A method for synthesizing humic acid complex for feed by biological enzyme method

Abstract

The invention discloses a method for synthesizing humic acid complex for feed by enzymatic method. First, macromolecular humic acid is extracted from weathered coal in the invention, then the macromolecular humic acid is degraded by biological enzyme and complexed with mineral salt, and the humic acid complex for feed is obtained after cooling and crystallization. The humic acid complex used in the invention replaces inorganic trace elements, and can reduce the emission of trace elements in the environment while ensuring the normal production performance of animals.

Description

Technical Field

[0001] The present invention relates to the technical field of organic trace element synthesis, and particularly relates to a method for synthesizing a humic acid complex for feed by a bio-enzymatic method. Background Art

[0002] Humic acid is a class of organic substances formed and accumulated through a series of processes of microbial decomposition and transformation of animal and plant remains, mainly plant remains, as well as geochemistry. As an additive, humic acid can effectively improve the immunity of livestock and poultry and reduce the dosage of antibiotics; at the same time, it has the advantages of promoting growth, improving feed conversion rate, improving the feeding environment, improving the quality of livestock and poultry products, and being environmentally friendly, and has become one of the best choices for current pharmaceutical feed additives and antibiotic substitutes. As is well known, weathered coal contains a relatively high content of humic acid. China has large reserves and a wide distribution of weathered coal resources. For a long time, weathered coal resources have been arbitrarily abandoned, seriously polluting the environment and affecting the improvement of the living standards of humans. Using weathered coal as the raw material for the development and utilization of humic acid resources to provide raw materials for the production of humic acid products not only solves the pollution problem of weathered coal resources, but also provides a new direction for the development of the coal industry.

[0003] Trace mineral elements are essential nutritional elements for animal organisms. In the feed industry, at first, inorganic salts were used to meet the body's needs, but the absorption rate of inorganic salts is low and there are many side effects, and they have gradually been replaced by some organic salts in the market. Organic salts are well-known for their higher biological potency, stronger stability, and easier absorption than inorganic salts. At present, amino acid complexes have become a new generation of trace element supplements for animals, not only having the advantages of organic acid salts, but also having the advantages of easier absorption and more stable structure.

[0004] Although the absorption rate of amino acid complexes is the highest, their price is relatively high. Currently, in the feed market, the competition is very fierce, and it is urgent to find a low-cost trace element supplement. Using humic acid as the main raw material and complexing it with mineral salts, the obtained humic acid complex not only has the effect of "1 + 1 > 2", but also can reduce the production cost of enterprises and improve the competitiveness of enterprises. Therefore, it is necessary to develop a preparation process for humic acid complexes that is environmentally friendly and has a high extraction efficiency.

[0005] Humic acid complexes, such as copper humate, iron humate, zinc humate, etc., are currently widely used in the fertilizer field, but have not been seen in the feed field. Due to the different absorption mechanisms of animals and plants, and animals having higher safety requirements, the preparation method of humic acid complexes for fertilizers is not applicable to the preparation of humic acid complexes for feeds. Moreover, the traditional chemical synthesis method has many steps, many by-products, and difficult product purification. Generally, it can only meet the requirements of small-scale laboratory preparation and is not suitable for industrial production. Therefore, there is an urgent need to find a new preparation method for humic acid complexes for feeds. Summary of the Invention

[0006] In view of the above problems, the present invention provides a method for synthesizing humic acid complexes for feeds by a biological enzyme method. The present invention first extracts macromolecular humic acid from weathered coal, then uses biological enzymes to degrade the macromolecular humic acid and complex it with mineral salts, and after cooling and crystallization, a humic acid complex for feeds is obtained. The humic acid complex used in the present invention replaces inorganic trace elements, and can reduce the emission of trace elements in the environment while ensuring the normal production performance of animals, which will play a positive guiding and promoting role in the development of the organic trace element industry and even the livestock industry.

