Soluble powder for treating livestock liver injury induced by glyphosate and preparation method thereof

Abstract

The invention belongs to a medicinal preparation characterized by form, and particularly relates to soluble powder for treating livestock liver injury induced by glyphosate and a preparation method thereof. The soluble powder comprises cane sugar, a liver protection compound and a detoxification degradation agent, and the liver protection compound is obtained by performing deodorization treatment on fresh spirulina and modifying the spirulina with epigallocatechin gallate; the detoxication degradation agent is obtained by modifying laccase with rosmarinic acid and further encapsulating an obtained compound with chitosan and trehalose, and the liver protection compound and the detoxication degradation agent can play a role in a synergistic manner. The soluble powder disclosed by the invention is suitable for being prepared on site before livestock eat forage / feed, and after the soluble powder is added into drinking water, livestock liver injury induced by glyphosate and degradation products thereof can be relieved, the function of a liver anti-oxidation system can be maintained as far as possible, and oxidative stress and the like caused by glyphosate can be effectively resisted.

Description

Technical Field

[0001] The present invention belongs to pharmaceutical preparations characterized by form, and particularly relates to a soluble powder for treating glyphosate-induced livestock liver injury and a preparation method thereof. Background Art

[0002] The chemical name of glyphosate is N-(phosphonomethyl)glycine, with the chemical formula C3H8NO5P, a relative molecular mass of 169.07. The pure product is a white solid, with a solubility in water of 1.2% at 25°C and low solubility in organic solvents. Glyphosate is more soluble when present in the form of salts, such as glyphosate isopropylamine salt, glyphosate sodium salt, glyphosate potassium salt, etc. As a systemic and conductive broad-spectrum non-selective herbicide, glyphosate can kill common weeds in agricultural and animal husbandry production. The systemic conductivity of glyphosate is extremely strong. After application, it can be rapidly absorbed by the roots, stems and leaves of plants, migrate and accumulate in metabolically active parts, and interfere with protein synthesis and plant metabolism by inhibiting the synthesis of 5-enolpyruvylshikimate-3-phosphate, preventing shikimic acid from being converted into aromatic amino acids (such as phenylalanine, tryptophan and tyrosine), ultimately leading to plant death. Due to the better herbicidal performance of glyphosate, it has not been completely banned yet, but its toxicity is inevitable. Its oral LD 50 for rats is 5000 mg / kg, and the oral LD 50 for mice is 10000 mg / kg, the LD 50 for goats is 3530 mg / kg, and the acute dermal LD 50 of rabbits > 2000 mg / kg.

[0003] Existing studies have paid little attention to the potential safety and health risks of livestock that use forage as their main source of nutrition after eating forage sprayed with glyphosate. Some evidence shows that the risks of this process may be underestimated. Based on the glyphosate residues in forage and the forage consumption of livestock, it is inferred that the concentration of glyphosate in pig intestine is about 0.005~0.02mg / mL, and the concentration of glyphosate in cattle intestine is about 0.002~0.003mg / mL. When further considering the actual situation, such as the fact that the forage fed to livestock is generally not cleaned, the actual forage consumption changes with the growth of livestock, the health status of the livestock themselves, and the supply of fresh forage in season, livestock may ingest more glyphosate during their growth and face the harm caused by excessive glyphosate. Most of the toxic substances in forage are metabolized by the liver, which transforms them through oxidation, reduction and hydrolysis, making the liver one of the main target organs for exogenous toxic substances. When livestock ingest too much glyphosate, it may induce liver damage. In addition, livestock feed prepared from forage will also bring the above-mentioned problems. Due to the complex source of forage, improper harvesting management, rough production process, etc., livestock face the problem of glyphosate-induced liver damage.

[0004] Liver damage in livestock may lead to decreased immune function, metabolic dysfunction, malnutrition, etc., which seriously affects the life cycle of livestock, and further affects the production benefits and economic benefits obtained from livestock breeding. Therefore, it is necessary to prepare a drug for treating glyphosate-induced liver damage in livestock. Summary of the invention

[0005] Based on the above problems, the present invention provides a soluble powder for treating glyphosate-induced liver damage in livestock. The soluble powder contains sucrose, a liver protection complex, and a detoxification and degradation agent. The liver protection complex is obtained by removing the fishy smell from fresh spirulina and then modifying it with epigallocatechin gallate; the detoxification and degradation agent is obtained by modifying laccase with rosmarinic acid, and the obtained complex is further encapsulated with chitosan and trehalose; and sucrose plays a role in attracting food. The liver protection complex and the detoxification and degradation agent can work synergistically. The soluble powder is prepared on the spot and added to drinking water to reduce glyphosate-induced liver damage in livestock, maintain the function of the liver's antioxidant system as much as possible, and effectively resist the oxidative stress caused by glyphosate.

[0006] The invention provides a soluble powder for treating glyphosate-induced liver damage in livestock, comprising the following raw materials in parts by weight: 0.5-1.5 parts of sucrose, 2-5 parts of a liver protection complex, and 3-5 parts of a detoxification and degradation agent.

