Cordyceps militaris polysaccharide fluorescent carbon dots capable of being used for detecting and degrading tetracycline antibiotics as well as preparation method and application of cordyceps militaris polysaccharide fluorescent carbon dots

Abstract

The invention discloses a cordyceps militaris polysaccharide fluorescent carbon dot capable of being used for detecting and degrading tetracycline antibiotics as well as a preparation method and application thereof, and belongs to the technical field of nano materials and environmental monitoring. The cordyceps militaris polysaccharide fluorescent carbon dots are prepared by the following steps: dissolving cordyceps militaris polysaccharide and zinc chloride in water, and then carrying out high-temperature reaction; and after the reaction is finished, filtering, dialyzing and drying to obtain the cordyceps militaris polysaccharide fluorescent carbon dots. The carbon quantum dot prepared by the invention realizes the integration of detection and degradation of tetracycline antibiotics, and has a series of advantages of simplicity in operation, high selectivity, low detection limit, environmental friendliness and the like, thereby having important value in the technical fields of nano materials and environmental monitoring.

Description

Technical Field

[0001] The invention belongs to the technical field of nanomaterials and environmental monitoring, and specifically relates to Cordyceps militaris polysaccharide fluorescent carbon dots that can be used for detecting and degrading tetracycline antibiotics, and a preparation method and application thereof. Background Art

[0002] Tetracycline antibiotics (TCs) are a typical class of broad-spectrum antibiotics, which are widely used to prevent and treat microbial diseases such as filariasis and giardiasis. At the same time, tetracycline can also be used to increase the yield of animal husbandry. However, the abuse of tetracycline can lead to serious consequences. Studies have shown that tetracycline is commonly used to treat a variety of diseases in mammals, but less than 50% of its dose is metabolized. A large part of tetracycline is discharged into water bodies and soil with animal feces and urine, causing pollution. Tetracycline remaining in animals will endanger human health through the food chain. Antibiotic residues in foods such as meat, milk and honey are highly toxic and can cause gastrointestinal disorders, allergies and central nervous system reactions. In order to deal with the above problems, many technologies for detecting tetracycline have been developed. Among the traditional tetracycline detection methods, the fluorescence method is widely used in the detection of tetracycline because of its high sensitivity, low detection limit and fast response. However, TCs actually include a variety of different antibiotics such as minocycline (MC), tetracycline (TC), oxytetracycline (OTC), doxycycline (DOC) and chlortetracycline (CTC). These antibiotics have similar structures but are difficult to detect simultaneously. Current research often only detects one specific antibiotic, and the detection methods for similar antibiotics are often complicated. Therefore, it is very important to find a simple and effective method to achieve the simultaneous detection of multiple tetracycline antibiotics. In addition, in practical applications, the detection and degradation of tetracycline antibiotics should complement each other, but there are very few related studies. Therefore, the development of a material that can be used to simultaneously detect and degrade tetracycline antibiotics faces huge challenges. Summary of the invention

[0003] The invention provides a Cordyceps militaris polysaccharide fluorescent carbon dot for detecting and degrading tetracycline antibiotics. The fluorescent carbon dot is a blue fluorescent quantum dot which takes Cordyceps militaris polysaccharide as a carbon source and is co-doped with zinc and chlorine.

[0004] The above Cordyceps militaris polysaccharide fluorescent carbon dots are prepared by the following method:

[0005] The Cordyceps militaris polysaccharide and zinc chloride are dissolved in water, and then subjected to a high temperature reaction; after the reaction is completed, the mixture is filtered, dialyzed, and dried to obtain the Cordyceps militaris polysaccharide fluorescent carbon dots.

[0006] In the above preparation method, each raw material is selected from the following parts by weight: 0.1-0.5 parts of Cordyceps militaris polysaccharide, 0.14-0.8 parts of zinc chloride, and 5-15 parts of water.