[0007] The technical solution of the present invention is: a method for synthesizing humic acid complexes for feeds by a biological enzyme method, which is characterized by including the following steps:

[0008] (1) Extracting macromolecular humic acid from weathered coal

[0009] Put weathered coal and water into a reaction vessel, stir and heat up, adjust the pH to 9-11 with sodium hydroxide to dissolve the weathered coal, filter out the insoluble substances to obtain a refined humic acid extract, then concentrate it, and add acid to the concentrated solution to adjust the pH to 3.5-4 to obtain a macromolecular humic acid feed liquid;

[0010] (2) Degrading and complexing macromolecular humic acid with biological enzymes

[0011] For the macromolecular humic acid feed liquid prepared in step (1), add sodium hydroxide to adjust the pH of the solution to 9-11.5, then add a biological enzyme solution, carry out an enzymatic reaction at 35-50 °C for 0.5-1.5 h, and then add mineral salts and stir and react for 1-4 h to obtain a humic acid complex solution;

[0012] (3) Cooling crystallization of the humic acid complex:

[0013] Cool the humic acid complex solution prepared in step (2) for crystallization, carry out centrifugal separation, and dry to obtain the humic acid complex. Its production process flow chart is as Figure 1 shown.

[0014] The above-mentioned biological enzyme solution is prepared as follows: Take biological enzymes and dissolve them in a quaternary ammonium ionic liquid. After complete dissolution, pre-cool the solution in an environment at 1 - 5°C for standby. The above-mentioned biological enzymes are one or more of tannin acyl hydrolase, phytase, laccase, pullulanase, and cellulase, and more preferably one or more of tannin acyl hydrolase, phytase, and laccase. The mass ratio of the biological enzyme to the ionic liquid is 1:2 - 10. The ionic liquid can be regarded as a network structure, and the biological enzyme can be evenly dispersed in the network structure of the ionic liquid, so that the biological enzyme can be better dispersed in the aqueous solution, thereby improving the stability of the enzyme and maintaining the catalytic activity and selectivity of the enzyme.

[0015] The above-mentioned quaternary ammonium ionic liquid includes one or more of diethylamine acetate ionic liquid, diethylamine nitrate ionic liquid, and diethylamine formate ionic liquid.

[0016] The above-mentioned mineral salts are one or more of chlorides, sulfates, nitrates, and acetates of copper, iron, zinc, manganese, chromium, cobalt, etc. Further preferably, they are one or more of chlorides, sulfates, and acetates.

[0017] Further, the mother liquor after centrifugal filtration in step (3) is pumped into the reaction vessel in step (1) to continue as the reaction solution (replacing part of the water) for recycling.

[0018] Preferably, the total humic acid content in the weathered coal in step (1) is 45 - 65%, more preferably 55%; the mass ratio of the weathered coal to water is 1:3 - 10.

[0019] Preferably, in step (1), the temperature is raised by stirring, and the temperature after raising is 80 - 90°C. Among them, the solution is concentrated until the solid content in the feed liquid is 18 - 20%.

[0020] Preferably, the addition amount of the biological enzyme in step (2) is 0.01 - 0.5% of the mass of the weathered coal, preferably 0.01 - 0.1%; among them, the enzyme activity of tannin acyl hydrolase is ≥500 U / g; the activity of phytase is ≥10000 U / g; the enzyme activity of laccase is ≥10000 U / g; the activity of pullulanase is ≥2500 U / g; the enzyme activity of cellulase is ≥3500 U / g.

[0021] Preferably, the pH value range of the solution in step (2) is 9.5 - 10.5, and the reaction temperature is 40 - 45°C.

[0022] Preferably, the mass ratio of the humic acid to the mineral salt in step (2) is 4:1 - 2.8, and the stirring reaction time after adding the mineral salt is 1 - 2 h.

[0023] The reaction mechanism of the present invention is:

[0024] R-(COOH)n + nNaOH → R-(COONa) n + nH₂O

[0025] R-(COONa) n + nHCl → R-(COOH) n + nNaCl

[0026] R-(COOH) n → nR-COOH

[0027] R-COOH + NaOH → R-COONa

[0028] R-COONa + MCl → R-COOM + NaCl

[0029] Humic acid is a macromolecular organic mixture that is easily soluble in alkali, difficult to dissolve in water, and insoluble in acid. The extraction of humic acid is based on its solubility, using an alkali solution for extraction and then precipitating with acid.

[0030] Biological enzymes act as catalysts to initiate and accelerate chemical reactions. They can break open large chemical chains and quickly convert various macromolecules into small organic molecules. Through the process of biological enzyme catalysis, the processing time is shortened, the reaction temperature is reduced, the oxidation rate of metals is slowed down, and the complexation of small molecules and mineral salts is accelerated.

