Immune globulin G nano-particles as well as industrial preparation method and application thereof

Abstract

The invention relates to the technical field of preparation of immune globulin G nanoparticles, in particular to an immune globulin G nanoparticle as well as a preparation method and application thereof. The invention discloses an immune globulin G nano-particle. The particle size of the immune globulin G nano-particle is 10 nm to 200 nm; the invention discloses an immunoglobulin G nano-particle as well as a preparation method and application thereof. The prepared immunoglobulin G nano-particle has better thermal stability, and the prepared immunoglobulin G nano-particle still keeps effectiveness at high temperature; compared with the traditional immune globulin G, the immune globulin G nanoparticles prepared by the method disclosed by the invention have the advantages that the half-life period is effectively prolonged, and frequent intake is not needed; the immunoglobulin G nano-particles prepared by the method disclosed by the invention have higher biological activity.

Description

Technical Field

[0001] The present application relates to the technical field of preparation of immunoglobulin G nanoparticles, and in particular to an immunoglobulin G nanoparticle and an industrial preparation method and application thereof. Background Art

[0002] Bovine colostrum contains a large number of nutrients and bioactive components. The bioactive components of bovine colostrum include immunoglobulins, growth factors, hormones, enzymes, lactoferrin, cytokines, leukocytes, nucleosides and nucleotides. Immunoglobulin G (IgG) is the most important antibody in bovine colostrum, accounting for 80-85% of the total immunoglobulins. It contains two subclasses, IgG1 (the most abundant immunoglobulin isotype) and IgG2. Before the invention of artificial antibodies, bovine colostrum was regarded as the key to unlocking the mechanism of preventing microbial infection. It is well known that immunoglobulins are beneficial to human health in terms of immunogenic response. When the host is exposed to foreign matter (antigen), antibodies will bind, recognize and destroy bacteria, toxins, viruses and other antigens. When the antigen enters the human body next time, it will stimulate the body to produce the same antibodies to clear the infection in the body.

[0003] Temperature, pH, enzyme degradation and interaction with other components can affect the activity of immunoglobulins. To address such issues, many nanoparticles synthesized from non-biological composite materials have begun to be used in biomedicine, such as drug delivery, enzyme immobilization, diagnostic imaging, cancer therapy and targeted gene delivery. Nanoparticles have better stability, targeted delivery, and enhanced biological activity, which can significantly affect human health.

[0004] Therefore, how to prepare immunoglobulin G into nanoparticles with thermal stability, biological activity and longer half-life has become an urgent problem to be solved. Summary of the invention

[0005] In view of the deficiencies in the prior art, the present application provides an immunoglobulin G nanoparticle and an industrial preparation method and application thereof.

[0006] This application aims to solve the technical problems of high temperature instability, short half-life and loss of biological activity of immunoglobulin. By extracting IgG nanoparticles from bovine colostrum, its effectiveness is maintained, its service life is extended, and its biological activity is enhanced. It can be applied to various biomedicines, including beverages, treatment plans and other product manufacturing processes.

[0007] In a first aspect, the present application provides an immunoglobulin G nanoparticle, which adopts the following technical solution: An immunoglobulin G nanoparticle, wherein the particle size of the immunoglobulin G nanoparticle is 10nm-200nm.

[0008] The particle size of conventional immunoglobulin G (i.e., conventional IgG that has not been nanosized) is >500 nm, and immunoglobulin G is prone to aggregate with proteins or itself to obtain aggregated IgG.

[0009] The immunoglobulin G nanoparticles prepared in the present application have a particle size of 10nm-200nm. The nanoscale immunoglobulin IgG prepared in the present application does not aggregate with proteins or itself, so the prepared immunoglobulin G nanoparticles have more exposed surface sites, which can more effectively bind to pathogens and promote their inactivation.

[0010] Preferably, the immunoglobulin G nanoparticles are extracted from bovine colostrum.

