Preparation method and application of copper protein

Abstract

The invention relates to a preparation method and application of copper protein, and belongs to the technical field of novel fertilizer synergists, and the preparation method comprises the following steps: S1, taking a copper-containing substance as a raw material or a mixture of the copper-containing substance and a zinc-containing substance as a raw material, dissolving the raw material in ammonia water at low temperature to obtain a reaction solution, and concentrating to obtain a protein synthesis solution; s2, mixing polysuccinimide with water, and uniformly stirring to obtain a protein base solution; and S3, dropwise adding the protein synthesis solution in the step S1 into the protein base solution in the step S2, and reacting to obtain the copper protein. The copper protein is obtained by the method aiming at the phenomenon that the effect is poor in the copper supplementing process, fertilizer damage is easily caused or the photosynthesis capacity of gramineous crops is poor, and consequently nutrient transfer is slow. According to the copper-containing protein obtained from the copper protein, structural instability or isomerization caused by addition of other substances is reduced in the composition, the method is simple, convenient and easy to implement, the copper-containing protein with stable composition is easily obtained, and therefore the real effect of the copper-containing protein is achieved.

Description

TECHNICAL FIELD

[0001] The application belongs to the technical field of new fertilizer synergists, and particularly relates to a preparation method and application of a copper protein. BACKGROUND

[0002] Copper in the soil, in addition to that brought by the parent material, is mainly brought by human activities, such as the application of fungicides, the comprehensive utilization of breeding waste and the use of trace element fertilizers; however, with the release of the Action Plan for Prevention and Control of Soil Pollution and the emergence of modern pesticides fungicides, the content of copper element in the soil in China has the following phenomena:

[0003] 1. Copper compounds (such as copper sulfate and copper hydroxide) as the original broad-spectrum fungicides are gradually replaced by synthetic organic fungicides due to their unstable effects and frequent phytotoxicity, resulting in a sharp decline in the accumulation of copper elements in the soil.

[0004] 2. The comprehensive utilization of breeding waste makes the originally directly used livestock manure safe after removing heavy metals, antibiotics and fermentation, which greatly improves the safety, but cannot effectively supplement the copper content in the soil, thereby causing potential harm to the yield and human health.

[0005] Copper is an essential trace element for crop growth and plays an important role in plant physiological processes; when crops lack copper, problems such as slow growth, leaf discoloration and decreased disease resistance occur, which is due to the lack of copper protein.

[0006] Copper protein is a kind of protein containing copper ions, which widely exists in organisms and participates in various important biological processes. Copper ions usually act as cofactors in related proteins such as cytochrome c oxidase, superoxide dismutase, multicopper oxidase and copper-zinc superoxide dismutase, and participate in key biochemical reactions such as electron transfer and redox reaction. The role of copper ions in copper protein mainly reflects its redox activity, which can switch between different Cu + and Cu 2+ . However, free copper ions have strong toxicity to cells, so crops will precisely control the utilization of copper ions through the regulation of copper chaperone and copper transporter.

[0007] In view of the above situation, the technical personnel of the company obtained the related protein by compounding copper sulfate chelation on the basis of polyaspartic acid, but the existence of sulfate in the protein structure caused great instability and isomerism, so that it cannot play the excellent effect of copper protein. Therefore, how to synthesize a copper protein that meets the application of crops and does not cause harm and can play its effect by using the structure and performance of polyaspartic acid has become a problem to be solved. SUMMARY

[0008] The present application aims at the poor effect in the use of copper protein, easy to cause fertilizer harm, and easy to appear dissociation phenomenon in the storage process, applies polyaspartic acid as the excellent adaptability and chemical stability of the artificial synthesis protein, selects poly succinimide one-step hydrolysis chelation method to synthesize related protein, reduces the structure instability or isomerization caused by the addition of other substances, so as to play its true effect.