[0031] The technical effects of the present invention are as follows:

[0032] 1. Traditional chemical synthesis methods have many steps, many by-products, and are difficult to purify the products. Generally, they can only meet the requirements of small-scale laboratory preparation and are not suitable for industrial production. The biological enzyme method adopted by the present invention has the advantages of high production efficiency, environmental friendliness, and safety to the human body. Moreover, weathered coal, which is cheap and easily available, is used as a raw material to prepare humic acid, reducing the production cost.

[0033] 2. The charge within the humic acid complex molecule of the present invention tends to be neutral, is easily absorbed under the pH environment of the digestive tract, has good stability, and high biological utilization rate. Using the humic acid complex to replace inorganic trace elements can reduce the emission of trace elements in the environment while ensuring the normal production performance and health of animals, improving environmental pollution and being more healthy and environmentally friendly.

[0034] 3. During the preparation process of the humic acid complex of the present invention, the mother liquor after centrifugal filtration is pumped into the reaction vessel to continue to be used as the reaction solution for recycling. This is a synthetic path of green chemistry, which reduces the production cost and improves the production efficiency at the same time. Brief Description of the Drawings

[0035] Figure 1 It is a process flow chart for the production of the humic acid complex of the present invention;

[0036] Figure 2 Infrared absorption spectrum of humic acid in the embodiment of the present invention;

[0037] Figure 3 Infrared absorption spectrum of copper humate prepared in Example 1 of the present invention;

[0038] Figure 4 Infrared absorption spectrum of iron humate prepared in Example 2 of the present invention;

[0039] Figure 5 Infrared absorption spectrum of zinc humate prepared in Example 3 of the present invention;

[0040] Figure 6 Infrared absorption spectrum of manganese humate prepared in Example 4 of the present invention;

[0041] Figure 7 Infrared absorption spectrum of chromium humate prepared in Example 5 of the present invention;

[0042] Figure 8 Infrared absorption spectrum of cobalt humate prepared in Example 6 of the present invention. Detailed implementation manners

[0043] The technical solutions of the present invention will be clearly and completely described below in conjunction with specific embodiments of the present invention. Obviously, the described embodiments are only a part of the embodiments of the present invention, rather than all of the embodiments. All other embodiments obtained by those of ordinary skill in the art based on the embodiments of the present invention without creative efforts shall fall within the protection scope of the present invention.

[0044] Synthesis of diethylamine nitrate ionic liquid: Slowly add 1 mol of nitric acid dropwise to a reaction vessel containing 1.2 mol of diethylamine, react at 50 °C for 5 h to obtain a yellow liquid. After the reaction is completed, unreacted diethylamine and a small amount of water are removed by rotary evaporation at 80 °C, and the ionic liquid is obtained by vacuum drying. The preparation of diethylamine acetate ionic liquid and diethylamine formate ionic liquid refers to the preparation of diethylamine nitrate ionic liquid.

[0045] Example 1: Preparation of copper humate complex for feed

[0046] (1) Extraction of macromolecular humic acid from weathered coal

[0047] Put 596 kg of weathered coal with a total humic acid content of 55% into 2000 L of water, heat up to 90 °C, add sodium hydroxide to adjust the pH of the solution to 9, dissolve the weathered coal for 2 h, filter off the insoluble matter to obtain a refined humic acid extract solution, concentrate the solution until the solid content in the feed liquid is 18%; add concentrated hydrochloric acid to the concentrated solution to adjust the pH to 3.5 to obtain a macromolecular humic acid feed liquid;

[0048] (2) Degradation and complexation of macromolecular humic acid by bio - enzymes

[0049] In step (1), the feed liquid is adjusted to pH 10 with sodium hydroxide and the temperature is 40 °C. Then, a tannin acyl hydrolase solution (150 g of tannin acyl hydrolase dissolved in 500 g of diethylamine acetate ionic liquid) is added, and an enzymatic reaction is carried out for 1 h. Then, 111 kg of copper chloride is added and stirred and mixed evenly for 1 h to obtain a copper humate complex solution;

[0050] (3) Cooling crystallization of humic acid complex

[0051] The reaction liquid is pumped into a crystallization kettle, the stirring paddle and cooling water are turned on. After stirring and crystallizing for 5 h, centrifugal separation is carried out, and drying gives copper humate complex with a copper content of 13.81% and a yield of 95.42%.