[0011] In a second aspect, the present application provides an industrial preparation method of immunoglobulin G nanoparticles, using the following technical solution: An industrial preparation method of immunoglobulin G nanoparticles, the preparation method comprises the following steps: (1) centrifugally defatting the bovine colostrum to obtain defatted bovine colostrum; (2) heating the defatted bovine colostrum to 40-50° C., adjusting the pH of the defatted bovine colostrum to 4.4-4.6, filtering, removing casein, and obtaining whey; (3) adjusting the pH value of the whey to 3-3.8 and filtering to obtain purified whey; (4) ultrafiltration and concentration of the purified whey to obtain nanoparticles; (5) Drying the nanoparticles to obtain immunoglobulin G nanoparticles.

[0012] By adopting the above technical scheme, in the present application, immunoglobulin G nanoparticles are prepared through the steps of centrifugal defatting, casein removal, whey purification, ultrafiltration concentration, and drying; the prepared immunoglobulin G nanoparticles have good dispersibility, thermal stability, hydrophobicity and non-polarity.

[0013] In the present application, an acid solution is first used to adjust the pH of defatted bovine colostrum to precipitate casein, and the casein is filtered out to obtain whey. The pH of the whey is adjusted to further purify the whey and further remove the protein precipitate to obtain purer whey.

[0014] Preferably, in step (1), the temperature of the bovine colostrum is controlled to be 4-18° C. before the bovine colostrum is centrifuged and defatted.

[0015] By adopting the above technical solution, the temperature of the bovine colostrum is controlled at 4-18°C before centrifugal degreasing, which can ensure the quality and biological activity of the bovine colostrum and effectively prevent the bacteria in the colostrum from affecting the quality of the bovine colostrum.

[0016] Preferably, during the centrifugal defatting in step (1), the centrifugal speed is 3000-6000 rpm, and the temperature of the bovine colostrum is controlled to be ≤25°C during the centrifugation process.

[0017] By adopting the above technical solution, by controlling the temperature of bovine colostrum during the centrifugation process, a lower temperature is helpful for fat separation, and effectively preventing fat liquefaction caused by increased temperature, which affects the separation efficiency.

[0018] Preferably, in step (2), the solution used to adjust the pH of the defatted bovine colostrum is an acid solution.

[0019] Preferably, the acid solution used is hydrochloric acid solution, citric acid solution, vitamin C solution or acetic acid solution.

[0020] Preferably, in step (2), the filtering step is as follows: filtering the pH-adjusted defatted bovine colostrum through a 60-200 mesh sieve.

[0021] Preferably, in step (3), the solution used to adjust the pH of whey is an acid solution.

[0022] Preferably, the acid solution used is hydrochloric acid solution, citric acid solution, vitamin C solution or acetic acid solution.

[0023] Preferably, in step (3), the filtering step is as follows: filtering the pH-adjusted whey through a sieve of ≥200 mesh.

[0024] Preferably, in the step (4), during the ultrafiltration concentration, an ultrafiltration device with a membrane molecular weight cutoff of 10 KDa is used for ultrafiltration concentration.

[0025] Preferably, the immunoglobulin G nanoparticles obtained after drying in step (5) have a particle size of 10 nm-200 nm.

[0026] Preferably, the drying method in step (5) includes freeze drying, spray drying and other drying methods.

[0027] In a third aspect, the present application provides an immunoglobulin G nanoparticle or an application of the immunoglobulin G nanoparticle prepared by the above method in the preparation of food, health care products or medicines, using the following technical solution: An immunoglobulin G nanoparticle or an application of the immunoglobulin G nanoparticle prepared by the above method in the preparation of food, health care products or medicines.

[0028] The immunoglobulin G nanoparticles prepared by the method of the present application have structural stability and high antibody activity, and can be widely used in food, health products, liquid beverages or other products.