[0007] The preparation method of the liver protection complex is as follows: L1. After washing fresh spirulina with water, blanch it in water at 80 °C for 1 - 2 min, then air-dry the surface moisture to obtain a spirulina sample. Crush ginger charcoal and sieve it to obtain ginger charcoal powder with a particle size of 80 - 100 μm. Disperse the ginger charcoal powder in water to obtain a ginger charcoal suspension with a mass percentage concentration of 4 - 8%. Immerse all of the spirulina sample in the ginger charcoal suspension and let it stand at room temperature for 2 - 3 h. After that, wash it and grind it to obtain spirulina powder with a particle size of 10 - 20 μm after complete drying; L2. Weigh the spirulina powder, xanthan gum aqueous solution, and epigallocatechin gallate obtained in step L1 according to a mass ratio of 8 - 10:10 - 15:1. After mixing the spirulina powder and the xanthan gum aqueous solution, ultrasonically disperse them until uniform, then add epigallocatechin gallate, adjust the pH to 7 - 8, heat to 30 - 40 °C and maintain for 4 - 8 h to obtain Solution 1; L3. Dialyze the Solution 1 obtained in step L2 at 4 °C for 10 - 12 h. Select a dialysis bag with a cut-off molecular weight of 9 - 12 kDa for dialysis. After that, vacuum freeze-dry the substances in the dialysis bag to obtain the liver protection complex.

[0008] Preferably, the mass percentage concentration of the xanthan gum aqueous solution in step L2 is 3 - 5%.

[0009] Preferably, the dialysis medium is changed 2 - 3 times during dialysis in step L3. The dialysis medium is deionized water. Vacuum freeze-drying is preferably carried out with pre-cooling at -60 °C, a baffle temperature of -50 °C, and a vacuum degree of 5 Pa.

[0010] The preparation method of the detoxifying and degrading agent is as follows: S1. Weigh rosmarinic acid and sodium hydroxide solution according to a mass-to-volume ratio of 40 - 50 mg:6 - 9 mL, stir until evenly dispersed, and adjust the pH to 7 - 7.5 to obtain Solution 2; S2. Weigh EDC, NHS, and the Solution 2 obtained in step S1 according to a mass-to-volume ratio of 100 - 120 mg:60 - 80 mg:10 - 15 mL, heat to 30 - 40 °C, stir until evenly dispersed, and then let it stand at room temperature for 20 - 30 min to obtain Solution 3; S3. Weigh laccase and the Solution 3 obtained in step S2 according to a mass-to-volume ratio of 80 - 100 mg:10 - 15 mL, heat to 32 - 36 °C, stir at a speed of 100 - 150 rpm for 18 - 20 h. After that, centrifuge at a centrifugal force of 5000 - 6000 g for 8 - 10 min, concentrate the supernatant to 0.4 - 0.5 times the original volume, and vacuum freeze-dry to obtain the complex; S4. Dissolve chitosan in acetic acid solution to obtain a chitosan solution with a mass percentage concentration of 3 - 5%. Adjust the pH to 5 - 6, add the complex obtained in step S3, with the addition amount of the complex being 8 - 10 mg / mL. Stir at a speed of 150 - 200 rpm at room temperature for 20 - 30 min. After that, centrifuge at a centrifugal force of 8000 - 10000 g for 10 - 15 min. Disperse the obtained precipitate in trehalose solution to obtain a mixture. Spray-dry the mixture to obtain the detoxification and degradation agent.

[0011] Preferably, in step S1, the concentration of the sodium hydroxide solution is 3 mol / L, and hydrochloric acid solution is preferably used to adjust the pH.

[0012] Preferably, in step S2, EDC is 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide with the CAS number 1892 - 57 - 5, and NHS is N-hydroxysuccinimide with the CAS number 6066 - 82 - 6.

[0013] Preferably, in step S3, laccase is derived from Coriolus versicolor, and the enzyme activity is preferably 200 - 300 U / g.

[0014] Preferably, in step S4, the mass percentage concentration of the acetic acid solution is preferably 2%, the mass-volume concentration of the precipitate in the trehalose solution is preferably 1 - 3 mg / mL, the mass percentage concentration of the trehalose solution is preferably 6%, and spray drying is preferably carried out at an air flow rate of 1.5 - 2 m 3 / min, an inlet air temperature of 70 - 80 °C, and an outlet air temperature of 50 - 60 °C.

[0015] The present invention also provides a preparation method of a soluble powder for treating glyphosate-induced livestock liver injury, and the specific steps are as follows: V1. Weigh sucrose, liver protection complex, and detoxification and degradation agent according to the corresponding weight parts. Dissolve sucrose in water to obtain a sucrose solution with a mass percentage concentration of 5 - 8%. Mix the sucrose solution with the detoxification and degradation agent. After spray drying, the obtained fine powder is frozen to obtain the core content. V2. Dissolve the liver protection complex in water to obtain solution 4 with a mass percentage concentration of 10 - 15%. Add the core content obtained in step V1 into solution 4. After quickly stirring until evenly dispersed, vacuum freeze-dry to obtain the soluble powder for treating glyphosate-induced livestock liver injury.

