Preferably, the method of improving the antioxidant capacity of aquatic animal liver comprises: (1) Increase the activity of CAT and SOD in liver tissue of aquatic animals; (2) Reduce the MDA content in the liver tissue of aquatic animals.

5. The use according to any one of claims 1 to 4, wherein: The aquatic animal is selected from one or more of fish, shrimp and crab; Preferably, the fish is largemouth bass.

6. A feed for aquatic animals, characterized in that: The aquatic animal feed comprises a low fish meal base feed and alkaline phosphatase.

7. The aquatic animal feed according to claim 6, wherein: The amount of alkaline phosphatase added is 300-500 mg per 1 kg of the low fish meal basic feed; Wherein, based on the total weight of the low fish meal basic feed, the low fish meal basic feed contains 25-35% by weight of fish meal.

8. A method for preparing aquatic animal feed, characterized in that: The method comprises the following steps: S1, performing a first mixing process on the solid raw materials in the low fish meal basic feed and water to obtain a first mixed feed; S2, subjecting the first mixed material to an expanded granulation process to obtain first granules; S3, performing a second mixing treatment on the oil phase raw material in the low fish meal base feed and alkaline phosphatase to obtain a second mixed feed; S4, spraying the second mixed material on the surface of the first particles and then drying them.

9. A method for raising aquatic animals, characterized in that: Raising aquatic animals in a breeding area and feeding the aquatic animal feed according to claim 6 or 7 to the aquatic animals; The feeding amount of the aquatic animal feed is 20-35 g / d relative to 1 kg of the aquatic animal; Among them, the breeding conditions in the breeding area include: water temperature of 27-32°C, pH value of 7.5-8.0, ammonia nitrogen content <0.2mg / L, nitrite content <0.01mg / L, and dissolved oxygen concentration ≥7.0mg / L.

10. The breeding method according to claim 9, wherein: The content of alkaline phosphatase fed is 6.0-17.5 mg / d per 1 kg of the aquatic animal.

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