[0029] In summary, the present application includes at least one of the following beneficial technical effects: 1. The present application discloses an immunoglobulin G nanoparticle and an industrial preparation method and application thereof. The prepared immunoglobulin G nanoparticle has better thermal stability and the prepared immunoglobulin G nanoparticle still maintains its effectiveness at high temperature; 2. The immunoglobulin G nanoparticles prepared by the method of the present application effectively extend the half-life compared with the traditional immunoglobulin G; 3. The immunoglobulin G nanoparticles prepared by the method of the present application have higher biological activity. BRIEF DESCRIPTION OF THE DRAWINGS

[0030] Figure 1 The transmission electron microscopy (TEM) image of the immunoglobulin G nanoparticles extracted by the method of Example 1; Figure 2 The SEM images are of immunoglobulin G nanoparticles extracted by the method of Example 1 and conventional IgG that has not been nano-processed; wherein, Figure 2 The left picture is the SEM image of IgG nanoparticles. Figure 2 The right picture is a SEM image of conventional IgG that has not been nanosized; Figure 3 Dynamic light scattering shows the size distribution of immunoglobulin G nanoparticles extracted using the method of Example 1; Figure 4 The thermal stability diagram of immunoglobulin G nanoparticles extracted by the method of Example 1 and conventional IgG that has not been nano-processed; wherein, Figure 4 (a) is the thermal stability diagram of IgG nanoparticles; Figure 4 (b) is the thermal stability diagram of conventional IgG without nano-processing; Figure 5 The denaturation diagram of immunoglobulin G nanoparticles extracted by the method of Example 1 and conventional IgG that has not been nano-processed; wherein, Figure 5 (a) is the denaturation diagram of IgG nanoparticles; Figure 5 (b) is the denaturation diagram of conventional IgG without nano-treatment; Figure 6 This is an assembly diagram of immunoglobulin G nanoparticles extracted using the method of Example 1 and conventional IgG that has not been nano-processed; Figure 7 The denaturation test results of immunoglobulin G nanoparticles extracted by the method of Example 1 and conventional IgG that has not been nano-processed; Figure 8The infrared spectra (FTIR) of immunoglobulin G nanoparticles extracted by the method of Example 1 and conventional IgG that has not been nano-processed; Fig. 9 The X-ray diffraction patterns (X-RD) of immunoglobulin G nanoparticles extracted by the method of Example 1 and conventional IgG that has not been nanosized; Fig.10 Thermogravimetric analysis (TGA) of immunoglobulin G nanoparticles extracted by the method of Example 1 and conventional IgG that has not been nano-processed. DETAILED DESCRIPTION

[0031] The technical solution of the present application is further illustrated by specific embodiments below. The specific embodiments do not limit the protection scope of the present application; some non-essential modifications and adjustments made by others based on the concept of the present application still fall within the protection scope of the present application.

[0032] The raw materials involved in this application are all commercially available products, among which: Conventional IgG without nano-treatment, i.e., reagent grade human purified immunoglobulin G, was purchased from Sigma-Aldrich (USA).

[0033] The present application is further described in detail below in conjunction with embodiments and comparative examples.

[0034] An industrial preparation method of immunoglobulin G nanoparticles, the steps are as follows: Step 1: Collect colostrum: milk the cows and collect colostrum, and control the temperature of the colostrum to 4-18°C; Step 2: Centrifugal degreasing: transport the colostrum to a centrifugal degreasing machine for degreasing to obtain defatted colostrum; Among them, the current setting value of the centrifugal degreasing machine is 7.5-8.5A, and the speed is 3000-6000rpm; during the centrifugal degreasing process, the temperature of the bovine colostrum is controlled to be ≤25°C.

[0035] Step 3, removing casein: raising the temperature of the defatted bovine colostrum to 40-50°C, and adjusting the pH value of the defatted bovine colostrum to 4.4-4.6 using an acid solution (such as hydrochloric acid solution, citric acid solution, vitamin C solution or acetic acid solution, etc.); at this time, the casein reaches its isoelectric point and begins to coagulate, causing the casein to precipitate, and filtering through a 60-200 mesh sieve to remove the casein and obtain whey; The step of filtering through a 60-200 mesh screen in this step can be replaced by centrifugation using a centrifugal device.