[0009] The technical scheme adopted by the present application to achieve the purpose is:

[0010] A preparation method of copper protein, comprising the following steps:

[0011] S1. Using copper-containing material as raw material, or using a mixture of copper-containing material and zinc-containing material as raw material, dissolving the raw material in ammonia water at low temperature to obtain a reaction solution, and concentrating to obtain a protein synthesis solution;

[0012] S2. Mixing poly succinimide with water, stirring uniformly, and dispersing into particles with particle size <10 μm to obtain a protein base solution;

[0013] S3. Dropping the protein synthesis solution of step S1 into the protein base solution of step S2, and reacting at 40-50℃ for 1-3h to obtain copper protein.

[0014] Preferably, the copper-containing material is one or both of cupric oxide and cupric hydroxide; the zinc-containing material is zinc oxide or zinc hydroxide, and the mass ratio of copper to zinc is 1:(0.01-0.2).

[0015] Preferably, the purity of cupric oxide is ≥99%, the purity of cupric hydroxide is ≥99%, the purity of zinc oxide is ≥99.5%, the purity of zinc hydroxide is ≥99.5%, and the mass concentration of ammonia water is 25% (calculated as NH3, the same below).

[0016] Preferably, the molar ratio of raw material to ammonia water is 1:(6.0-6.5), and the molar ratio of the above substances is changed to 1:(4.0-4.2) after concentration in the protein synthesis solution stage.

[0017] Preferably, the reaction temperature of the reaction solution is controlled at ≤40℃; and the concentration temperature of the protein synthesis solution is 55-65℃.

[0018] Preferably, the mass ratio of poly succinimide to water is 1:(1-2), and the dispersion is carried out in two stages, the first stage is the coarse mixing of the material, the stirring speed is 100-200 r / min, and the duration is 10-15 min; the second stage is the grinding stage of the material, the stirring speed is 3000-4500 r / min, and the stirring time is 30-45 min, usually water cooling is needed to control the material temperature not to exceed 65℃.

[0019] Preferably, the time for adding the protein synthesis solution to the protein base solution in step S3 is controlled at 15-25 minutes. Too long a time will cause fluctuations in the ammonia ratio and affect the subsequent results. The mass ratio of the protein synthesis solution to the protein base solution is 1:(2-4).

[0020] A fertilizer enhancer comprising the copper protein prepared according to any one of claims 1-7.

[0021] When the copper protein is added to nitrogen-containing fertilizers, and the nitrogen-containing fertilizer is one or any combination of urea, ammonium sulfate, ammonium chloride, and ammonium carbonate, the amount of copper protein added is 4.5-6.5 kg / T of fertilizer; for other nitrogen-containing fertilizers, the amount of copper protein additive is 10-25 kg / T of fertilizer.

[0022] Copper-containing protein fertilizers are applied during the cultivation of gramineous crops.

[0023] The beneficial effects of this invention are:

[0024] This invention provides a method for preparing copper protein, using polysuccinimide as the main raw material to obtain a superior polyaspartic acid derivative. This method avoids the harmful effects of free copper / zinc on crops, reducing problems such as slow growth, leaf chlorosis, and decreased disease resistance / stress. It also avoids the instability, high structural isomerism, and inability to fully realize the intended effects of copper protein. The resulting product is a readily available polyaspartic acid-based copper protein with outstanding effects. This protein demonstrates remarkable performance in supplementing copper / zinc and regulating crop physiological and biochemical processes. Attached Figure Description

[0025] Figure 1 This is the FTIR spectrum of the copper-containing synthetic protein produced in Example 1.

[0026] Figure 2 This is the FTIR spectrum of the copper-containing comparative protein I produced in Comparative Example 1. Detailed Implementation

[0027] The present invention will now be described in detail with reference to specific embodiments. Obviously, the described embodiments are only a part of the embodiments of the present invention, not all of them. All other embodiments obtained by those skilled in the art based on the embodiments of the present invention without creative effort should fall within the scope of protection of the present invention. Any aspects not described herein are prior art.

[0028] Example 1

[0029] Preparation method of copper ion-containing synthetic protein I:

[0030] Dissolve 20g of 99% copper monoxide in 105.19g of 25% ammonia water. After dissolving at low temperature, it is designated as reaction solution I. The reaction temperature of the reaction solution is controlled at 35-38℃. The reaction solution is concentrated to obtain 90.76g of protein synthesis solution I. The temperature at which the concentration is converted into protein synthesis solution is 59-61℃.