[0052] The infrared absorption spectrum of the copper humate prepared in Example 1 is as Figure 3 shown. Compared with the infrared spectrum of humic acid Figure 2 , the characteristic absorption peak of OH at 3427.93 cm -1 of humic acid disappears, indicating that the atoms in humic acid participate in the coordination reaction, and humic acid does undergo a complexation reaction with copper.

[0053] Example 2: Preparation of iron humate complex for feed

[0054] (1) Extraction of macromolecular humic acid from weathered coal

[0055] 656 kg of weathered coal with a total humic acid content of 50% is put into 2000 L of water, the temperature is raised to 90 °C, and the pH of the solution is adjusted to 9 with sodium hydroxide. The weathered coal is dissolved for 2 h, and the insoluble substances are filtered off to obtain a refined humic acid extract. The solution is concentrated until the solid content in the feed liquid is 19%. Concentrated hydrochloric acid is added to the concentrated solution to adjust the pH to 3.8 to obtain a macromolecular humic acid feed liquid;

[0056] (2) Degradation and complexation of macromolecular humic acid by bio - enzymes

[0057] In step (1), the feed liquid is adjusted to pH 10 with sodium hydroxide and the temperature is 40 °C. Then, a bio - composite enzyme solution (100 g of tannin acyl hydrolase and 50 g of pullulanase dissolved in 800 g of diethylamine nitrate ionic liquid) is added, and an enzymatic reaction is carried out for 1 h. Then, 228 kg of ferrous sulfate is added and stirred and mixed evenly for 2 h to obtain an iron humate complex solution;

[0058] (3) Cooling crystallization of humic acid complex

[0059] The reaction liquid is fed into the crystallization kettle. The stirring paddle and cooling water are turned on. After stirring and crystallizing for 5 h, centrifugal separation is carried out, and then drying is performed to obtain the iron humate complex. The iron content is 11.95%, and the yield is 97.36%.

[0060] The infrared absorption spectrum of the iron humate prepared in Example 2 is as Figure 4 shown. Compared with the infrared spectrum of humic acid Figure 2 , the characteristic absorption peak of OH at 3427.93 cm -1 of humic acid disappears, indicating that the atoms in humic acid participate in the coordination reaction, and humic acid indeed undergoes a complexation reaction with iron.

[0061] Example 3: Preparation of zinc humate complex for feed

[0062] (1) Extracting macromolecular humic acid from weathered coal

[0063] 534 kg of weathered coal with a total humic acid content of 60% is put into 2000 L of water. The temperature is raised to 90 °C, and sodium hydroxide is added to adjust the pH of the solution to 9. The weathered coal is dissolved for 2 h, and the insoluble matter is filtered off to obtain a refined humic acid extract. The solution is concentrated until the solid content in the feed liquid is 20%. Concentrated hydrochloric acid is added to the concentrated solution to adjust the pH to 4 to obtain a macromolecular humic acid solution;

[0064] (2) Degrading and complexing macromolecular humic acid with bio-enzyme

[0065] The feed liquid from step (1) is adjusted to a pH of 10.5 and a temperature of 35 °C with sodium hydroxide, and a phytase solution (250 g of phytase dissolved in 2500 g of diethylamine formate ionic liquid) is added for an enzymatic reaction for 1 h. 143 kg of zinc sulfate is added, and the mixture is stirred evenly and reacted for 2 h to obtain a zinc humate complex feed liquid;

[0066] (3) Cooling crystallization of the humic acid complex

[0067] The reaction liquid is fed into the crystallization kettle. The stirring paddle and cooling water are turned on. After stirring and crystallizing for 5 h, centrifugal separation is carried out, and then drying is performed to obtain the zinc humate complex. The zinc content is 13.95%, and the yield is 92.5%.

[0068] The infrared absorption spectrum of the zinc humate prepared in Example 3 is as Figure 5 shown. Compared with the infrared spectrum of humic acid Figure 2 , the characteristic absorption peak of OH at 3427.93 cm -1 of humic acid disappears, indicating that the atoms in humic acid participate in the coordination reaction, and humic acid indeed undergoes a complexation reaction with zinc.