[0036] Step 4, whey purification: purifying the obtained whey to obtain purified whey; Specifically, the pH value of the whey is adjusted to 3-3.8 using an acid solution (such as hydrochloric acid solution, citric acid solution, vitamin C solution or acetic acid solution, etc.), and filtered through a sieve of ≥200 mesh to finally obtain purified whey; The step of filtering through a sieve of ≥200 mesh in this step can be replaced by centrifugation using a centrifugal device.

[0037] Step 5, ultrafiltration and concentration: ultrafiltration and concentration of the purified whey to remove lactose, small molecules and regularly dispersed immunoglobulin IgG, and convert the immunoglobulin IgG into nanoparticles; Among them, ultrafiltration equipment is used for ultrafiltration concentration, and the membrane cutoff molecular weight used by the ultrafiltration equipment is 10KDa.

[0038] Step 6: Drying: Drying the obtained nanoparticles to obtain immunoglobulin G nanoparticles; the particle size of the immunoglobulin G nanoparticles is 10nm-200nm.

[0039] Example 1: Immunoglobulin G nanoparticles extracted from bovine colostrum An industrial preparation method of immunoglobulin G nanoparticles extracted from bovine colostrum comprises the following steps: (1) Collecting bovine colostrum: Milk the cows and collect the colostrum, then immediately refrigerate at 4°C; (2) Centrifugal degreasing: The colostrum is transported to a centrifugal degreasing machine for degreasing to obtain defatted colostrum; The current setting value of the centrifugal skimmer is 8.2A, the speed is 5000rpm, and the initial temperature of the milk during the centrifugation process is controlled to be ≤25℃; (3) Removal of casein: The temperature of the defatted bovine colostrum was raised to 45° C., the pH of the defatted bovine colostrum was adjusted to 4.6 using a 0.1 M citric acid solution to precipitate casein, and the casein was removed by filtering through a 120-mesh sieve to obtain whey; (4) Whey purification: The pH value of the whey was adjusted to 3.8 using 0.1 M citric acid solution and filtered through a 200-mesh sieve to obtain purified whey.

[0040] (5) Ultrafiltration concentration: Purified water is added to the ultrafiltration device for ultrafiltration, the membrane cutoff molecular weight used in the ultrafiltration device is 10 kDa, and the purified whey obtained in step (4) is added with 2 times the volume of ultrapure water, and ultrafiltered to the original volume through a 10 kDa ultrafiltration membrane, the retentate is recovered, and 2 times the original volume of ultrapure water is added, and ultrafiltration is performed again. This process is repeated 3 times or more until the absorbance of the permeate measured at 280 nm by an ultraviolet spectrophotometer is less than 0.05, and then it is concentrated.

[0041] (6) Drying: The ultrafiltration concentrate obtained was freeze-dried for 48 hours to obtain immunoglobulin G nanoparticles (i.e., IgG nanoparticles).

[0042] Embodiment 2: The difference from Example 1 is that in step (4), the pH value of the whey is adjusted to 3.5.

[0043] Embodiment 3: The difference from Example 1 is that in step (4), the pH value of the whey is adjusted to 3.

[0044] Comparative Example 1: The difference from Example 1 is that in step (4), the pH value of the whey is adjusted to 3.9.

[0045] Comparative Example 2: The difference from Example 1 is that in step (4), the pH value of the whey is adjusted to 2.9.

[0046] The above method was used to extract immunoglobulin G nanoparticles from 100 kg of bovine milk, and the immunoglobulin G nanoparticles obtained in Examples 1-3, Comparative Example 1, and Comparative Example 2 were measured using a transmission electron microscope (TEM). The results are shown in Table 1.

[0047] Table 1 Test results Combined with the detection results in Table 1, it can be seen that the size distribution of the immunoglobulin G nanoparticles prepared in Examples 1-3 is between 10 nm and 200 nm.

[0048] Combining Example 1 and Comparative Example 1, it can be seen that although the yield of Comparative Example 1 is greater than that of immunoglobulin G nanoparticles, the particle size of Comparative Example 1 is greater than that of Example 1. This is mainly because the pH becomes higher, which makes the prepared immunoglobulin G nanoparticles contain a certain amount of impurities, thereby making the yield of Comparative Example 1 higher than that of Example 1. Therefore, the detection result of Example 1 is better.