[0016] Preferably, in step V1, spray drying is preferably carried out at an air flow rate of 2 - 3 m 3 / min, an inlet air temperature of 50 - 60 °C, and an outlet air temperature of 30 - 40 °C. Freezing is carried out below 0 °C, preferably freezing at -20 °C for 48 h.

[0017] Preferably, in step V2, vacuum freeze-drying is preferably carried out at a pre-cooling temperature of -20°C, with a shelf temperature of -10°C and a vacuum degree of 8 Pa.

[0018] The beneficial effects of the present invention are as follows: The present invention prepares a soluble powder, which is preferably dissolved in water and given to livestock before they eat grass or feed, and can better alleviate the liver damage induced by glyphosate and its degradation products in livestock. The soluble powder of the present invention contains sucrose, a liver protection complex, and a detoxification and degradation agent. Sucrose plays a role in attracting food. The liver protection complex is obtained by deodorizing fresh spirulina and then modifying it with epigallocatechin gallate, and finally retaining the low molecular weight part. The detoxification and degradation agent is obtained by modifying laccase with rosmarinic acid, and the obtained complex is further encapsulated with chitosan and trehalose. The soluble powder of the present invention has a core-shell structure, with the liver protection complex as the shell material and the detoxification and degradation agent as the inner core material. In the actual application process, the liver protection complex is first released into water, and then the detoxification and degradation agent is released. The soluble powder of the present invention can be completely dissolved within 60 s without solid residues. Toxicological experiments show that it has a high safety level and does not affect the normal life of mice. Using white leghorn chickens for livestock experiments shows that when the soluble powder is prepared and used immediately and added to drinking water, it can reduce the liver damage induced by glyphosate, maintain the function of the liver antioxidant system as much as possible, and effectively resist the oxidative stress brought by glyphosate, etc. Description of the Drawings

[0019] The drawings described herein are used to provide a further understanding of the present application and constitute a part of the present application. The schematic embodiments and descriptions thereof are used to explain the present application and do not constitute an improper limitation to the present application. In the drawings: Figure 1 It is a graph of the dissolution time results of the liver protection complex, the detoxification and degradation agent, and the soluble powder; Figure 2 It is a graph of the body weight change of male mice during the toxicological experiment; Figure 3 It is a graph of the body weight change of female mice during the toxicological experiment; Figure 4 It is a graph of the results of the contents of ALT and AST in the serum after the experiment on sexually mature white leghorn chickens; Figure 5 It is a graph of the results of the contents of ALP and γ-GT in the serum after the experiment on sexually mature white leghorn chickens; Figure 6 It is a graph of the results of the contents of ALT and AST in the serum after the experiment on 2-month-old white leghorn chickens; Figure 7 It is a graph of the results of the contents of ALP and γ-GT in the serum after the experiment on 2-month-old white leghorn chickens; Figure 8Results graph of the contents of CAT, SOD, and GPx in the chicken liver after the experiment on sexually mature white - feather chickens; Figure 9 Results graph of the contents of CAT, SOD, and GPx in the chicken liver after the experiment on 2 - month - old white - feather chickens; Figure 10 Results graph of the contents of MDA, PCO, and AOPP in the chicken liver after the experiment on sexually mature white - feather chickens; Figure 11 Results graph of the contents of MDA, PCO, and AOPP in the chicken liver after the experiment on 2 - month - old white - feather chickens. Detailed implementation manners

[0020] In order to more clearly explain the overall concept of this application, the following will be described in detail by way of examples in conjunction with the accompanying drawings of the specification. In the following description, a large number of specific details are given to provide a more thorough understanding of the present invention. However, it is obvious to those skilled in the art that the present invention can be implemented without one or more of these details. In other examples, in order to avoid confusion with the present invention, some well - known technical features in the art are not described.

[0021] Example 1: This example provides a preparation method of a liver - protecting complex, and the specific steps are as follows: L1. After washing fresh spirulina with water, blanch it in 80°C water for 1 min, then air - dry the surface moisture to obtain a spirulina sample. Crush ginger charcoal and sieve it to obtain ginger charcoal powder with a particle size of 80 μm. Disperse the ginger charcoal powder in water to obtain a ginger charcoal suspension with a mass percentage concentration of 4%. Immerse all the spirulina samples in the ginger charcoal suspension, let it stand at room temperature for 2 h, then wash and completely dry it, and grind it to obtain spirulina powder with a particle size of 10 μm; L2. Weigh the spirulina powder obtained in step L1, an aqueous solution of xanthan gum with a mass percentage concentration of 3%, and epigallocatechin gallate according to a mass ratio of 8:10:1. After mixing the spirulina powder and the aqueous solution of xanthan gum, ultrasonically disperse it until it is uniform, then add epigallocatechin gallate, adjust the pH to 7, heat it to 30°C and keep it for 4 h to obtain solution 1; L3. Dialyze the solution 1 obtained in step L2 at 4°C for 10 h. Select a dialysis bag with a cut - off molecular weight of 9 kDa for dialysis. Replace the dialysis medium (deionized water) 2 times during dialysis. After that, pre - cool the substances in the dialysis bag at - 60°C for 15 h, and perform vacuum freeze - drying at a partition temperature of - 50°C and a vacuum degree of 5 Pa to obtain the liver - protecting complex.