[0007] In a specific embodiment, the raw materials are selected from the following parts by weight: 0.12 parts of Cordyceps militaris polysaccharide, 0.6 parts of zinc chloride, and 10 parts of water.

[0008] In the above preparation method, the conditions of the high temperature reaction are selected from: hydrothermal reaction at 160-240°C for 8-14h; preferably: hydrothermal reaction at 180°C for 10h.

[0009] In the above preparation method, the filtration is performed using a 0.22 μm filter membrane; and the dialysis is performed using a dialysis bag with a molecular weight cut-off of 1000 Da.

[0010] The present invention provides the use of the Cordyceps militaris polysaccharide fluorescent carbon dots in detecting and / or degrading tetracycline antibiotics.

[0011] In the present invention, the tetracycline antibiotic is selected from one or more of minocycline (MC), tetracycline (TC), chlortetracycline (CTC), oxytetracycline (OTC), and doxycycline (DOC).

[0012] The present invention provides a method for detecting tetracycline antibiotics, comprising the following steps:

[0013] The Cordyceps militaris polysaccharide fluorescent carbon dots were mixed with tetracycline antibiotic standard solutions of different concentration gradients, and then the emission intensity was measured at an excitation wavelength of 366 nm, and the fluorescence intensity at the peak was recorded; the fluorescence intensity at the peak was used as the ordinate, and the tetracycline antibiotic concentration was used as the abscissa to draw a graph and obtain a linear equation;

[0014] Then, the test sample solution is mixed with the Cordyceps militaris polysaccharide fluorescent carbon dots, and the emission intensity is measured at an excitation wavelength of 366 nm. The fluorescence intensity at the peak of the test solution is recorded and substituted into the linear equation to obtain the concentration of tetracycline antibiotics in the test sample solution.

[0015] In the above detection method, the usage ratio of the Cordyceps militaris polysaccharide fluorescent carbon dots to the tetracycline antibiotic standard solution is 1 mg:1 mL; the usage ratio of the Cordyceps militaris polysaccharide fluorescent carbon dots to the sample solution to be tested is 1 mg:1 mL.

[0016] The present invention provides a detection method for distinguishing tetracycline antibiotics, comprising the following steps:

[0017] Cordyceps militaris polysaccharide fluorescent carbon dots were mixed with aqueous solutions of various tetracycline antibiotics of the same concentration, and then the color of the carbon dots in each solution was photographed under ultraviolet light using the colorimeter APP on a smartphone, and the obtained fluorescent colors were digitized and output to obtain RGB values; principal component analysis was performed using the obtained experimental data to obtain a PCA graph; PCA was used to analyze the differences in fluorescent colors;

[0018] The sample solution to be tested was mixed with the Cordyceps militaris polysaccharide fluorescent carbon dots, and then the color of the carbon dots in each solution under ultraviolet light was photographed using the colorimeter APP on a smartphone. The obtained fluorescent color was digitized and output to obtain the RGB value. The type of antibiotics in the sample solution to be tested was obtained after principal component analysis.

[0019] In the above detection method, the usage ratio of the Cordyceps militaris polysaccharide fluorescent carbon dots to the tetracycline antibiotic aqueous solution is 1 mg:1 mL; the usage ratio of the Cordyceps militaris polysaccharide fluorescent carbon dots to the sample solution to be tested is 1 mg:1 mL.

[0020] In the above detection method, the concentration of the tetracycline antibiotic aqueous solution is selected from 60 to 120 μmol / L.

[0021] The present invention provides a method for degrading tetracycline antibiotics, comprising the following steps:

[0022] The Cordyceps militaris polysaccharide fluorescent carbon dots were mixed with a sample solution containing tetracycline antibiotics and then subjected to light treatment to achieve the degradation of tetracycline antibiotics.

[0023] In the above degradation method, the usage ratio of the Cordyceps militaris polysaccharide fluorescent carbon dots to the sample solution is 1 mg:1 mL.

