Method for preparing carbon dots from bean clear liquid as well as carbon dots and application thereof

Abstract

The invention discloses a processing method for preparing carbon dots from bean clear liquid, the carbon dots and application thereof, and belongs to the technical field of nano materials. The method comprises the following steps: carrying out a hydrothermal reaction on the bean clear liquid, cooling after the hydrothermal reaction is ended, centrifuging the cooled reaction liquid, taking the supernate, filtering, taking the filtrate, dialyzing, and drying the dialysate, so as to obtain the bean clear liquid carbon dots. According to the method, the bean clear liquid is utilized, so that the effect of waste utilization can be achieved, and the environmental pollution can be reduced. The prepared bean supernatant carbon dots are good in dispersity and stability, can penetrate through the blood brain barrier and inhibit Abeta40 aggregation, and are wide in application range and high in additional value.

Description

Technical Field

[0001] The present invention relates to the technical field of nanomaterials, and particularly to a processing method for preparing carbon dots from soybean whey, the carbon dots prepared thereby, and their applications. Background Art

[0002] Carbon dots are small carbon nanoparticles with a particle size less than 10 nm, and have recently emerged as a kind of benign nanoparticles, which have the potential to replace heavy metals contained in toxic quantum dots. Carbon dots have characteristics such as excitation wavelength dependence, photoluminescence, good biocompatibility, low cytotoxicity, and optical stability, and can easily penetrate cell membranes, so they have potential application prospects in bioimaging and clinical treatment.

[0003] The synthesis methods of carbon dots are mainly classified into two methods: "top-down" and "bottom-up". The top-down method reduces large carbon materials from large to small by physical or chemical methods until the size reaches the nanoscale. The bottom-up method uses small-sized carbon-containing organic compounds as carbon sources, and combines small molecules in an orderly or disorderly manner to grow from small to large, so as to reach a certain size to prepare carbon dots. The top-down method generally includes: arc discharge method, laser ablation method, electrochemical method, chemical oxidation method, etc.; the bottom-up method generally includes: solvothermal synthesis method, combustion method, high-temperature pyrolysis method, microwave method, ultrasonic method, etc. The carbon dots prepared by these two types of methods all exhibit excellent luminescence characteristics.

[0004] Biomass is any material extracted from plants or animals and is usually used for energy production. Research shows that biomass carbon dots not only have the chemical properties of traditional carbon dots, but also are more environmentally friendly and have the potential to reduce toxicity and improve biocompatibility due to their environmentally friendly synthesis method and high yield, and are considered to have the potential to improve the limitations of traditional carbon dots.

[0005] Tofu is made from soybeans and is a widely consumed daily food. However, in the large-scale production of tofu, there is an environmental problem, namely wastewater, i.e., soybean whey. And this natural substance, soybean whey, can be used as a biomass carbon source to green-synthesize fluorescent carbon dots. This raw material is easily available and can also achieve waste recycling, which can greatly reduce environmental pollution. Moreover, the present invention also finds that the carbon dots prepared using soybean whey can inhibit the aggregation of Aβ40, reduce the neurocytotoxicity and reactive oxygen species level caused by the aggregates of Aβ40, and at the same time have good biofilm permeability and other functions.

[0006] In the prior art, there is a patent CN 110155984 A (Method for hydrothermally synthesizing biomass fluorescent carbon dots using soybean dregs as raw material and its application), which discloses a method for hydrothermally synthesizing carbon quantum dots using soybean dregs as raw material, including: mixing soybean dregs with water to obtain a precursor solution containing soybean dregs, placing the precursor solution in an autoclave, heating and reacting at 100-500 °C, after the reaction is completed, cooling, removing insoluble substances, and freeze-drying the supernatant after dialysis to obtain carbon quantum dot powder. However, the patent does not mention that the obtained carbon quantum dot powder has the effect of inhibiting Aβ40 aggregation. Summary of the Invention

[0007] In order to overcome the deficiencies of the prior art, the present invention provides a method for preparing carbon dots from soybean whey, its carbon dots and applications. The preparation method uses soybean whey to green-synthesize fluorescent carbon dots, with easily available raw materials, which can reduce environmental pollution, and the preparation process of this method is green, non-toxic, environmentally friendly and safe.