[0022] Example 2: This example provides a preparation method of a liver - protecting complex, and the specific steps are as follows: L1. After washing fresh spirulina with water, blanch it in water at 80 °C for 1.5 min, then air-dry the surface moisture to obtain a spirulina sample. Crush ginger charcoal and sieve it to obtain ginger charcoal powder with a particle size of 90 μm. Disperse the ginger charcoal powder in water to obtain a ginger charcoal suspension with a mass percentage concentration of 5%. Immerse all of the spirulina sample in the ginger charcoal suspension, let it stand at room temperature for 2.5 h, then wash it after completion, and grind it after completely drying to obtain spirulina powder with a particle size of 15 μm; L2. Weigh the spirulina powder obtained in step L1, an aqueous solution of xanthan gum with a mass percentage concentration of 4%, and epigallocatechin gallate according to a mass ratio of 9:11:1. After mixing the spirulina powder and the aqueous solution of xanthan gum, ultrasonically disperse them until uniform, then add epigallocatechin gallate, adjust the pH to 7.5, heat to 35 °C and maintain for 6 h to obtain Solution 1 after completion; L3. Dialyze the Solution 1 obtained in step L2 at 4 °C for 11 h. Select a dialysis bag with a cut-off molecular weight of 10 kDa for dialysis, and change the dialysis medium (deionized water) 2 times during dialysis. After completion, pre-cool the substance in the dialysis bag at -60 °C for 12 h, and perform vacuum freeze-drying at a partition temperature of -50 °C and a vacuum degree of 5 Pa to obtain the liver protection complex.

[0023] Example 3: This example provides a method for preparing a liver protection complex, and the specific steps are as follows: L1. After washing fresh spirulina with water, blanch it in water at 80 °C for 2 min, then air-dry the surface moisture to obtain a spirulina sample. Crush ginger charcoal and sieve it to obtain ginger charcoal powder with a particle size of 100 μm. Disperse the ginger charcoal powder in water to obtain a ginger charcoal suspension with a mass percentage concentration of 8%. Immerse all of the spirulina sample in the ginger charcoal suspension, let it stand at room temperature for 3 h, then wash it after completion, and grind it after completely drying to obtain spirulina powder with a particle size of 20 μm; L2. Weigh the spirulina powder obtained in step L1, an aqueous solution of xanthan gum with a mass percentage concentration of 5%, and epigallocatechin gallate according to a mass ratio of 10:15:1. After mixing the spirulina powder and the aqueous solution of xanthan gum, ultrasonically disperse them until uniform, then add epigallocatechin gallate, adjust the pH to 8, heat to 40 °C and maintain for 8 h to obtain Solution 1 after completion; L3. Dialyze the Solution 1 obtained in step L2 at 4 °C for 12 h. Select a dialysis bag with a cut-off molecular weight of 10 kDa for dialysis, and change the dialysis medium (deionized water) 3 times during dialysis. After completion, pre-cool the substance in the dialysis bag at -60 °C for 14 h, and perform vacuum freeze-drying at a partition temperature of -50 °C and a vacuum degree of 5 Pa to obtain the liver protection complex.

[0024] Example 4: This example provides a method for preparing a detoxification and degradation agent, and the specific steps are as follows: S1. Weigh rosmarinic acid and 3 mol / L sodium hydroxide solution according to the mass-volume ratio of 40 mg: 6 mL, stir until evenly dispersed, and adjust the pH to 7 with 1 mol / L hydrochloric acid solution to obtain Solution 2; S2. Weigh EDC, NHS and the Solution 2 obtained in Step S1 according to the mass-volume ratio of 100 mg: 60 mg: 10 mL. EDC is 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide with the CAS number 1892-57-5, and NHS is N-hydroxysuccinimide with the CAS number 6066-82-6. Heat to 30 °C, stir until evenly dispersed, and then let it stand at room temperature for 20 min to obtain Solution 3; S3. Weigh laccase and the Solution 3 obtained in Step S2 according to the mass-volume ratio of 80 mg: 10 mL. The laccase is derived from Coriolus versicolor with an enzyme activity of 200 U / g. Heat to 32 °C, stir at a speed of 100 rpm for 18 h, and after completion, centrifuge at a centrifugal force of 5000 g for 8 min. Concentrate the supernatant to 0.4 times the original volume and obtain the complex after vacuum freeze-drying; S4. Dissolve chitosan in acetic acid solution with a mass percentage concentration of 2% to obtain a chitosan solution with a mass percentage concentration of 3%. Adjust the pH to 5, add the complex obtained in Step S3, and the addition amount of the complex is 8 mg / mL. Stir at a speed of 150 rpm at room temperature for 20 min. After completion, centrifuge at a centrifugal force of 8000 g for 10 min. Disperse the obtained precipitate in a trehalose solution with a mass percentage concentration of 6%. The mass-volume concentration of the precipitate in the trehalose solution is 1 mg / mL. The obtained mixture is spray-dried at an air flow rate of 1.5 m 3 / min, an inlet air temperature of 70 °C, and an outlet air temperature of 50 °C to obtain the detoxification and degradation agent.