[0024] In the above-mentioned degradation method, a xenon lamp is used for illumination.

[0025] The method for detecting and degrading tetracycline antibiotics of the present invention is applicable to matrix samples such as aqueous solutions and foods, such as water, milk, honey, etc.; solid samples can be prepared in the form of aqueous solutions, or can be detected after filtering to remove water-insoluble components.

[0026] The beneficial effects of the present invention are:

[0027] The present invention uses Cordyceps militaris polysaccharide as a carbon source, doped with zinc chloride, and prepares a fluorescent carbon dot with a lattice spacing of about 0.33nm. The carbon dot will produce obvious fluorescence signal changes as the concentration of tetracycline antibiotics changes, and abundant color changes can be observed with the naked eye under ultraviolet light, thereby realizing sensitive detection of tetracycline antibiotics. At the same time, the fluorescent carbon dot can also be used as a catalyst for photocatalytic degradation to realize efficient degradation of tetracycline antibiotics. Thus, the fluorescent carbon dot can be used as a portable product to realize real-time, on-site visualization, high sensitivity, and environmental protection detection of tetracycline antibiotics (TCs), and realize rapid degradation of tetracycline antibiotics.

[0028] In addition, the fluorescent carbon dots prepared by the present invention can also be used to distinguish TCs of the same concentration by combining with a smartphone with an easily accessible color scanning application. The Colorimeter installed in the smartphone can convert the red, green and blue (RGB) channels of the fluorescent image into digital values, and utilize the rich color changes of the fluorescent carbon dots during the TCs detection process, without the need for expensive instruments and skilled operators, to quickly and conveniently achieve on-site visual detection and differentiation of TCs.

[0029] The main raw material Cordyceps militaris polysaccharide used in the present invention is a natural product extracted from nature, which avoids the introduction of traditional chemical agents. The design of this system embodies the sustainable development concept of "from nature, for nature, and integrating into nature", and provides technical support for tetracycline antibiotic pollution source control, environmental planning and governance, and public health and safety protection.

[0030] In summary, the carbon quantum dots prepared in the present invention realize the integrated detection and degradation of TCs, and have a series of advantages such as simple operation, easy to carry, high selectivity, low detection limit, and environmental friendliness, thus having important value in the field of nanomaterials and environmental monitoring technology. BRIEF DESCRIPTION OF THE DRAWINGS

[0031] Figure 1 The transmission electron microscopy image (including high-resolution image) (a) and particle size (b) of the Cordyceps militaris polysaccharide fluorescent carbon dots described in Example 2;

[0032] Figure 2 This is the fluorescence emission graph of the Cordyceps militaris polysaccharide fluorescent carbon dots described in Example 2;

[0033] Figure 3 This is the X-ray photoelectron spectrum of the Cordyceps militaris polysaccharide fluorescent carbon dots described in Example 2;

[0034] Figure 4 The fluorescence spectra and linear relationship diagram obtained by detecting different concentrations of antibiotics with Cordyceps militaris polysaccharide fluorescent carbon dots described in Example 2;

[0035] Figure 5 The mechanism of TC quenching the fluorescence carbon dots of Cordyceps militaris polysaccharide;

[0036] Figure 6 PCA diagram of Cordyceps militaris polysaccharide fluorescent carbon dots combined with smartphones for detecting tetracycline antibiotics;

[0037] Figure 7 The effects of different ions and amino acids on the mixed system of Cordyceps militaris polysaccharide fluorescent carbon dots;

[0038] Figure 8 This is the UV spectrum of the fluorescent carbon dots of Cordyceps militaris polysaccharide during the degradation of TCs;

[0039] Fig. 9 The degradation kinetics of Cordyceps militaris polysaccharide fluorescent carbon dots in the process of degrading TCs;

[0040] Fig.10 This is the mechanism of TCs degradation by Cordyceps militaris polysaccharide fluorescent carbon dots. DETAILED DESCRIPTION

[0041] In the present invention, Cordyceps militaris polysaccharide is prepared by the following method:

[0042] The Cordyceps militaris extract was dissolved in an ethanol-water solution and subjected to graded alcohol precipitation, wherein the volume ratio of water to ethanol was successively reduced from 4:1 to 1:1, and Cordyceps militaris polysaccharide was obtained after four times of graded alcohol precipitation.