[0069] Example 4: Preparation of manganese humate complex for feed

[0070] (1) Extraction of macromolecular humic acid from weathered coal

[0071] Put 534 kg of weathered coal with a total humic acid content of 60% into 2000 L of water, heat it to 85 °C, add sodium hydroxide to adjust the pH of the solution to 10.5, dissolve the weathered coal for 2 h, filter off the insoluble matter to obtain a refined humic acid extract solution, concentrate the solution until the solid content in the feed liquid is 18%; add hydrochloric acid to the concentrated solution to adjust the pH to 3.5 to obtain a macromolecular humic acid feed liquid;

[0072] (2) Degradation and complexation of macromolecular humic acid by bioenzymes

[0073] Adjust the pH of the feed liquid in step (1) to 10.5 and the temperature to 50 °C with sodium hydroxide, add a bio - composite enzyme solution (100 g of phytase and 150 g of tannin acyl hydrolase dissolved in 1500 g of a mixed ionic liquid of diethylamine formate and diethylamine acetate (the mass ratio of the two ionic liquids is 1:1)), carry out an enzymatic reaction for 1 h, add 101 kg of manganese chloride, stir and mix evenly, and react for 2 h to obtain a humic acid - manganese complex solution;

[0074] (3) Cooling crystallization of humic acid complex

[0075] Transfer the reaction liquid to a crystallization kettle, turn on the stirring paddle and cooling water, stir and crystallize for 5 h, then carry out centrifugal separation and drying to obtain a humic acid - manganese complex, with a manganese content of 12.09% and a yield of 90.5%.

[0076] The infrared absorption spectrum of the humic acid - manganese prepared in Example 4 is as Figure 6 shown. Compared with the infrared spectrum of humic acid Figure 2 the characteristic absorption peak of OH at 3427.93 cm -1 of humic acid disappears, indicating that the atoms in humic acid participate in the coordination reaction, and humic acid does undergo a complexation reaction with manganese.

[0077] Example 5: Preparation of humic acid - chromium complex for feed

[0078] (1) Extraction of macromolecular humic acid from weathered coal

[0079] Put 526 kg of weathered coal with a total humic acid content of 57% into 2000 L of water, heat it to 90 °C, add sodium hydroxide to adjust the pH of the solution to 10, dissolve the weathered coal for 2 h, filter off the insoluble matter to obtain a refined humic acid extract solution, concentrate the solution until the solid content in the feed liquid is 18%; add concentrated hydrochloric acid to the concentrated solution to adjust the pH to 3.5 to obtain a macromolecular humic acid feed liquid;

[0080] (2) Degradation and complexation of macromolecular humic acid by bioenzymes

[0081] Step (1): Adjust the pH of the solution to 11 and the temperature to 40 °C by adding sodium hydroxide to the feed liquid. Add a bio - composite enzyme solution (100 g of phytase and 150 g of cellulase dissolved in 2000 g of diethylamine acetate ionic liquid), carry out an enzymatic reaction for 1 h, add 200 kg of chromium chloride, stir and mix evenly, and react for 1 h to obtain a humic acid - chromium complex solution;

[0082] (3) Cooling crystallization of the humic acid complex

[0083] Transfer the reaction liquid to a crystallization kettle, turn on the stirring paddle and cooling water, stir and crystallize for 5 h, then perform centrifugal separation and drying to obtain the humic acid - chromium complex. The chromium content is 7.98% and the yield is 91.4%.

[0084] The infrared absorption spectrum of the humic acid - chromium prepared in this Example 5 is as Figure 7 shown. Compared with the infrared spectrum of humic acid Figure 2 , the characteristic absorption peak of OH at 3427.93 cm -1 of humic acid disappears, indicating that the atoms in humic acid participate in the coordination reaction, and humic acid does undergo a complexation reaction with chromium.

[0085] Example 6: Preparation of humic acid - cobalt complex for feed

[0086] (1) Extraction of macromolecular humic acid from weathered coal

[0087] Put 508 kg of weathered coal with a total humic acid content of 65% into 2000 L of water, heat up to 90 °C, add sodium hydroxide to adjust the pH of the solution to 10, dissolve the weathered coal for 2 h, filter out the insoluble substances to obtain a refined humic acid extraction solution, concentrate the solution until the solid content in the feed liquid is 20%; add concentrated hydrochloric acid to the concentrated solution to adjust the pH to 3.5 to obtain a macromolecular humic acid solution;

[0088] (2) Degradation and complexation of macromolecular humic acid by bio - enzyme

[0089] In step (1), add sodium hydroxide to the feed liquid to adjust the pH of the solution to 11 and the temperature to 40 °C, add a cellulase solution (450 g of cellulase dissolved in 1250 g of diethylamine acetate ionic liquid), carry out an enzymatic reaction for 1 h, add 124 kg of cobalt sulfate, stir and mix evenly, and react for 1 h to obtain a humic acid - cobalt complex solution;

[0090] (3) Cooling crystallization of the humic acid complex

[0091] Transfer the reaction liquid to a crystallization kettle, turn on the stirring paddle and cooling water, stir and crystallize for 5 h, then perform centrifugal separation and drying to obtain the humic acid - cobalt complex. The cobalt content is 11.43% and the yield is 93.1%.