[0049] Combining Example 1 and Comparative Example 2, it can be seen that the output of Comparative Example 2 is less than that of Comparative Example 1, so the detection result of Example 1 is better.

[0050] Therefore, according to Table 1, when purifying whey, the pH value of the whey is adjusted to 3-3.8, and the immunoglobulin G nanoparticles prepared have the best effect, and the immunoglobulin G nanoparticles prepared in Example 1 have the highest yield.

[0051] Performance testing: The immunoglobulin G nanoparticles prepared in Example 1 were measured.

[0052] 1. Transmission electron microscopy (TEM) determination: The immunoglobulin G nanoparticles obtained in Example 1 were measured using a transmission electron microscope (TEM). The results showed that the size of the immunoglobulin G nanoparticles can be seen in the image to be distributed between 10 nm and 200 nm, and the size of each immunoglobulin G nanoparticle is 10 nm; most of the immunoglobulin G nanoparticles exist in the form of clusters, each cluster contains four to ten nanoparticles, and the size does not exceed 200 nm; the specific results are shown in Figure 1 .

[0053] 2. Scanning electron microscope (SEM) determination The morphology of the immunoglobulin G nanoparticles prepared in Example 1 was characterized by scanning electron microscopy (SEM). Figure 2 Shown is the morphology comparison between IgG nanoparticles and conventional IgG without nano-processing.

[0054] Figure 2 Left (IgG nanoparticles): This is a SEM image of IgG nanoparticles at a magnification of 350,000 times. The microstructure of the nanoparticles can be clearly observed in the image. The particles are in the shape of regular spheres and are closely arranged, indicating that these IgG particles have been successfully nanosized.

[0055] Figure 2 Right (conventional IgG without nanoparticle processing): This is a SEM image of conventional IgG without nanoparticle processing, with a magnification of 400 times. The image shows large, irregular flake structures, indicating that the IgG sample has a larger particle size and a relatively flat surface, which is significantly different from the fine structure of nanoparticles.

[0056] pass Figure 2 It can be seen that the IgG nanoparticles prepared by the method of the present application are more uniform and fine, while conventional IgG that has not been nano-processed presents a larger block or flake morphology, further proving the effectiveness of the nano-process.

[0057] 3. Dynamic light scattering (DLS) measurement The immunoglobulin G nanoparticles obtained in Example 1 were further measured using a dynamic light scattering particle size analyzer. The DLS measurement results showed that the size of the immunoglobulin G nanoparticles was between 10 nm and 200 nm. Figure 3 As shown, the DLS measurement results not only provide the average value of the particle size, but also show the distribution range and uniformity of the particle size, proving that the IgG nanoparticles maintain good dispersion and stability during the preparation process.

[0058] 4. Thermal stability determination (1) Wavelength and absorbance detection The wavelength and absorbance of the immunoglobulin G nanoparticles prepared in Example 1 and conventional IgG without nano-treatment were detected by ultraviolet-visible spectrometer at 85°C for 0, 5 min, 10 min, 20 min, 40 min, 60 min, and 120 min, respectively. The detection results are as follows: Figure 4 shown.

[0059] according to Figure 4 It can be seen that the absorbance of immunoglobulin G nanoparticles differs little at different time points, which indicates that after nano-treatment, the sample has a higher stability in the UV-visible region (200-400 nm).

[0060] Comparing the absorbance curves at different time points, the absorbance of immunoglobulin G nanoparticles changed little during the heat treatment (85°C), while the absorbance of conventional IgG that had not been nano-processed showed a significant downward trend over time. This shows that nano-processing can improve the stability of IgG at high temperatures and slow down its denaturation or degradation.

[0061] The absorbance changes of conventional IgG that has not been treated with nano-sizing are significantly greater at different time points (especially 5 minutes, 10 minutes and thereafter), indicating that its structural stability is poor and it is more likely to undergo denaturation or polymerization under high temperature conditions; and the absorbance of conventional IgG that has not been treated with nano-sizing gradually decreases with time, which may be due to the denaturation of its structure caused by high temperature, and some IgG molecules may undergo cross-linking reactions with other molecules, thereby affecting the absorbance; while the IgG after nano-sizing shows a higher resistance to denaturation under the same conditions, indicating that it has better structural integrity and a lower tendency to polymerization during the heat treatment process.