[0031] 123.70g of polysuccinimide was mixed with 185.55g of water and stirred continuously for 15min at a stirring speed of 150r / min; then stirred for 40min at a stirring speed of 4000r / min. The entire stirring process was water-cooled to control the temperature of the material to not exceed 65℃, thus obtaining protein base liquid I.

[0032] Protein synthesis solution I was added dropwise to protein base solution I, and the reaction time was controlled at 20 min; after reacting at 45℃ for 2 h, copper-containing synthetic protein I was obtained.

[0033] Example 2

[0034] Preparation method of copper ion-containing synthetic protein II:

[0035] Dissolve 20g of 99% copper monoxide in 100.98g of 25% ammonia water. After dissolving at low temperature, it is designated as reaction solution II. The reaction temperature of the reaction solution is controlled at 35-38℃. The reaction solution is concentrated to obtain 90.76g of protein synthesis solution II. The concentration temperature for converting the protein synthesis solution to protein synthesis solution is 59-61℃.

[0036] 113.45g of polysuccinimide was mixed with 113.45g of water and stirred continuously for 15 minutes at a stirring speed of 100r / min. Then, the mixture was stirred for 30 minutes at a stirring speed of 4500r / min. The mixture was cooled with water throughout the stirring process to keep the temperature of the mixture below 65℃, thus obtaining protein base liquid II.

[0037] Protein synthesis solution II was added dropwise to protein base solution II for 15 minutes; after reacting at 40°C for 3 hours, copper-containing synthetic protein II was obtained.

[0038] Example 3

[0039] Preparation method of copper ion-containing synthetic protein III:

[0040] Dissolve 20g of 99% copper monoxide in 109.40g of 25% ammonia water. After dissolving at low temperature, it is designated as reaction solution III. The reaction temperature of the reaction solution is controlled at 35-38℃. The reaction solution is concentrated to obtain 90.76g of protein synthesis solution III. The concentration temperature for converting the protein synthesis solution to protein synthesis solution is 59-61℃.

[0041] 121.01g of polysuccinimide was mixed with 242.02g of water and stirred continuously for 10 minutes at a stirring speed of 200r / min. Then, the mixture was stirred for 45 minutes at a stirring speed of 3000r / min. The mixture was cooled with water throughout the stirring process to keep the temperature of the mixture below 65℃, thus obtaining protein base liquid III.

[0042] Protein synthesis solution III was added dropwise to protein base solution III, and the reaction time was controlled at 25 min; after reacting at 50℃ for 1 h, copper-containing synthetic protein III was obtained.

[0043] Example 4

[0044] Preparation method of copper ion-containing synthetic protein IV:

[0045] 24.14 g of 99% copper hydroxide and 0.20 g of 99.5% zinc oxide were dissolved in 105.19 g of 25% ammonia water and kept at low temperature for later use. This solution was designated as reaction solution IV. The remaining conditions were the same as in Example 1 to obtain copper ion-containing synthetic protein IV.

[0046] Example 5

[0047] Preparation method of copper ion-containing synthetic protein V:

[0048] 22.27g of 99% copper hydroxide and 1.83g of 99.5% zinc oxide were dissolved in 105.19g of 25% ammonia water and kept at low temperature for later use. This solution was designated as reaction solution V. The remaining conditions were the same as in Example 1 to obtain copper ion-containing synthetic protein V.

[0049] Example 6

[0050] Preparation method of copper ion-containing synthetic protein VI:

[0051] 20.42g of 99% copper hydroxide and 4.10g of 99.5% zinc hydroxide were dissolved in 105.19g of 25% ammonia water and kept at low temperature for later use. This solution was designated as reaction solution VI. The remaining conditions were the same as in Example 1 to obtain copper ion-containing synthetic protein VI.