[0092] The infrared absorption spectrum of the humic acid - cobalt prepared in this Example 6 is asFigure 8 As shown, compared with the infrared spectrum of humic acid Figure 2 the characteristic absorption peak of OH at 3427.93 cm -1 disappeared, indicating that the atoms in humic acid participated in the coordination reaction, and humic acid did undergo a complexation reaction with cobalt.

[0093] Application and implementation test

[0094] 1. Experimental animals and experimental design

[0095] The experiment was conducted from April to June 2022 in Shandong Asia-Pacific Haihua Biotechnology Co., Ltd. The experimental animals were 240-day-old healthy Hailan brown-shell laying hens. The experimental diet was based on the "Chicken Feed Standard" NY / T 33 - 2004, and a compound diet for laying hens during the laying period was selected. Eighty 240-day-old laying hens after the peak laying period, with similar body weights, good health, and normal appetites, were divided into 4 groups, with 20 hens in each group. In the basal diet, 6 mg / kg Cu (CuSO4·5H2O), 75 mg / kg Fe (FeSO4·H2O), 60 mg / kg Zn (ZnSO4·H2O), and 60 mg / kg Mn (MnSO4·H2O) of feed-grade inorganic trace elements were added as the inorganic group, and the addition amounts referred to the trace element addition amounts in the premix produced by a large feed company. The experimental groups were divided into a group adding humic acid complex, a 70% humic acid complex group, and a 50% humic acid complex group, adding 100%, 70%, and 50% of the humic acid complex of the trace element addition amounts in the inorganic group, respectively. The pre-feeding period was 10 days, and the formal test period was 43 days. The chicken house was semi-open, and the hens were fed in three-tier stepped laying hen cages, with 5 hens in each cage. The feeding and management conditions of each group were the same. The hens were fed twice a day and the eggs were collected twice a day, with free access to feed and water. Deworming and immunization were carried out according to the routine management procedures of the chicken farm. During the pre-experiment period, all groups were fed the basal diet uniformly, and during the formal experiment period, each group was fed the diet supplemented with different doses of trace elements.

[0096] 2. Sources of experimental materials

[0097] The feed-grade inorganic trace elements were: CuSO4·5H2O, FeSO4·H2O, ZnSO4·H2O, MnSO4·H2O, purchased from the market, and the humic acid complex was developed by Shandong Asia-Pacific Haihua Biotechnology Co., Ltd. (Examples 1 - 4).

[0098] 3. Measurement methods and indicators

[0099] At 6:00 p.m. on the 14th, 28th, and 42nd days of the experiment, all eggs produced on the same day were collected, marked with the date and treatment group, and 10 eggs were randomly selected from each group according to the replicates for measuring egg quality on the same day. For egg quality, an EggShellThickness from Orka Technology was used to measure the eggshell thickness, an Egg Force Reader to measure the eggshell strength, and an Egg analyzer to measure the egg weight, yolk color, albumen height, and Haugh unit; Feces of laying hens were collected every day during the last 3 days of the experimental period and properly stored in a dark and dry place for subsequent determination of trace element content. The determination of trace elements in fecal samples was carried out by ICP-OES to measure the contents of copper, iron, zinc, and manganese in the feces.

[0100] 4. Experimental Results

[0101] The experimental results are shown in Table 1-2. It can be seen from the experimental results that compared with the addition amount of inorganic trace elements in the conventional laying hen premix, the humic acid complex has the characteristic of high absorption, and reducing the dosage of copper, iron, zinc, and manganese can also maintain the normal production of laying hens and egg quality; moreover, the combined use of humic acid complexes can significantly reduce the emission of trace elements in feces, and the trace elements are better utilized.