[0062] As the heating time increases, the absorbance of conventional IgG that has not been treated with nanoparticles drops rapidly in the early stage (within 10 minutes), and then the absorbance changes slowly. This indicates that in the initial stage, IgG undergoes a more intense denaturation reaction, and then gradually reaches a relatively stable state; while the absorbance of nano IgG particles changes less at each time point, indicating that it maintains a relatively stable state throughout the heating process.

[0063] The immunoglobulin G nanoparticles prepared in Example 1 and conventional IgG that has not been treated with nano-sizing were heated at 85°C for 0, 5 min, 10 min, 20 min, 40 min, 60 min, and 120 min, respectively, and the denaturation of the immunoglobulin G nanoparticles and the conventional IgG that has not been treated with nano-sizing was observed. The denaturation results of the immunoglobulin G nanoparticles and the conventional IgG that has not been treated with nano-sizing are shown in FIG. Figure 5 shown.

[0064] according to Figure 5 It can be seen that conventional IgG that has not been nano-processed is heated at 85°C for 5 minutes, denatured, and precipitated, and all the precipitation is heated to 40 minutes; the immunoglobulin G nanoparticles prepared in the present application do not denature or precipitate after heating at 85°C for 40 minutes. Compared with conventional IgG, the IgG nanoparticles prepared in the present application have better stability.

[0065] In summary, the thermal and structural stability of immunoglobulin G is significantly improved after nano-processing, and it is not easy to denature and aggregate under high temperature conditions. This method also ensures that IgG is seamlessly integrated into liquid beverages and other delivery systems without degradation, paving the way for new multifunctional biomedical applications. The enhanced properties of IgG nanoparticles give them better prospects in biomedical applications such as drug delivery and targeted therapy, which can provide more effective treatment methods and improve human health. IgG nanoparticles retain and improve their ability to effectively bind antigens, which is crucial for diagnosis, immunotherapy and vaccine development.

[0066] (2) Denaturation result detection The immunoglobulin G nanoparticles prepared in Example 1 and the conventional IgG without nano-treatment were heated at 60°C, 65°C, 70°C, 75°C, 80°C, and 85°C for 0-120 min, respectively. The heated products were centrifuged, and the percentage of the solid content after centrifugation to the total content was calculated and recorded as the denaturation result. The test results are shown in Figure 7 .

[0067] according to Figure 7 It can be seen that there are significant differences in the thermal stability results between immunoglobulin G nanoparticles and conventional IgG that have not been nano-processed.

[0068] The activity of the IgG nanoparticles was not affected when the IgG nanoparticles were heated at 60°C-85°C for 0-120 minutes.

[0069] When conventional IgG that has not been treated with nanomaterials is heated at 60°C-85°C for 0-120 minutes, IgG denatures and precipitates. Conventional IgG that has not been treated with nanomaterials gradually loses its activity as the temperature rises and the heating time increases.

[0070] Therefore, compared with conventional IgG that has not been nano-processed, the IgG nanoparticles prepared by the method of the present application maintain good dispersibility, stability and activity, extend the half-life, and can be widely used in food, health products, liquid beverages, medical applications, etc.

[0071] With the advancement of nanotechnology, the stability and functionality of IgG nanoparticles have been significantly enhanced, providing new opportunities for applications in various industries, including the medical field.

[0072] 5. Infrared spectroscopy (FTIR) determination The absorption peaks of the immunoglobulin G nanoparticles prepared in Example 1 and conventional IgG that has not been nano-processed were detected using an infrared spectrometer.