[0052] Comparative Example 1: Polyaspartic acid reacts directly with copper sulfate

[0053] Preparation method of copper-containing contrast protein I:

[0054] Dissolve 63.16g of 98% copper sulfate pentahydrate in 37.90g of water to obtain control protein reaction solution I;

[0055] 416.54g of 40% ammonium polyaspartate was stirred at a speed of 4000r / min for 40min. The entire stirring process was water-cooled to control the temperature of the material to not exceed 65℃, thus obtaining the control protein base liquid I.

[0056] Comparative protein synthesis solution I was added dropwise to comparative protein base solution I, and the reaction time was controlled at 20 min; after reacting at 45℃ for 2 h, copper-containing comparative protein I was obtained.

[0057] Comparative Example 2 exceeded the controlled reaction temperature

[0058] Preparation method of copper-containing contrast protein II:

[0059] Dissolve 20g of 99% copper monoxide in 105.19g of 25% ammonia water, and set aside as control reaction solution II. The reaction temperature of the solution is controlled at 55-57℃. The reaction solution is concentrated to obtain 90.76g of control protein synthesis solution II. The concentration temperature is 65-70℃.

[0060] The conditions for generating the contrast protein base solution II were the same as those in Example 1;

[0061] The contrast protein synthesis solution II was added dropwise to the contrast protein base solution II, and the reaction time was controlled at 20 min; after reacting at 75℃ for 2 h, the contrast protein II containing copper ions was obtained.

[0062] Comparative Example 3 was not concentrated; the material ratio was outside the range.

[0063] Preparation method of copper-containing contrast protein III:

[0064] Dissolve 20g of 99% copper monoxide in 105.19g of 25% ammonia water, and set aside at low temperature to obtain 125.19g of protein synthesis solution III. The reaction temperature of the reaction solution is controlled at 35-38℃.

[0065] The conditions for generating the contrast protein base solution III were the same as those in Example 1;

[0066] The contrast protein synthesis solution III was added dropwise to the contrast protein base solution III, and the reaction conditions were the same as in Example 1, to obtain the contrast protein III containing copper ions.

[0067] Comparative Example 4: Protein synthesis conditions were not controlled; the two solutions were directly mixed.

[0068] Preparation method of copper-containing contrast protein IV:

[0069] The preparation conditions for the control protein synthesis solution IV and the control protein base solution IV were the same as in Example 1;

[0070] After pouring the contrast protein synthesis solution IV into the contrast protein base solution IV and stirring evenly, copper-containing contrast protein IV was obtained.

[0071] Comparative Example 5: Long-term stirring reaction

[0072] Preparation method of copper ion-containing contrast protein V:

[0073] The preparation conditions for the control protein synthesis solution V and the control protein base solution V were the same as in Example 1;

[0074] The contrast protein synthesis solution V and the contrast protein base solution V were reacted at the rate of Example 1, and then stirred continuously for 6 hours to obtain contrast protein V containing copper ions.

[0075] Comparative Example 6 without two-stage mixing

[0076] Preparation method of copper ion-containing contrast protein VI:

[0077] The protein synthesis solution VI is the same as that in Example 1;

[0078] The material ratio of the comparative protein base liquid VI was the same as that of Example 1; the difference was that the stirring speed was 150 r / min, the duration was 15 min, and the temperature of the material was controlled not to exceed 65°C to obtain the comparative protein base liquid VI.

[0079] The contrast protein synthesis solution VI was added dropwise to the contrast protein base solution VI, and the reaction conditions were the same as in Example 1, to obtain contrast protein VI containing copper ions.

[0080] Experimental Example 1

[0081] The copper-containing synthetic proteins prepared in Examples 1 to 3 (referred to as T1 to T3) and Comparative Examples 1 to 6 (referred to as CK1 to CK6) were used as test samples. Meanwhile, the treatment without the above proteins but with copper ion supplementation was used as CK. Under the same nutrient conditions, indoor potted plant experiments were conducted to test the advantages of the present invention under the condition of copper ion deficiency followed by supplementation.