[0102] Table 1 Residual amounts of trace elements in feces under different treatments

[0103] Inorganic salt Humic acid complex 70% Humic acid complex 50% Humic acid complex Copper (μg / g) 61.56±8.12 48.37±6.95 31.99±8.14 24.71±3.15 Iron (μg / g) 470±35.12 450.53±14.66 420.36±18.64 398.94±25.22 Zinc (μg / g) 343±39.15 308.55±27.62 246.97±29.71 209.42±20.27 Manganese (μg / g) 259.68±18.56 217.71±10.59 182.12±15.58 148.79±12.55

[0104] Table 2 Comparison of laying rate and egg quality under different treatments

[0105] Inorganic salt Humic acid complex 70% Humic acid complex 50% Humic acid complex Egg production rate (%) 80.31±5.71 83.95±5.41 87.66±4.73 82.48±5.15 Eggshell specific gravity (%) 38.19±6.06 39.55±5.06 39.22±5.64 38.94±5.22 Eggshell thickness (mm) 0.410±0.027 0.421±0.036 0.418±0.033 0.042±0.054 Haugh unit 68.95±7.42 70.94±8.59 69.13±7.82 71.07±6.38 Yolk color 5.09±1.11 5.10±0.92 5.22±0.95 5.08±0.83

Claims

1. A method for synthesizing a humic acid complex for feed by a biological enzyme method, characterized in that, It includes the following steps: (1) Extracting macromolecular humic acid from weathered coal Put weathered coal and water into a reaction vessel, stir and heat up to 80 - 90 °C, adjust the pH to 9 - 11 with alkali to dissolve the weathered coal, filter out the insoluble substances to obtain a refined extract of humic acid, then concentrate it, and add acid to the concentrated solution to adjust the pH to 3.5 - 4 to obtain a macromolecular humic acid feed liquid; (2) Degrading and complexing macromolecular humic acid with bio - enzymes For the macromolecular humic acid feed liquid prepared in step (1), add sodium hydroxide to adjust the pH of the solution to 9 - 11.5, then add the bio - enzyme solution, carry out an enzymatic reaction at 35 - 50 °C for 0.5 - 1.5 h, then add mineral salts, stir and react for 1 - 4 h to obtain a humic acid complex solution; The bio - enzyme solution is prepared as follows: Dissolve the bio - enzyme in a quaternary ammonium ionic liquid, and after complete dissolution, pre - cool it in an environment of 1 - 5 °C for standby; The bio - enzyme is one or more of tannin acyl hydrolase, phytase, and laccase; The quaternary ammonium ionic liquid is one or more of diethylamine acetate ionic liquid, diethylamine nitrate ionic liquid, and diethylamine formate ionic liquid; For the mineral salts, the mineral is copper, iron, zinc, manganese, chromium, or cobalt; The salt is one or more of chloride, sulfate, nitrate, and acetate; The mass ratio of humic acid to mineral salts is 4:1 - 2.8; (3) Cooling crystallization of humic acid complex Cool the humic acid complex solution prepared in step (2) for crystallization, carry out centrifugal separation, and dry it to obtain the humic acid complex.

2. The method for synthesizing a humic acid complex for feed by a bioenzymatic method according to claim 1, characterized in that, The mass ratio of the bio - enzyme to the ionic liquid is 1:2 - 10.

3. The method for synthesizing a humic acid complex for feed by a biological enzyme method as described in claim 1, characterized in that, The mother liquor after centrifugal separation in step (3) is pumped into the reaction vessel in step (1) to continue to be used as the reaction liquid for recycling.

4. The method for synthesizing a humic acid complex for feed by a biological enzyme method according to claim 1, characterized in that, The total humic acid content in the weathered coal in step (1) is 45% - 65%.

5. The method for synthesizing a humic acid complex for feed by a biological enzyme method according to claim 1, characterized in that, In step (1), the solution is concentrated to a solid content of 18% - 20% in the feed liquid.

6. The method for synthesizing a humic acid complex for feed by a biological enzyme method as described in claim 1, characterized in that, The addition amount of the bio - enzyme in step (2) is 0.01% - 0.5% of the mass of the weathered coal.

7. A method for synthesizing a humic acid complex for feed by a biological enzyme method according to any one of claims 1-6, characterized in that, In step (2), add sodium hydroxide to adjust the pH of the solution to 9.5 - 10.5, and the enzymatic reaction temperature is 40 - 45 °C.

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