[0073] The infrared spectrum shows the absorption peak characteristics of IgG nanoparticles and conventional IgG samples without nano-treatment, such as Figure 8 As shown. According to the absorption peaks of the conventional IgG sample that has not been nano-processed, it can be seen that the NH stretching vibration (amide A) causes a broad and strong absorption band centered at 3300 cm-1. Other strong absorption bands appear near 1650 cm-1 and 1534 cm-1, respectively, caused by C=O stretching vibration (amide I) and NH bending vibration (amide II). The absorption bands centered at 3300 cm-1, 1650 cm-1, 1534 cm-1 and 1075 cm-1 correspond to protein NH stretching (amide A), C=O stretching (amide I), NH bending (amide II) and CO stretching vibration, respectively.

[0074] There are significant differences in the spectra of IgG nanoparticles and conventional IgG that has not been nanosized. Pure IgG nanoparticles without added chemical components have narrower peaks, indicating a lower degree of structural variation. In contrast, conventional IgG samples that have not been nanosized show broader peaks at 3300cm-1, 1650cm-1, and 1075cm-1, reflecting that the structures of C=O and NH functional groups are more stable, so conventional IgG that has not been nanosized is easy to combine with other components.

[0075] 1235cm -1 and 1075cm -1 The absorption band at 1235 cm corresponds to the antisymmetric stretching vibration of the ester group or the CO stretching of the alcohol, indicating that conventional IgG without nano-treatment is more likely to undergo cross-linking reactions with other compounds to form a polymer structure formed by immune protein G and other components, namely, polymerized IgG; and the filament peak (1235 cm -1 and 1075cm -1 ) did not appear in the spectrum of IgG nanoparticles, which indicates that IgG nanoparticles have more monomers, trimers or tetramers than conventional IgG that has not been nanosized, rather than polymer structures formed with other components (see Figure 6 ).

[0076] 6. X-ray diffraction (XRD) determination The immunoglobulin G nanoparticles prepared in Example 1 and conventional IgG that has not been nanosized were detected using an X-ray diffractometer.

[0077] According to the XRD measurement results, the average crystallinity value of conventional IgG is 42.52; the average value of IgG nanoparticles (NPs) is 2.22; the IgG nanoparticles prepared by the present invention do not undergo obvious aggregation, and compared with conventional IgG that has not undergone nano-treatment, the crystallinity is significantly improved.

[0078] In addition, due to the reduced particle size and improved dispersibility, IgG nanoparticles showed high stability. Fig. 9 ) showed the characteristic diffraction peaks of IgG nanoparticles, proving their good crystalline structure and highly dispersed state.

[0079] 7. Use thermogravimetric analysis (TGA) to measure The immunoglobulin G nanoparticles prepared in Example 1 and conventional IgG that has not been nanosized were detected using a thermogravimetric analyzer.

[0080] TGA results show that during the heating process, the weight loss curves of IgG nanoparticles and conventional IgG that have not been nano-processed are significantly different. Fig.10 As shown in the figure, the weight loss of IgG nanoparticles at 600°C was 23.7%, while the weight loss of conventional IgG without nano-processing was 21.8%. This shows that IgG nanoparticles have high thermal stability, further verifying that they can maintain structural integrity and stability during the preparation process.

[0081] The present application discloses an innovative technology for extracting IgG nanoparticles from bovine colostrum, which nanoparticles have strong thermal stability, extended half-life and improved biological activity. Immunoglobulin G nanoparticles prepared by the present application are used to manufacture nanoparticles that remain active at high temperatures, making them suitable for thermally processed products. By extending its half-life, frequent intake is not required, which improves commercial feasibility. In addition, the method used in the present application focuses on retaining and enhancing the antigenic, antibacterial and immune-enhancing properties of IgG during processing and storage. The method also ensures that IgG is seamlessly integrated into liquid beverages and other delivery systems without degradation, paving the way for new multifunctional biomedical applications. The enhanced properties of IgG nanoparticles give it better prospects in biomedical applications, such as drug delivery and targeted therapy, which can provide more effective treatment methods and improve human health. IgG nanoparticles retain and improve their ability to effectively bind antigens, which is crucial for diagnosis, immunotherapy and vaccine development.