[0008] The aggregation of amyloid-β protein is closely related to cognitive ability and memory. When it aggregates abnormally, toxic aggregates can be produced, and these toxic aggregates cause cell death by destroying the permeability, elasticity and synaptic conduction of cell membranes, etc., thereby making memory, thought, emotion, skills and body movements uncontrollable. The carbon dots of soybean whey prepared by the present invention can inhibit Aβ40 aggregation, reduce the neurocytotoxicity and reactive oxygen species level caused by the aggregates of Aβ40, and at the same time have functions such as good biofilm permeability.

[0009] In order to achieve the above object, the technical solution adopted by the present invention is: a processing method for preparing carbon dots from soybean whey, including the following steps:

[0010] Perform hydrothermal reaction on soybean whey, cool after completion, then centrifuge the cooled reaction solution, take the supernatant for filtration, then take the filtrate for dialysis, and then dry the dialysis solution to obtain carbon dots of soybean whey.

[0011] Further, the soybean whey is also called soybean clear water or yellow slurry water, which is the upper clear liquid produced after the soy products are "cured" during the fixed molding process, and is a milky yellow or milky white translucent liquid.

[0012] Further, the soy products include tofu, dried bean curd sheets or dried bean curd.

[0013] Further, the "curing" is also called "swelling slurry" or "nurturing slurry", which means that after the soybean milk is curdled (adding a coagulant), it is left standing for a period of time; the standing time is 10-30 minutes.

[0014] Specifically, the preparation process of the soybean whey is as follows: Mix soybeans with water at a mass ratio of 1:2 to 5 and soak for 10 to 24 hours. Then, add water to the soaked soybeans at a mass ratio of 1:3 to 10 to make raw soy milk. Heat the raw soy milk to obtain cooked soy milk. After the cooked soy milk is cooled to 75 to 90 °C, add magnesium chloride or an aqueous solution of magnesium chloride. The mass ratio of magnesium chloride to soybeans is 1:0.4 to 0.8. After stirring, perform curd retting for 10 to 30 minutes. The supernatant is the soybean whey.

[0015] Further, the temperature of the hydrothermal reaction is 150 to 200 °C.

[0016] Further, the time of the hydrothermal reaction is 10 to 15 hours.

[0017] Further, the temperature of the cooling is 10 to 40 °C.

[0018] Further, the conditions for centrifugation are centrifugation for 5 to 30 minutes at a rotational speed of 3000 to 10000 r / min.

[0019] Further, the filtration is carried out using a filter membrane with a pore size of 0.2 to 0.3 μm.

[0020] Further, the dialysis is carried out using a dialysis bag with a molecular weight cut-off of 300 to 1000 Da; the time of dialysis is 12 to 48 hours.

[0021] Further, the drying includes freeze-drying or hot-air drying.

[0022] Further, the average particle size of the carbon dots in the obtained soybean whey is 8.21 nm; the maximum excitation wavelength is 410 nm, and the maximum emission wavelength is 500 nm; it is brownish-yellow under visible light and emits green fluorescence under ultraviolet light irradiation.

[0023] The present invention provides carbon dots of soybean whey prepared according to the above method.

[0024] The present invention provides the application of the carbon dots of soybean whey in brain imaging.

[0025] Further, the application in brain imaging means that it can be used as a contrast agent for fluorescence imaging and photoacoustic imaging.

[0026] The present invention provides the application of the carbon dots of soybean whey in inhibiting Aβ40 aggregation.

[0027] Further, the application in inhibiting Aβ40 aggregation means using the carbon dots of soybean whey to prepare a drug or a drug carrier for preventing and treating Alzheimer's disease.

[0028] A kind of carbon dots for realizing brain imaging and inhibiting Aβ40 aggregation, which are prepared by hydrothermal reaction using soybean whey as raw material.

[0029] In one embodiment of the present invention, the preparation method of the carbon dots is specifically as follows: carry out hydrothermal reaction on the soybean whey, cool it after completion, then centrifuge the cooled reaction solution, take the supernatant for filtration, then take the filtrate for dialysis, and then dry the dialysis solution to obtain soybean whey carbon dots.

[0030] In one embodiment of the present invention, the soybean whey is also called soybean clear water or yellow slurry water, which is the upper clear liquid produced after the soybean products are retted during the fixed molding process, and is a milky yellow or milky white translucent liquid.