[0082] This experiment was conducted in the chemical laboratory of Hebei Xietong Chemical Co., Ltd. 50-mesh vermiculite was used as the culture medium and Jimai 526 was used as the experimental crop. Each treatment was replicated in 3 replicates and 6 pots were used for each replicate treatment. At the same time, the direct seeding method was adopted and 25 wheat seeds were planted in each pot. During the seedling stage, the moisture content of the culture medium was maintained at about 50% and was adjusted to about 25% of the soil moisture content in the later stage of seedling emergence (as a mild drought).

[0083] To verify the effectiveness of the treatments, wheat seedlings were harvested 7 days after each treatment was applied. The fresh weight of the winter wheat seedlings was determined by weighing, and the dry weight was determined by drying and weighing as an indicator of growth rate. Chlorophyll content was measured using a portable SPAD502 chlorophyll meter as an indicator of leaf chlorosis. Enzyme activities (superoxide dismutase (SOD) and proline) were used as indicators of copper protein regulation of crop disease resistance and physiological and biochemical properties, respectively. SOD activity was measured using the xanthine oxidase-cytochrome c method (McCord & Fridovich method), and proline activity was measured using the hydroxylamine colorimetric method. Data were statistically analyzed using EXCEL, and one-way ANOVA was performed using SPSS 22.0, with Duncan's test for significance. Specific results are shown in the table below.

[0084] Table 1. Test results for different copper proteins

[0085]

[0086] The data in the table above show that in the indoor experiment, the photosynthesis of wheat treated with T1-T3 was improved (referring to chlorophyll content data) and the photosynthetic nutrient translocation rate (referring to dry weight data) was also improved. Compared with CK5, which performed best among the other control treatments, the increases were 6.57-10.35% (chlorophyll content) and 8.39-10.77% (dry weight), respectively, which were significant. This indicates that the T1-T3 treatment can significantly improve photosynthetic capacity and photosynthetic nutrient translocation rate, and reduce the rate of leaf chlorosis. In addition to the above indicators, the data on superoxide dismutase and proline also show that the three copper proteins can improve the disease resistance and drought resistance of wheat. Specifically, the increases in superoxide dismutase and proline-related data (T1-T3 treatment) compared with CK5, which performed best among the other control treatments (CK1-CK6, CK), were 15.89-18.96% and 32.30-40.76%, respectively.

[0087] Experimental Example 2

[0088] In 2024, a field experiment was conducted in Jinzhou City, Shijiazhuang City, Hebei Province (N37°95′55″, E115°09′84″). The experimental area was a typical brown soil with soil organic matter content of 1.93%, total nitrogen of 1.23 g / kg, available phosphorus of 31.57 mg / kg, available potassium of 117.23 mg / kg, and available copper of 1.2 mg / kg. It was a plot of land with low organic matter, medium to low copper content, and medium nitrogen, phosphorus, and potassium levels. The area had low rainfall and had experienced severe development of diseases such as Fusarium head blight during previous plantings, which met the requirements for experimental land.

[0089] This experiment consisted of 10 treatments, each with one concentration, with three replicates. Each plot was 150 m² in area. 2 The samples were randomly distributed across different plots. Synthetic copper proteins prepared in Examples 1-3 (referred to as T1-T3 treatments) and control copper proteins prepared in Comparative Examples 1-6 (referred to as CK1-CK6 treatments) were used as samples and applied via foliar spray at a concentration of 100 g / 667 m³. 2 Meanwhile, a blank control group was set up, which did not use copper protein and was sprayed with water only, and was recorded as CK; the application was carried out during the greening and jointing stage of winter wheat.

[0090] In the experiment, yield, effective spike number, thousand-grain weight, and number of grains per spike were designed and analyzed according to the relevant requirements of general agronomy. Superoxide dismutase, peroxidase, and proline, which are related to drought resistance, were measured using the nitroblue tetrazolium (NBT) photochemical reduction method, the guaiacol method, and the sulfosalicylic acid-ninhydrin method, respectively (the measurement period was before the next irrigation after the greening irrigation, during the drought stage of winter wheat). Data were statistically analyzed using EXCEL, and one-way ANOVA was performed using SPSS 22.0 data analysis system. The Duncan method was used to test for significance. The results are shown in Table 2.

[0091] Table 2. Effects of different copper protein sprays on winter wheat yield and drought resistance.