[0025] Example 5: This example provides a preparation method of a detoxification and degradation agent, and the specific steps are as follows: S1. Weigh rosmarinic acid and 3 mol / L sodium hydroxide solution according to the mass-volume ratio of 45 mg: 8 mL, stir until evenly dispersed, and adjust the pH to 7.2 with 1 mol / L hydrochloric acid solution to obtain Solution 2; S2. Weigh EDC, NHS and the Solution 2 obtained in Step S1 according to the mass-volume ratio of 110 mg: 70 mg: 12 mL. EDC is 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide with the CAS number 1892-57-5, and NHS is N-hydroxysuccinimide with the CAS number 6066-82-6. Heat to 35 °C, stir until evenly dispersed, and then let it stand at room temperature for 24 min to obtain Solution 3; S3. Weigh laccase and the solution 3 obtained in step S2 according to the mass-to-volume ratio of 90 mg: 12 mL. The laccase is derived from Coriolus versicolor with an enzyme activity of 250 U / g. Heat it to 34 °C and stir it at a speed of 120 rpm for 19 h. After that, centrifuge it at a centrifugal force of 5400 g for 9 min. Concentrate the supernatant to 0.45 times the original volume and obtain the complex after vacuum freeze-drying. S4. Dissolve chitosan in an acetic acid solution with a mass percentage concentration of 2% to obtain a chitosan solution with a mass percentage concentration of 4%. Adjust the pH to 5.5, and add the complex obtained in step S3. The addition amount of the complex is 9 mg / mL. Stir it at a speed of 180 rpm at room temperature for 24 min. After that, centrifuge it at a centrifugal force of 9000 g for 12 min. Disperse the obtained precipitate in a trehalose solution with a mass percentage concentration of 6%. The mass-to-volume concentration of the precipitate in the trehalose solution is 2 mg / mL. The obtained mixture is spray-dried at an air flow rate of 1.8 m 3 / min, an inlet air temperature of 75 °C, and an outlet air temperature of 55 °C to obtain the detoxifying and degrading agent.

[0026] Example 6: This example provides a method for preparing a detoxifying and degrading agent, and the specific steps are as follows: S1. Weigh rosmarinic acid and a 3 mol / L sodium hydroxide solution according to the mass-to-volume ratio of 50 mg: 9 mL, stir until evenly dispersed, and adjust the pH to 7.5 with a 1 mol / L hydrochloric acid solution to obtain solution 2. S2. Weigh EDC, NHS and the solution 2 obtained in step S1 according to the mass-to-volume ratio of 120 mg: 80 mg: 15 mL. EDC is 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide with a CAS number of 1892-57-5, and NHS is N-hydroxysuccinimide with a CAS number of 6066-82-6. Heat it to 40 °C, stir until evenly dispersed, and then let it stand at room temperature for 30 min to obtain solution 3. S3. Weigh laccase and the solution 3 obtained in step S2 according to the mass-to-volume ratio of 100 mg: 15 mL. The laccase is derived from Coriolus versicolor with an enzyme activity of 300 U / g. Heat it to 36 °C and stir it at a speed of 150 rpm for 20 h. After that, centrifuge it at a centrifugal force of 6000 g for 10 min. Concentrate the supernatant to 0.5 times the original volume and obtain the complex after vacuum freeze-drying. S4. Dissolve chitosan in an acetic acid solution with a mass percentage concentration of 2% to obtain a chitosan solution with a mass percentage concentration of 5%. Adjust the pH to 6, add the complex obtained in step S3, with the addition amount of the complex being 10 mg / mL, stir at a speed of 200 rpm at room temperature for 30 min, and then centrifuge at a centrifugal force of 10000 g for 15 min. The obtained precipitate is dispersed in a trehalose solution with a mass percentage concentration of 6%, and the mass-volume concentration of the precipitate in the trehalose solution is 3 mg / mL. The obtained mixture is spray-dried at an air flow rate of 2 m 3 / min, an inlet air temperature of 80 °C, and an outlet air temperature of 60 °C to obtain the detoxifying and degrading agent.

[0027] Example 7: This example provides a preparation method for a soluble powder for treating glyphosate-induced livestock liver injury. The specific steps are as follows: V1. Weigh 0.5 parts of sucrose, 2 parts of liver protection complex, and 3 parts of detoxifying and degrading agent by weight. The liver protection complex is prepared from Example 1, and the detoxifying and degrading agent is prepared from Example 4. Dissolve sucrose in water to obtain a sucrose solution with a mass percentage concentration of 5%. Mix the sucrose solution with the detoxifying and degrading agent, and spray-dry at an air flow rate of 2 m 3 / min, an inlet air temperature of 50 °C, and an outlet air temperature of 30 °C. The obtained fine powder is frozen at -20 °C for 48 h to obtain the core content; V2. Dissolve the liver protection complex in water to obtain solution 4 with a mass percentage concentration of 10%. Add the core content obtained in step V1 to solution 4, quickly stir until evenly dispersed, pre-cool at -20 °C for 15 h, and then perform vacuum freeze-drying at a partition temperature of -10 °C and a vacuum degree of 8 Pa to obtain the soluble powder for treating glyphosate-induced livestock liver injury.