[0031] In one embodiment of the present invention, the temperature of the hydrothermal reaction is 150 - 200 °C, and the time is 10 - 15 hours.

[0032] In one embodiment of the present invention, the cooling temperature is 10 - 40 °C.

[0033] In one embodiment of the present invention, the centrifugation conditions are centrifugation for 5 - 30 minutes under the condition of a rotation speed of 3000 - 10000 r / min.

[0034] In one embodiment of the present invention, the filtration is carried out using a filter membrane with a pore size of 0.2 - 0.3 μm.

[0035] In one embodiment of the present invention, the dialysis is carried out using a dialysis bag with a molecular weight cut-off of 300 - 1000 Da; the dialysis time is 12 - 48 hours.

[0036] Beneficial effects

[0037] 1. The present invention uses soybean whey, a by-product of tofu processing, as a carbon source, and uses the hydrothermal method to green-synthesize fluorescent carbon dots. Compared with the top-down method, this method has the advantage of simple preparation. At the same time, compared with organic reagent carbon sources, the raw materials of the present invention are easily available and green and non-toxic.

[0038] 2. The output of soybean whey is large, and its random discharge will cause environmental pollution. The present invention uses it to prepare carbon dots, realizing waste recycling and reducing the environmental impact of soybean whey.

[0039] 3. The soybean whey carbon dots obtained in the present invention have good dispersibility, with an average particle size of about 8 nm, and have good pH stability under weak acid, weak base and neutral conditions. In addition, the soybean whey carbon dots can also penetrate the blood-brain barrier and have the function of inhibiting Aβ40 aggregation, with a wide application range and high added value. Brief description of the drawings

[0040] Figure 1It is the high-resolution transmission electron microscope image of carbon dots from soybean whey.

[0041] Figure 2 It is the XPS spectrum of carbon dots from soybean whey.

[0042] Figure 3 It is the ultraviolet-visible absorption spectrum.

[0043] Figure 4 It is the photo of carbon dots from soybean whey solution under visible light (left A) and ultraviolet light (right B).

[0044] Figure 5 It is the fluorescence emission spectrum of carbon dots from soybean whey at different excitation wavelengths.

[0045] Figure 6 It is the graph of the change in fluorescence intensity of carbon dots from soybean whey under different pH conditions.

[0046] Figure 7 It is the infrared spectrum of carbon dots from soybean whey.

[0047] Figure 8 It is the XRD pattern of carbon dots from soybean whey.

[0048] Figure 9 It is the aggregation situation detected by the ThT fluorescence method for 40 μmol / L Aβ40 and Aβ40 with different concentrations of carbon dots from soybean whey added.

[0049] Figure 10 A is the atomic force microscope image of 40 μmol / L Aβ40 after incubation at 37 °C for 24 hours; Figure 10 B is the atomic force microscope image of 40 μmol / L Aβ40 in the presence of an equal volume of 100 μg / mL carbon dots from soybean whey after incubation at 37 °C for 24 hours.

[0050] Figure 11 A is the cytotoxicity of carbon dots from soybean whey at different concentrations; Figure 11 AB is the effect of carbon dots from soybean whey on the neurocytotoxicity caused by Aβ40 aggregates, including Aβ40 alone, Aβ40 + different concentrations of carbon dots from soybean whey.

[0051] Figure 12 It is the effect of carbon dots from soybean whey on the production of reactive oxygen species caused by Aβ40 aggregates, including the situation of reactive oxygen species production after the cells were incubated alone for 24 hours, the cells + Aβ40 were incubated for 24 hours, and the cells + Aβ40 + 100 μg / mL carbon dots from soybean whey were incubated for 24 hours.

[0052] Figure 13 It is the fluorescence imaging map of carbon dots from soybean whey in the mouse brain.

[0053] Figure 14A is the high-resolution transmission electron microscope image of the soybean dregs carbon dots; Figure 14 B shows the aggregation of 40 μmol / L Aβ40 detected by the ThT fluorescence method and after adding different concentrations of soybean dregs carbon dots to Aβ40. Specific implementation manners

[0054] The present invention is not limited to the following specific implementation manners. Those of ordinary skill in the art can implement the present invention in other various specific implementation manners according to the content disclosed in the present invention, or those that adopt the design structure and idea of the present invention and make simple changes or modifications all fall within the protection scope of the present invention.