[0043] In the present invention, the Cordyceps militaris extract was purchased from Xi'an Best Biotechnology Co., Ltd.

[0044] Other materials used in the present invention, if not otherwise stated, can be obtained through commercial channels. Other terms used in the present invention, unless otherwise specified, generally have the meanings commonly understood by those of ordinary skill in the art. The present invention is further described in detail below in conjunction with specific examples and with reference to data. The following examples are only for illustrating the present invention, and are not intended to limit the scope of the present invention in any way.

[0045] Example 1

[0046] Preparation of Cordyceps militaris polysaccharide fluorescent carbon dots, the steps are as follows:

[0047] 180 mg of Cordyceps militaris polysaccharide and 740 mg of zinc chloride were dissolved in 10 mL of ultrapure water and ultrasonicated for 10 min. The solution was thoroughly mixed, transferred to a 25 mL polytetrafluoroethylene-lined stainless steel autoclave, and kept at 180 ° C for 10 h. After cooling to room temperature, the resulting dark yellow solution was filtered through a 0.22 μm filter membrane, and the filtrate was further dialyzed in ultrapure water for 24 h with a dialysis bag (cut-off molecular weight of 1000 Da), and the water was renewed every 6 h to remove small molecules. Subsequently, freeze-dried for 24 h, Cordyceps militaris polysaccharide fluorescent carbon dot powder (Zn, Cl-CDs) was obtained.

[0048] Example 2

[0049] Preparation of Cordyceps militaris polysaccharide fluorescent carbon dots, the steps are as follows:

[0050] 120 mg of Cordyceps militaris polysaccharide and 600 mg of zinc chloride were dissolved in 10 mL of ultrapure water and ultrasonicated for 10 min. The solution was thoroughly mixed, transferred to a 25 mL polytetrafluoroethylene-lined stainless steel autoclave, and kept at 180 ° C for 10 h. After cooling to room temperature, the resulting dark yellow solution was filtered through a 0.22 μm filter membrane, and the filtrate was further dialyzed in ultrapure water for 24 h with a dialysis bag (cut-off molecular weight of 1000 Da), and the water was renewed every 6 h to remove small molecules. Subsequently, freeze-dried for 24 h, Cordyceps militaris polysaccharide fluorescent carbon dot powder (Zn, Cl-CDs) was obtained.

[0051] The transmission electron microscopy (TEM) and high-resolution images of the fluorescent carbon dots of Cordyceps militaris polysaccharide are shown in Figure 1. Figure 1 The morphology and microstructure of the fluorescent carbon dots of Cordyceps militaris polysaccharide were characterized by TEM. Figure 1 It can be seen that the Cordyceps militaris polysaccharide fluorescent carbon dots are uniformly distributed spherical nanoparticles with a diameter range of 3.0-5.0nm and an average particle size of 4.3nm. Further careful observation from the HRTEM image shows that the Cordyceps militaris polysaccharide fluorescent carbon dots show obvious lattice fringes with a lattice spacing of 0.33nm.

[0052] In order to explore the fluorescence properties of Cordyceps militaris polysaccharide fluorescent carbon dots, the fluorescence emission spectra of Cordyceps militaris polysaccharide fluorescent carbon dots at different excitation wavelengths of 290-400 nm were tested by fluorescence spectroscopy. Figure 2 As shown, it can be observed that the Cordyceps militaris polysaccharide fluorescent carbon dots obtain the strongest fluorescence emission at 430nm under 360nm excitation.