[0028] Example 8: This example provides a preparation method for a soluble powder for treating glyphosate-induced livestock liver injury. The specific steps are as follows: V1. Weigh 1 part of sucrose, 3 parts of liver protection complex, and 4 parts of detoxifying and degrading agent by weight. The liver protection complex is prepared from Example 2, and the detoxifying and degrading agent is prepared from Example 5. Dissolve sucrose in water to obtain a sucrose solution with a mass percentage concentration of 6%. Mix the sucrose solution with the detoxifying and degrading agent, and spray-dry at an air flow rate of 2.5 m 3 / min, an inlet air temperature of 55 °C, and an outlet air temperature of 35 °C. The obtained fine powder is frozen at -20 °C for 48 h to obtain the core content; V2. Dissolve the liver protection complex in water to obtain Solution 4 with a mass percentage concentration of 12%. Add the nuclear content obtained in Step V1 to Solution 4, quickly stir until evenly dispersed, pre-cool at -20°C for 18 h, and then perform vacuum freeze-drying at a partition temperature of -10°C and a vacuum degree of 8 Pa to obtain the soluble powder for treating glyphosate-induced liver injury in livestock.

[0029] Example 9: This example provides a preparation method for a soluble powder for treating glyphosate-induced liver injury in livestock. The specific steps are as follows: V1. Weigh 1.5 parts of sucrose, 5 parts of liver protection complex, and 5 parts of detoxification and degradation agent by weight. The liver protection complex is prepared according to Example 3, and the detoxification and degradation agent is prepared according to Example 6. Dissolve the sucrose in water to obtain a sucrose solution with a mass percentage concentration of 8%. Mix the sucrose solution with the detoxification and degradation agent, and perform spray drying at an air flow rate of 3 m 3 / min, an inlet air temperature of 60°C, and an outlet air temperature of 40°C. The obtained fine powder is frozen at -20°C for 48 h to obtain the nuclear content; V2. Dissolve the liver protection complex in water to obtain Solution 4 with a mass percentage concentration of 15%. Add the nuclear content obtained in Step V1 to Solution 4, quickly stir until evenly dispersed, pre-cool at -20°C for 20 h, and then perform vacuum freeze-drying at a partition temperature of -10°C and a vacuum degree of 8 Pa to obtain the soluble powder for treating glyphosate-induced liver injury in livestock.

[0030] Comparative Example 1: This comparative example provides a preparation method for a liquid preparation. The specific steps are as follows: Weigh 5 parts of the liver protection complex obtained in Example 3 and 1152 parts of water by weight, mix them until evenly dispersed, and store at room temperature for 30 days to obtain the liquid preparation of this comparative example.

[0031] Comparative Example 2: This comparative example provides a preparation method for a liquid preparation. The specific steps are as follows: Weigh 5 parts of the detoxification and degradation agent obtained in Example 6 and 1152 parts of water by weight, mix them until evenly dispersed, and store at room temperature for 30 days to obtain the liquid preparation of this comparative example.

[0032] Comparative Example 3: This comparative example provides a preparation method for a liquid preparation. The specific steps are as follows: Weigh 5 parts of the soluble powder obtained in Example 9 and 500 parts of water by weight, mix them until evenly dispersed, and store at room temperature for 30 days to obtain the liquid preparation of this comparative example.

[0033] Experimental test: I. Determination of dissolution time: Detect the dissolution time of the substances obtained in Examples 1-9. Taking Example 1 as an example, weigh 25 g of the liver protection complex in Example 1 into a 500 mL beaker, add 200 mL of cold boiled water (10 °C), stir, and calculate the time from adding the cold boiled water to complete dissolution.

[0034] The dissolution times of the liver protection complexes / detoxifying and degrading agents / soluble powders obtained in Examples 1-9 are as Figure 1 shown. It can be seen from Figure 1 that all of the above substances can be completely dissolved within 60 s.

[0035] II. Toxicology experiment: Select 100 Kunming mice, with 50 males and 50 females. Feed them adaptively for one week in an environment with a temperature of 20-26 °C and a humidity of 40-60%. The daily diet is the SPF-sized mouse growth and reproduction feed produced by Beijing Keao Xieli Feed Co., Ltd., production license number: SCXK(Beijing)2012-0019. Sterilized corncob bedding is used in the cages and replaced twice a week, and the cages are also replaced and washed and disinfected. The use of experimental animals (mice) follows the measures for the use and management of experimental animals and complies with animal welfare policies and relevant requirements.