[0055] Raw material sources

[0056] Weigh 500 g of soybeans and soak them in water at room temperature (25 ± 1 °C) for 12 h according to a mass ratio of 1:3. The soaked soybeans are ground into raw soy milk with water at a mass ratio of 1:6 and then steamed until boiling to obtain cooked soy milk. The cooked soy milk is cooled to about 85 °C, and 68 mL of a magnesium chloride aqueous solution with a concentration of 0.05 mol / mL is added. Stir while adding until flocculent precipitation appears. Let the curdled soy milk stand for 25 min, and then press it into shape. The liquid generated during the pressing process is the soy whey.

[0057] Example 1: Preparation of soy whey carbon dots

[0058] Transfer the soy whey into a polytetrafluoroethylene inner liner and seal it in a stainless steel autoclave. Tighten the stainless steel autoclave lid, place the autoclave in an oven, and carry out a hydrothermal reaction at 180 °C for 12 h. After that, naturally cool it to room temperature, then centrifuge the reaction solution at a speed of 5000 r / min for 15 min. Then take the supernatant, filter it with a 0.22 μm filter membrane and retain the filtrate. Then use a 500 Da dialysis bag to dialyze the filtrate for 24 hours, and then freeze-dry the dialysate to obtain soy whey carbon dots.

[0059] Example 2: Characterization of soy whey carbon dots

[0060] Use a high-resolution transmission electron microscope to observe the morphology of the carbon dots prepared in Example 1. The obtained carbon dots have good dispersibility, and the average particle size is about 8.21 nm, as specifically shown in Figure 1 shown.

[0061] Use X-ray photoelectron spectroscopy to analyze the elemental composition of the carbon dots prepared in Example 1. From the Figure 2 XPS full spectrum of A, three sharp peaks of C1s (282.9 eV), N1s (400.1 eV) and O1s (534.4 eV) can be seen, indicating that the obtained carbon dots are composed of C, N and O elements. Figure 2B, 2C, and 2D are the high-resolution XPS spectra of C1s, N1s, and O1s, respectively.

[0062] Figure 3 It is the UV-Vis absorption spectrum of the carbon dots prepared from the soybean whey in Example 1. It can be seen from the figure that its absorption bands are located near 280 nm and 350 nm.

[0063] As Figure 4 shown, the prepared carbon dots from the soybean whey are light yellow under visible light and emit green fluorescence under ultraviolet light irradiation.

[0064] As Figure 5 shown, the maximum emission wavelength of the prepared carbon dots from the soybean whey continuously redshifts with the increase of the laser wavelength. When the excitation wavelength is 410 nm, its fluorescence intensity is the largest. Thus, it can be known that its maximum excitation wavelength is 410 nm and the maximum emission wavelength is 500 nm.

[0065] Prepare carbon dot dispersions with different pH conditions (1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, and 14) and incubate them, and then measure their fluorescence intensities at an excitation wavelength of 410 nm and an emission wavelength of 500 nm. The results are as Figure 6 shown. The fluorescence intensity of the carbon dots decreases under strong acidic (pH = 1 - 4) and strong basic (pH = 13 - 14) conditions, while it changes little in a relatively large range of pH = 5 - 12, indicating good stability.

[0066] The infrared spectrum of the carbon dots prepared from the soybean whey in Example 1 is as Figure 7 shown. It can be seen from the figure that the peak at 1421 cm -1 is caused by the bending vibration of C-H, the peak at 1616 cm -1 comes from the stretching vibration of C=O, the peak at 2932 cm -1 is caused by the stretching vibration of C-H, and the relatively broad peak in this range at 3383 cm -1 comes from the stretching vibration of O-H. The absorption peak at 1076 cm -1 is the absorption peak of the C-O bond.

[0067] Figure 8 It is the XRD spectrum of the carbon dots prepared from the soybean whey in Example 1. It can be seen from the figure that when the 2-Theta angle is 22.05°, there is a significant diffraction peak, indicating the existence of an amorphous carbon structure in the carbon dots.