[0092]

[0093]

[0094] As shown in the table above, the copper protein obtained by spraying this solution can effectively increase crop yield and has a good effect on crop drought resistance. Firstly, in terms of yield, the T1-T3 treatments were significantly higher than other treatments, with yield increases ranging from 8.48% to 9.62% compared to the control. Analysis of yield components, including effective panicle number, thousand-grain weight, and number of grains per panicle, revealed that the main reasons for the yield increase were the increase in effective panicle number (6.83%-7.93%) and thousand-grain weight (6.39%-6.71%). The effective panicle number is related to crop tillering, primarily affecting the jointing stage. While polyaspartic acid (PAA) has a similar function, the control treatment did not show similar results. Therefore, the technicians concluded that copper protein as a whole promoted crop tillering, not just PAA. Conversely, simply adding PAA and copper ions would denature PAA and render it ineffective. The increase in thousand-grain weight further demonstrates that copper protein promotes photosynthesis and nutrient transport, thus increasing thousand-grain weight. Secondly, regarding drought resistance, the data from the three groups of superoxide dismutase, peroxidase, and proline showed a similar trend, namely, the T1-T3 treatment was significantly higher than other treatments. From this perspective, it is precisely because of the improved drought resistance that the crop's water use efficiency is improved, which leads to better replenishment of the thousand-grain weight and thus increases the yield.

[0095] Experimental Example 3

[0096] Examples 1-3 and Comparative Examples 1-6 were applied to a field maize (Zhengdan 958) planting experiment. The experimental method is as follows: Following fertilization practices, topdressing was applied during the maize experiment. The urea synergists obtained in the above examples and comparative examples were coated onto the surface of urea at an addition rate of 5.5 kg / T. To ensure uniform treatment, food additive tartrazine 60 was also coated onto the surface at an addition rate of 60 g / T urea. These treatments were designated as T1-3 and CK1-6, respectively. The total nitrogen content of the obtained products was 46.0%. To eliminate interference from tartrazine 60, a tartrazine + urea treatment was also set up, with the same treatment method as above, designated as CK7. The total nitrogen content of the obtained product was 46.1%. Finally, a blank urea treatment was set up, designated as CK, with a total nitrogen content of 46.2%. In the maize experiment, urea was applied alone at a rate of 20 kg / 667 m². 2 .

[0097] This experiment was conducted in 2024 in Zanhuang County, Shijiazhuang City, Hebei Province (114°52′E, 37°69′N). The selected plots had an organic matter content of 2.70-3.00 g / kg, and were classified as low-clay, high-sand brown soil. The soil contained 1.16 g / kg total nitrogen, 30.14 mg / kg available phosphorus, 122.69 mg / kg available potassium, and 1.4 mg / kg available copper. Prior to the experiment, head smut had occurred annually. During the experiment, all plant protection, horticulture, and water management were carried out by the homeowner on the same day, reducing [potential risks]. To minimize experimental errors caused by a few related factors, this experiment comprehensively calculated the number of grains per ear by statistically analyzing yield, thousand-grain weight, and number of ears per hectare (converted from plots). Simultaneously, the incidence of head smut was statistically analyzed to characterize disease resistance. One-way ANOVA was performed using SPSS 22.0 data analysis system, and the Duncan method was used for significance testing. All indicators were measured according to the Plant Biology Experiment Guide (2020), and the number of grains per ear and seed setting rate were calculated. The experiment was designed with four replicates per treatment, each replicate being 300m². 2 The experiment was conducted using a randomized block distribution, and the experimental results are shown in the table below.