[0036] After the adaptive feeding period, start the experiment. Randomly divide the mice into ten groups and label them as the first group to the tenth group in sequence. The first group is the control group, and the second group to the tenth group correspond to Examples 1-9 respectively. The dosage of the substances obtained in Examples 1-9 is 4 g / kg. Calculate the dosage of the substances obtained in Examples 1-9 according to the body weight of the mice, dissolve and make up the volume to 1.2 mL with water, and then intragastrically administer. Replace the control group with the same amount of normal saline for intragastric administration. Repeat once a day, observe the daily food intake, water intake and activity of the mice, count the death situation. If a mouse dies, immediately dissect it to determine the cause of death and observe the volume, color, texture, etc. of the main organs. The experiment lasts for 21 days. The results of the death situation of the mice at the end of the experiment are as follows: Table 1 Death situation of mice

[0037] During the experiment, the weight change situations of male mice and female mice are respectively as Figure 2 , Figure 3 shown. It can be seen from Figure 2 , Figure 3 that during the experiment, the mice in the second group to the tenth group lived normally and there was no significant difference in body weight from the mice in the control group, indicating that the liver protection complexes / detoxifying and degrading agents / soluble powders obtained in Examples 1-9 are non-toxic.

[0038] III. Livestock experiment: Considering the growth cycles of different livestock, chickens were selected to conduct livestock experiments. Sexually mature white - feather chickens and 2 - month - old white - feather chickens were respectively selected for the experiments. After 10 days of adaptive feeding, the experiments began. Taking the sexually mature white - feather chickens as an example, they were randomly divided into 10 groups, with 15 chickens in each group. The first group was the control group, the second group was the drug group, and the third to tenth groups were all experimental groups, corresponding to Example 3, Example 6, Examples 7 - 9, and Comparative Examples 1 - 3 respectively. In Example 3 and Example 6, they were dissolved at the same ratio as in Comparative Example 1 or Comparative Example 2, while in Examples 7 - 9, they were dissolved at the same ratio as in Comparative Example 3. The examples were used immediately after dissolution.

[0039] The white - feather chickens were not provided with drinking water from 0:00 to 7:00 every day. Before feeding the chicken feed in the morning, the white - feather chickens were given special water. The water in the control group and the drug group was water without any drugs, and the water in the experimental groups was water dissolved with liver - protection complex / detoxification and degradation agent / soluble powder respectively. The supply amount of the special water was 200 mL per chicken. Subsequently, chicken feed was provided. The feed in the control group was normal chicken feed, and the feed in the drug group and the experimental groups was chicken feed containing 200 mg of glyphosate (Merck Chemical Technology (Shanghai) Co., Ltd., analytical pure). The experiment lasted for three months, and then the white - feather chickens were sacrificed. The contents of alanine aminotransferase (ALT), aspartate aminotransferase (AST), alkaline phosphatase (ALP), and γ - glutamyl transpeptidase (γ - GT) in the serum were detected (using a kit for detection, and the kits were purchased from Shanghai Meiao Biotechnology Co., Ltd., with the product numbers being MO - J30625P, MO - J30626P, MO - J30660P, and MO - J31226P in sequence). Chicken livers were obtained, and at the same time, the contents of catalase (CAT) (MO - J30715P), superoxide dismutase (SOD) (MO - J30494P), glutathione peroxidase (GPx) (Nanjing Dulai Biotechnology Co., Ltd., E0117), and the contents of malondialdehyde (MDA) (MO - J30490P), protein carbonyl compounds (PCO) (Nanjing Jiancheng Bioengineering Institute, A087), and advanced oxidation protein products (AOPP) (Jianglai Bio, ELISA detection kit) in the chicken livers were detected.

[0040] ALT, AST, ALP, and γ - GT are four common liver damage indicators. After feeding the chicken feed containing glyphosate, the results of the above - mentioned indicators of sexually mature white - feather chickens are as Figure 4 、 Figure 5 shown, and the results of the above - mentioned indicators of 2 - month - old white - feather chickens are as Figure 6 、 Figure 7 shown. From Figures 4 - 7It can be seen that glyphosate will cause a significant increase in the content of the above indicators, showing a significant difference from the control group. After drinking the water prepared in Examples 7-9 in advance, the above indicators of the fifth, sixth, and seventh groups are close to the values of the control group. From the comparison of the values of the third and eighth groups, it can be seen that the effect of the freshly dissolved liver protection complex is better than that of the liquid preparation after being stored for 30 days. From the comparison of the values of the fourth and ninth groups, it can be seen that the detoxification and degradation agent has similar properties, and the freshly dissolved effect is also better. From the comparison of the values of the third, fourth, seventh groups, and the eighth, ninth, tenth groups, it can be seen that the special water obtained from a single liver protection complex or detoxification and degradation agent has a worse effect than the special water prepared from the soluble powder containing both of the above two substances. Therefore, the comparison of the values of the liver damage indicators shows that the soluble powder obtained in the present invention has a better liver protection effect, and the best usage method is to prepare and use it immediately as needed.

[0041] CAT, SOD, and GPx characterize the antioxidant system of the liver. The relevant results of sexually mature white - feather chickens and 2 - month - old white - feather chickens are as Figure 8 、 Figure 9 shown. From Figure 8 、 Figure 9 it can be seen that after the white - feather chickens drink the special water prepared from the soluble powder immediately, their antioxidant system can be maintained at a better level, and then continuously play the role of degrading and removing toxic substances.