[0068] Example 3: Application of Carbon Dots from Soybean Whey

[0069] 1. Inhibition of Aβ0 Aggregation by Carbon Dots from Soybean Whey

[0070] Preparation of Aβ40 monomer solution: Dissolve Aβ40 polypeptide powder in hexafluoroisopropanol solution at a concentration of 1 mg / mL. Place the solution in an ice bath and treat it with microwave ultrasonic waves for 1 h to remove pre-existing Aβ40 aggregates, and then let it stand in the ice bath for another 2 h to fully dissolve Aβ40. The standing solution is centrifuged at 4 °C and 15,000 g for 20 min to remove existing Aβ40 aggregates. After centrifugation, aspirate 75% of the supernatant into a centrifuge tube, and add 100 μL to each centrifuge tube. Use nitrogen to dry the liquid in the centrifuge tube, and Aβ40 is adsorbed on the inner wall of the centrifuge tube in the form of a thin film and stored at -20 °C until use. During the experiment, dissolve Aβ40 in a small amount of dimethyl sulfoxide and ultrasonicate it for 10 min to completely dissolve it, and then add PBS to prepare an Aβ40 solution with a final concentration of 40 μM for standby.

[0071] Determine the effect of soybean whey carbon dots on the aggregation of Aβ40 monomers. Mix different concentrations of soybean whey carbon dot solutions (5 μg / mL, 10 μg / mL, 25 μg / mL, 50 μg / mL, and 100 μg / mL) with Aβ40, and incubate them with shaking at 37 °C and 170 rpm for 24 hours. And use thioflavin T (25 μM) to label Aβ40, and measure the fluorescence intensity of the mixture of the sample and thioflavin T (λ EX : 445 nm, λ EM : 460 - 600 nm). Set the sample containing only Aβ40 as 100% to normalize other values, and repeat each experiment three times and take the average value. Figure 9 The fibrillation results of Aβ40 measured by thioflavin T are shown, that is, soybean whey carbon dots can significantly inhibit the fibrillation of Aβ40 monomers, and the higher the concentration of carbon dots, the more obvious the inhibitory effect.

[0072] 2. Inhibition of the formation of Aβ40 fibers by soybean whey carbon dots

[0073] Incubate Aβ40 and the Aβ40 + 100 μg / mL carbon dot mixture with shaking at 37 °C and 170 rpm for 24 hours. Then drop the solution onto mica sheets (10 μL) respectively, and perform atomic force microscopy detection after drying at room temperature. As Figure 10 shown in A, after Aβ40 was incubated alone for 24 hours, typical long and intertwined strip-like fibers (hundreds of nanometers long and several nanometers wide) appeared. When an equal volume of 100 μg / mL carbon dots was present, the aggregation morphology of Aβ40 changed significantly, and some short and loose fibers appeared, indicating that soybean whey carbon dots inhibited the formation of Aβ40 fibers.

[0074] 3. Inhibition of the neurocytotoxicity caused by Aβ40 aggregates by soybean whey carbon dots

[0075] Mouse hippocampal neuronal cells (HT22) were cultured in a CO2 incubator with the CO2 concentration controlled at 5% and the temperature at 37°C. Medium selection: DMEM, the medium contained 10% FBS and 100 U / mL of double antibiotics; when the cells reached about 80% confluence, trypsin was used to detach them from the culture dish. After being dispersed, they were counted under a microscope using a counter, and then transferred to a 96-well plate at a concentration of 10,000 cells per well; after culturing overnight in the CO2 incubator, the cells were checked under a microscope to see if they had adhered. After adhesion was completed, the medium in each well was removed, and carbon dots from soybean whey solution dissolved in the medium (5 μg / mL, 10 μg / mL, 25 μg / mL, 50 μg / mL, and 100 μg / mL), Aβ40, and Aβ40 + carbon dots from soybean whey solution (5 μg / mL, 10 μg / mL, 25 μg / mL, 50 μg / mL, and 100 μg / mL) were added. Then, the cells in the 96-well plate were placed in the CO2 incubator and cultured for 24 hours; again, the medium was removed, 100 μL of a solution containing 0.5 mg / mL of indigo carmine was added to each well, and after being placed back in the incubator and cultured for 4 hours, the upper layer of liquid was carefully removed. Then, 150 μL of DMSO solution was added to each well to dissolve the formazan produced by the reaction of live cells with MTT, and the cell viability was detected using an enzyme-linked immunosorbent assay (ELISA) reader with the ultraviolet wavelength set at 490 nm.