[0082] The present application discloses an immunoglobulin G nanoparticle and an industrial preparation method and application thereof. The method fills the gap in the domestic industrial preparation of high-purity immunoglobulins. Currently, the purity of immunoglobulin G industrially prepared in China is only 20-30%. The high-purity immunoglobulin G nanoparticles obtained by the method of the present invention can be as high as 80%.

[0083] The findings presented herein demonstrate that IgG nanoparticles possess a high degree of stability and efficiency, making them promising candidates for biomedical and pharmaceutical applications requiring stable and bioactive formulations.

[0084] IgG nanoparticles maintain structural integrity and amino acid stability under various conditions, making them suitable for biomedical and pharmaceutical applications. In addition, the environmentally friendly and cost-effective production method adopted in this application highlights the potential for large-scale application of IgG nanoparticles in functional foods, diagnostics and therapeutic development. Research can study the encapsulation of other bioactive molecules with IgG nanoparticles to create multifunctional nanocarriers. In addition, the interaction of IgG nanoparticles with various cell types and their immunomodulatory effects need to be thoroughly examined to evaluate their safety and effectiveness in clinical settings.

[0085] This study provides valuable insights into the potential industrial applications and versatility of IgG nanoparticles, highlighting their enhanced stability and bioactivity compared to traditional chemical nanoparticles.

Claims

1. An immunoglobulin G nanoparticle, characterized in that: The particle size of the immunoglobulin G nanoparticles is 10nm-200nm.

2. The immunoglobulin G nanoparticle according to claim 1, characterized in that: The immunoglobulin G nanoparticles are extracted from bovine colostrum.

3. An industrial preparation method of immunoglobulin G nanoparticles, characterized in that: The preparation method steps are as follows: (1) centrifugally defatting the bovine colostrum to obtain defatted bovine colostrum; (2) heating the skimmed bovine colostrum to 40-50° C., adjusting the pH of the skimmed bovine colostrum to 4.4-4.6, filtering, removing casein, and obtaining whey; (3) adjusting the pH value of the whey to 3-3.8 and filtering to obtain purified whey; (4) ultrafiltration and concentration of the purified whey to obtain nanoparticles; (5) Drying the nanoparticles to obtain immunoglobulin G nanoparticles.

4. The industrial preparation method of immunoglobulin G nanoparticles according to claim 3, characterized in that: In the step (1), before the bovine colostrum is centrifuged and defatted, the temperature of the bovine colostrum is controlled to be 4-18°C; During the centrifugal defatting in step (1), the centrifugal speed is 3000-6000 rpm, and the temperature of the bovine colostrum is controlled to be ≤25°C during the centrifugation process.

5. The industrial preparation method of immunoglobulin G nanoparticles according to claim 3, characterized in that: In the step (2), the solution used to adjust the pH of the defatted bovine colostrum is an acid solution; in the step (3), the solution used to adjust the pH of the whey is an acid solution.

6. The industrial preparation method of immunoglobulin G nanoparticles according to claim 5, characterized in that: The acid solution is hydrochloric acid solution, citric acid solution, vitamin C solution or acetic acid solution.

7. The industrial preparation method of immunoglobulin G nanoparticles according to claim 3, characterized in that: In the step (2), the filtering step is as follows: filtering the pH-adjusted defatted bovine colostrum through a 60-200 mesh sieve.

8. The industrial preparation method of immunoglobulin G nanoparticles according to claim 3, characterized in that: In the step (3), the filtering step is as follows: filtering the whey after adjusting the pH through a sieve of ≥200 mesh.

9. The industrial preparation method of immunoglobulin G nanoparticles according to claim 3, characterized in that: In the step (4), during the ultrafiltration concentration, an ultrafiltration device with a membrane molecular weight cutoff of 10KDa is used for ultrafiltration concentration; the immunoglobulin G nanoparticles obtained after drying in the step (5) have a particle size of 10nm-200nm.

10. Use of the immunoglobulin G nanoparticles according to claim 1 or 2, or the immunoglobulin G nanoparticles prepared by the method according to any one of claims 3 to 9 in the preparation of food, health products or medicines.

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