[0042] MDA, PCO, and AOPP can reflect the degree of peroxidative damage of liver tissue, or can be described as the degree of oxidative stress suffered. Figure 10 、 Figure 11 are the test results of the above indicators of sexually mature white - feather chickens and 2 - month - old white - feather chickens respectively. From Figure 10 、 Figure 11 it can be seen that for the white - feather chickens in the second group without any protective measures, after eating the chicken feed containing glyphosate, the contents of MDA, PCO, and AOPP increase significantly. In contrast, the above indicators of the fifth to seventh groups are still controlled within an acceptable range. Therefore, the soluble powder obtained in the present invention can effectively resist the oxidative stress caused by glyphosate, and oxidative stress is one of the manifestations of liver damage.

[0043] In addition, the above data show that at the best stage of the growth and reproduction of white - feather chickens, intervening to resist glyphosate as early as possible can effectively avoid problems such as oxidative stress and liver damage caused by glyphosate, and then help the white - feather chickens gain weight and increase economic benefits.

[0044] The present invention also used aminomethylphosphonic acid with a purity of 99% purchased from Merck Chemical Technology (Shanghai) Co., Ltd. to conduct the above livestock experiments, and obtained experimental results similar to those of glyphosate drugs, indicating that the soluble powder of the present application can more comprehensively cope with the liver damage caused by glyphosate and its degradation products.

[0045] As described above, these are only several embodiments of the present invention and do not impose any form of limitation on the present invention. Although the present invention is disclosed above with preferred embodiments, it is not intended to limit the present invention. Any person skilled in the relevant art, without departing from the technical solution of the present invention, making some changes or modifications using the disclosed technical content is equivalent to equivalent implementation cases and all fall within the scope of protection of the technical solution of the present invention.

Claims

1. A soluble powder for treating glyphosate-induced liver damage in livestock, characterized in that: Contains the following raw materials in parts by weight: 0.5-1.5 parts of sucrose, 2-5 parts of liver protection complex, 3-5 parts of detoxification and degradation agent.

2. The soluble powder for treating glyphosate-induced liver damage in livestock according to claim 1, characterized in that: The preparation method of the liver protection complex is as follows: L1. After washing fresh spirulina with water, blanching, and then air-drying the surface moisture, obtain a spirulina sample, crush ginger charcoal and sieve it to obtain ginger charcoal powder, disperse the ginger charcoal powder in water to obtain a ginger charcoal suspension with a mass percentage concentration of 4-8%, immerse the spirulina sample in the ginger charcoal suspension, let it stand at room temperature, wash it after completion, and grind it to obtain spirulina powder with a particle size of 10-20μm; L2. The spirulina powder, xanthan gum aqueous solution and epigallocatechin gallate obtained in step L1 are weighed in a mass ratio of 8-10:10-15:1, the spirulina powder and the xanthan gum aqueous solution are mixed, ultrasonically dispersed until uniform, and then epigallocatechin gallate is added, the pH is adjusted, and heating and maintaining is performed to obtain solution 1 after completion; L3. The solution 1 obtained in step L2 is dialyzed, and after the dialyzation, the substance in the dialysis bag is vacuum freeze-dried to obtain a liver protection complex.

3. The soluble powder for treating glyphosate-induced liver damage in livestock according to claim 2, characterized in that: The mass percent concentration of the xanthan gum aqueous solution in step L2 is 3-5%.

4. The soluble powder for treating glyphosate-induced liver damage in livestock according to claim 3, characterized in that: The preparation method of the detoxification degradation agent is as follows: S1. Weigh rosmarinic acid and sodium hydroxide solution according to a mass volume ratio of 40-50 mg: 6-9 mL, stir until uniformly dispersed, adjust the pH, and obtain solution 2; S2. Weigh EDC, NHS and solution 2 obtained in step S1 according to a mass volume ratio of 100-120 mg: 60-80 mg: 10-15 mL, heat, stir until uniformly dispersed, and then stand at room temperature to obtain solution 3; S3. Laccase and solution 3 obtained in step S2 were weighed according to a mass volume ratio of 80-100 mg: 10-15 mL, heated, stirred, and centrifuged after completion. The supernatant was concentrated to 0.4-0.5 times the original volume, and the complex was obtained after vacuum freeze-drying; S4. Dissolve chitosan in acetic acid solution to obtain a chitosan solution with a mass percentage concentration of 3-5%, adjust the pH, add the complex obtained in step S3, the amount of the complex added is 8-10 mg / mL, stir at room temperature, centrifuge after completion, disperse the obtained precipitate in the trehalose solution to obtain a mixture, spray-dry the mixture to obtain a detoxification and degradation agent.

5. The soluble powder for treating glyphosate-induced liver damage in livestock according to claim 4, characterized in that: The concentration of the sodium hydroxide solution in step S1 is 3 mol / L.

6. A method for preparing a soluble powder for treating glyphosate-induced liver damage in livestock according to any one of claims 1 to 5, characterized in that: The specific steps are as follows: V1. dissolving sucrose in water to obtain a sucrose solution, mixing the sucrose solution with a detoxifying and degrading agent, spray drying, and freezing the obtained micropowder to obtain the core contents; V2. Dissolve the liver protection complex in water to obtain solution 4, add the core content obtained in step V1 to solution 4, quickly stir until uniformly dispersed, and vacuum freeze-dry to obtain a soluble powder for treating glyphosate-induced livestock liver damage.

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