[0076] Figure 11 As shown in Figure A, the carbon dots from soybean whey solution itself have no toxicity to nerve cells; Figure 11 As shown in Figure B, when HT22 cells were treated with aggregated Aβ40 alone, the cell viability was 45.69%; while when Aβ40 and carbon dots from soybean whey solution at 5 μg / mL, 10 μg / mL, 25 μg / mL, 50 μg / mL, and 100 μg / mL were used to treat HT22 cells, the cytotoxicity was significantly reduced, and the cell viability increased to 78.21% (100 μg / mL).

[0077] 4. Effect of carbon dots from soybean whey solution on the ROS level induced by Aβ40 aggregates

[0078] The reactive oxygen species were detected using a reactive oxygen species detection kit with the fluorescent probe DCFH-DA. Mouse hippocampal neuronal cells (HT22) were cultured in a CO2 incubator with the CO2 concentration controlled at 5% and the temperature at 37°C. Medium selection: DMEM, the medium containing 10% FBS and 100 U / mL of double antibiotics; when the cells reached about 80% confluence, trypsin was used to detach them from the culture dish, and after being dispersed, they were transferred to a 6-well plate; after culturing overnight in the CO2 incubator, the cells were checked under a microscope for adherence. After the adherence was completed, the medium in each well was taken out, and Aβ40 and Aβ40 + soybean whey carbon dots dissolved in the medium (5 μg / mL, 10 μg / mL, 25 μg / mL, 50 μg / mL, and 100 μg / mL) were added. Then, the cells in the 6-well plate were placed in the CO2 incubator and cultured for 24 hours; again, the medium was taken out, 500 μL of 10 μmol / L DCFH-DA working solution was added to each well, and the plate was placed in the incubator and incubated for 20 min. The DCFH-DA working solution was collected, rinsed three times with PBS buffer, and the excess staining solution was washed away as much as possible. Finally, 1 mL of PBS buffer was added to each well, and photos were taken with a fluorescence inverted microscope. The process needed to be carried out in the dark, and finally, the results were calculated using Image J software.

[0079] As Figure 12 shown, compared with the control group, the green fluorescence intensity of DCFH-DA in the cells in the Aβ40 aggregate treatment group was stronger, indicating that the treatment of cells with Aβ40 aggregates significantly increased the content of intracellular ROS. After treatment with 100 μg / mL of soybean whey carbon dots, the green fluorescence intensity of DCFH-DA in the cells gradually decreased. This shows that soybean whey carbon dots have an inhibitory effect on the production of intracellular ROS caused by Aβ40 aggregates.

[0080] 5. Determination of the blood-brain barrier penetration ability of soybean whey carbon dots in mice

[0081] A carbon dot solution was prepared using sterile saline, and mice were gavaged (2 g / kg body weight). After 1, 2, 4, 8, and 24 hours, the mice were sacrificed and the whole brain tissues were taken out and imaged using a small animal in vivo imaging system.

[0082] As Figure 13 shown, after soybean whey carbon dots were gavaged to mice, they could cross the blood-brain barrier and be detected in the brain. Signals could be detected in the brain tissue 2 hours after gavage, and fluorescent signals could still be detected in the brain tissue at 24 hours. The above results indicate that the soybean whey carbon dots described in the present invention can cross the blood-brain barrier through oral administration.

[0083] Comparative Example 1

[0084] The fresh soybean dregs remaining after preparing soybean milk are evenly mixed with ultrapure water at a ratio of 1:4 to form a mixed solution. Transfer 60 mL of the mixed solution into a 100 mL polytetrafluoroethylene inner liner and seal it in a stainless-steel autoclave. Tighten the lid of the stainless-steel autoclave, place the autoclave in an oven, and carry out a hydrothermal reaction at 180 °C for 12 h. After completion, let it cool naturally to room temperature. Then, centrifuge the reaction solution at a rotational speed of 5000 r / min for 15 min. After that, take the supernatant, filter it using a 0.22 μm filter membrane, and retain the filtrate. Then, dialyze the filtrate using a 500 Da dialysis bag for 24 h. After that, freeze-dry the dialysate to obtain soybean dreg carbon dots.