[0098] Table 3 Effects of different copper proteins on maize seed setting rate

[0099]

[0100] The data in the table above shows that, in terms of yield, the TI-T3 treatment showed a significant difference compared to the other treatments, indicating that the copper protein produced by this scheme can significantly increase corn yield under the given production conditions. The reasons for this yield increase can be analyzed from the remaining data: Regarding thousand-grain weight, the TI-T3 treatment increased by 9.98-12.37%, indicating that the copper protein obtained by this scheme effectively promotes crop photosynthesis and nutrient transport, thus effectively increasing thousand-grain weight. Other treatments also showed some increase, such as CK1-CK6, which increased by 2.13-8.50%, with large fluctuations. Among them, the CK4 treatment (without controlled synthesis conditions) and CK5 (exceeding the specified stirring time) showed relatively better results, while the other treatments showed significant differences. This indicates that directly mixing copper sulfate, excessive temperature reaction, excessive concentration reaction, and failure to perform secondary stirring were the main reasons for the poor performance of the control protein. A similar pattern was observed in the number of grains per ear. Regarding the incidence data of seroconvertible tumors, we can see that the T1-T3 treatments are significantly better than the others, while the CK1-CK6 treatments show varying results. However, there are patterns to be found. Firstly, the CK4-CK6 treatments do not cause serious differences in the synthesized protein, at most resulting in insufficient conversion. Therefore, although their disease resistance is weakened, they still have a certain effect. The other three treatments have a more significant impact on copper protein, which directly leads to a weakening of disease resistance.

[0101] Test Example 4

[0102] The copper-containing synthetic protein I produced in Example 1 and the copper-containing comparative protein I produced in Comparative Example 1 were subjected to FTIR detection (see details). Figure 1 and Figure 2 And by comparing them, we found that:

[0103] The copper-containing synthetic protein I showed seven peaks and one peak group consisting of many dense peaks, with values ​​of 3231.39 cm⁻¹. -1 1632.78cm -1 1574.32cm -1 1391.64cm -1 1284.00cm -1 1199.80cm -1 1129.83cm -1 and 400-435cm -1 The characteristic region contains four peaks, representing the stretching vibration of the NH bond (3231.39 cm⁻¹). -1 ), stretching vibration of C=O bond (1632.78cm) -1 Asymmetric stretching vibration of the COO- bond (1574.32 cm⁻¹) -1In-plane bending shear vibration of CH bonds (1391.64 cm) -1 These four peaks form the basic framework of polyaspartic acid and exhibit strong chelating properties; while in the fingerprint region, four peaks (or peak groups) appear, representing the twisted helical vibration of the CH(CH2) bond (1284.00 cm⁻¹). -1 Out-of-plane vibrations of CH(CH2) bonds (1199.80 cm⁻¹) -1 The stretching vibration of the CN bond (1129.83cm) -1 ) and copper protein characteristic chelate bonds 400-435cm -1 These peaks all indicate that, while retaining the basic structure of polyaspartic acid, copper-containing synthetic protein I has produced a new structure, especially at 1200 cm⁻¹. -1 The structural peaks on the left and right.

[0104] The corresponding copper-containing contrast protein I showed 11 peaks and a peak group consisting of many dense peaks, with values ​​of 3235.16 cm⁻¹. -1 2096.18cm -1 1584.43cm -1 1442.14cm -1 1399.84cm -1 1325.31cm -1 1267.78cm -1 1105.25cm -1 1006.98cm -1 973.11cm -1 926.24cm -1 and 400-470cm -1 The characteristic region contains five peaks, representing the stretching vibration of the NH bond (3235.16 cm⁻¹). -1 The variation peak of the C=C bond (2096.18 cm⁻¹) -1 Asymmetric stretching vibration of the COO- bond (1584.43 cm) -1 ), stretching vibration of C-OH bond (1442.14 cm⁻¹) -1 The in-plane bending shear vibration of the CH bond (1399.84 cm) and the CH bond. -1 These five peaks form the basic framework of polyaspartic acid, but they still show differences compared to the sample in Example 1, especially the stretching vibration of the additional C-OH bond (1442.14 cm⁻¹). -1 ) and reduced C=O bond stretching vibrations (1632.78 cm) -1The results revealed the instability and low chelation efficiency of the structure in Comparative Example 1; while eight peaks (or peak groups) appeared in the fingerprint region, which were the twisted helical vibrations of the CH(CH2) bond (1325.31 cm⁻¹). -1 Asymmetric twisting of CH(CH2) bonds (1267.78cm) -1 The stretching vibration of the CN bond (1105.25cm) -1 and 1006.98cm -1 Irregular changes in the =CH bond (973.11cm) -1 and 926.24cm -1 ) and copper protein characteristic chelate bonds 400-470cm -1 These peaks indicate that, while retaining the basic structure of polyaspartic acid, there are some free carboxylate groups and variations in many locations, resulting in irregular peak variations. This also explains the low chelation efficiency and possible dissociation phenomena in Comparative Example 1. These peaks show significant instability compared to the spectra of Example 1, thus demonstrating the difference between the two.