[0085] The morphology of the soybean dreg carbon dots was observed using a high-resolution transmission electron microscope. The obtained carbon dots had good dispersibility, and the average particle size was about 16.28 nm, as shown in Figure 13 Figure A. Its particle size was larger than that of the soybean clear liquid carbon dots.

[0086] The effect of soybean dreg carbon dots on the aggregation of Aβ40 monomers was measured. Different concentrations of soybean dreg carbon dot solutions (5 μg / mL, 10 μg / mL, 25 μg / mL, 50 μg / mL, and 100 μg / mL) were mixed with Aβ40 and incubated with shaking at 37 °C and 170 rpm for 24 hours. And thioflavin T (25 μM) was used to label Aβ40, and the fluorescence intensity of the mixture of the sample and thioflavin T was measured (λ EX : 445 nm, λ EM : 460 - 600 nm). The sample containing only Aβ40 was set as 100% to normalize other values, and each experiment was repeated three times and the average value was taken. Figure 13 Figure B shows the fibrillation results of Aβ40 measured by thioflavin T, that is, soybean dreg carbon dots can significantly inhibit the fibrillation of Aβ40 monomers, and the higher the concentration of carbon dots, the more obvious the inhibitory effect. However, the inhibitory ability is less than that of the soybean clear liquid carbon dots at the same concentration.

[0087] Table 1

[0088] Soybean residue carbon dots Soybean clear liquid carbon dots 100 μg / mL 65.26％ 53.26％ 50 μg / mL 76.35％ 70.35％ 25 μg / mL 87.05％ 82.05％ 10 μg / mL 94.32％ 86.32％ 5 μg / mL 95.95％ 93.95％

[0089] The embodiments provided above are not intended to limit the scope covered by the present invention, nor are the described steps intended to limit the order of their execution. Obvious improvements made by those skilled in the art in combination with the existing well-known common sense also fall within the protection scope defined by the claims of the present invention.

Claims

1. A processing method for preparing carbon dots from soybean whey, characterized in that, It includes the following steps: Perform hydrothermal reaction on the soybean whey, cool it after completion, then centrifuge the cooled reaction solution, take the supernatant for filtration, then take the filtrate for dialysis, and then dry the dialysate to obtain soybean whey carbon dots.

2. The processing method according to claim 1, characterized in that, The soybean whey is the supernatant produced after retting during the fixed molding process of soy products.

3. The processing method according to claim 1, characterized in that, The preparation process of the soybean whey is as follows: Mix soybeans with water at a mass ratio of 1:2 - 5 and soak for 10 - 24 hours, then add water to the soaked soybeans at a mass ratio of 1:3 - 10 to make raw soy milk, heat the raw soy milk to obtain cooked soy milk, after the cooked soy milk cools to 75 - 90 °C, add magnesium chloride or an aqueous solution of magnesium chloride, the mass ratio of magnesium chloride to soybeans is 1:0.4 - 0.8, stir and then perform retting for 10 - 30 minutes, and the supernatant is the soybean whey.

4. The processing method according to claim 1, wherein The temperature of the hydrothermal reaction is 150 - 200 °C; the time of the hydrothermal reaction is 10 - 15 hours.

5. According to the processing method described in claim 1, characterized in that, The temperature of the cooling is 10 - 40 °C; the filtration is carried out using a filter membrane with a pore size of 0.2 - 0.3 μm.

6. The processing method according to claim 1, wherein The dialysis is carried out using a dialysis bag with a molecular weight cut-off of 300 - 1000 Da; the time of the dialysis is 12 - 48 hours.

7. A carbon dot of soybean clear liquid, characterized in that, The soybean whey carbon dots are prepared by the processing method according to any one of claims 1 - 6.

8. Use of the soybean clear liquid carbon dots described in claim 7 in brain imaging, characterized in that, The application in in-brain imaging refers to being used as a contrast agent for fluorescence imaging and photoacoustic imaging.

9. The application of the soybean whey carbon dots described in claim 7 in the preparation of a drug or drug carrier for preventing and treating Alzheimer's disease.

10. A carbon dot for realizing intracranial imaging and inhibiting the aggregation of Aβ40, characterized in that, The carbon dots are prepared from soybean whey as the raw material through hydrothermal reaction.

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