[0105] Finally, it should be noted that the above-described embodiments are merely specific implementations of the present invention, used to illustrate the technical solutions of the present invention, and not to limit them. The scope of protection of the present invention is not limited thereto. Although the present invention has been described in detail with reference to the foregoing embodiments, those skilled in the art should understand that any person skilled in the art can still modify or easily conceive of changes to the technical solutions described in the foregoing embodiments within the scope of the technology disclosed in the present invention, or make equivalent substitutions for some of the technical features; and these modifications, changes, or substitutions do not cause the essence of the corresponding technical solutions to deviate from the spirit and scope of the technical solutions of the embodiments of the present invention, and should all be covered within the scope of protection of the present invention.

Claims

1. A method for preparing copper protein, characterized in that, Includes the following steps: S1. Using copper-containing substances as raw materials, or a mixture of copper-containing substances and zinc-containing substances as raw materials, the raw materials are dissolved in ammonia water at low temperature to obtain a reaction solution, which is then concentrated to obtain a protein synthesis solution; S2. Mix polysuccinimide with water and stir until it is dispersed into particles with a diameter of <10μm to obtain the protein base solution; S3. Add the protein synthesis solution from step S1 to the protein base solution from step S2, and react at 40-50℃ for 1-3 hours to obtain copper protein.

2. The method for preparing copper protein according to claim 1, characterized in that, The copper-containing substance is one or both of copper monoxide and copper hydroxide; the zinc-containing substance is zinc oxide or zinc hydroxide, and the mass ratio of copper to zinc is 1:(0.01-0.2).

3. The method for preparing copper protein according to claim 2, characterized in that, The purity of copper monoxide is ≥99%, the purity of copper hydroxide is ≥99%, the purity of zinc oxide is ≥99.5%, the purity of zinc hydroxide is ≥99.5%, and the mass concentration of ammonia is 25%.

4. The method for preparing copper protein according to claim 1, characterized in that, The molar ratio of raw materials to ammonia is 1:(6.0-6.5); in the protein synthesis liquid stage, the molar ratio of raw materials to ammonia is concentrated to 1:(4.0-4.2).

5. The method for preparing copper protein according to claim 1, characterized in that, The reaction temperature of the reaction solution is controlled at ≤40℃; the temperature at which it is concentrated and transformed into a protein synthesis solution is 55-65℃.

6. The method for preparing copper protein according to claim 1, characterized in that, The mass ratio of polysuccinimide to water is 1:(1-2), and the dispersion is carried out in two stages. In the first stage, the stirring speed is 100-200 r / min and the duration is 10-15 min. The second stage involves mixing at a speed of 3000-4500 r / min for 30-45 min, while controlling the material temperature to not exceed 65℃.

7. The method for preparing copper protein according to claim 1, characterized in that, In step S3, the time for adding the protein synthesis solution to the protein base solution is controlled at 15-25 min, and the mass ratio of the protein synthesis solution to the protein base solution is 1:(2-4).

8. A fertilizer synergist, characterized in that, Including the copper protein prepared according to any one of claims 1-7.

9. The application of copper protein in fertilizers, characterized in that, When the copper protein is added to nitrogen-containing fertilizers, and the nitrogen-containing fertilizer is one or any combination of urea, ammonium sulfate, ammonium chloride, and ammonium carbonate, the amount of copper protein added is 4.5-6.5 kg / T of fertilizer; for other nitrogen-containing fertilizers, the amount of copper protein additive is 10-25 kg / T of fertilizer.

10. Copper-containing protein fertilizers are applied in the cultivation of gramineous crops.

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