[0053] In order to further confirm the elemental composition and chemical bonds of the fluorescent carbon dots from Cordyceps militaris polysaccharide, XPS measurements were performed, such as Figure 3 As shown. In the full spectrum scan, five main peaks were found: C1s, O 1s, N 1s, Cl 2p, and Zn 2p ( Figure 3a). Cordyceps militaris polysaccharide fluorescent carbon dots are composed of C (71.6%), O (19.9%), N (3.9%), Zn (2.5%) and Cl (2.1%) ( Figure 3 a), indicating that zinc and chlorine co-doped carbon quantum dots were successfully synthesized. Figure 3 As shown in b, the high-resolution Zn 2p peaks can be divided into two peaks, which correspond to Zn-O (1045.29 eV, 1022.37 eV). Figure 3 The three peaks in c) correspond to C=O (288.75 eV), CO / CN (286.69 eV), and CC (284.80 eV). Figure 3 d) is divided into three peaks, corresponding to Cl 2p3 / 2 (201.55eV, 198.38eV) and Cl 2p1 / 2 (199.89eV). High resolution N1s ( Figure 3 The two peaks in e) correspond to NH (401.69 eV) and NC (400.03 eV). High-resolution O1s ( Figure 3 The two peaks in f) correspond to HOC (533.60 eV) and CO (532.05 eV), respectively.

[0054] Example 3

[0055] Preparation of Cordyceps militaris polysaccharide fluorescent carbon dots, the steps are as follows:

[0056] 200 mg of Cordyceps militaris polysaccharide and 800 mg of zinc chloride were dissolved in 10 mL of ultrapure water and ultrasonicated for 10 min. The solution was thoroughly mixed, transferred to a 25 mL polytetrafluoroethylene-lined stainless steel autoclave, and kept at 180 ° C for 10 h. After cooling to room temperature, the resulting dark yellow solution was filtered through a 0.22 μm filter membrane, and the filtrate was further dialyzed in ultrapure water for 24 h with a dialysis bag (cut-off molecular weight of 1000 Da), and the water was renewed every 6 h to remove small molecules. Subsequently, freeze-dried for 24 h, Cordyceps militaris polysaccharide fluorescent carbon dot powder (Zn, Cl-CDs) was obtained.

[0057] 1. Design of the test for tetracycline antibiotics

[0058] Add 4.0 mg of Cordyceps militaris polysaccharide fluorescent carbon dots prepared in Example 2 to 4 mL of antibiotic aqueous solution of different concentrations (0-100 μM) and mix well. Antibiotics include tetracycline antibiotics (MC, TC, CTC, OTC, DOC) and non-tetracycline antibiotics (penicillin (PNG), erythromycin (ETC), ceftriaxone sodium (CRO), kanamycin (KAN)). Then transfer to a cuvette, measure its emission intensity at an excitation wavelength of 366 nm, and record the emission spectrum of the mixture. The slit widths for emission and excitation are both 10 nm. Plot the recorded fluorescence spectrum test data and obtain its linear relationship, as shown in FIG. Figure 4 shown.

[0059] Depend on Figure 4 It can be seen that with the increase of tetracycline antibiotic concentration, the fluorescence intensity of Cordyceps militaris polysaccharide fluorescent carbon dot system at 434nm decreased significantly, and as the concentration continued to increase, it was accompanied by varying degrees of red shift. Figure 4 The small figures above in the middle bf figures show from left to right the fluorescence color changes of the same mass of Cordyceps militaris polysaccharide fluorescent carbon dots (4.0 mg) when TCs solutions with concentrations of 10 μM, 20 μM, 30 μM, 40 μM, 50 μM, 60 μM, 80 μM, and 100 μM were added respectively. It can be seen that with the increase of TCs concentration, the fluorescence of the Cordyceps militaris polysaccharide fluorescent carbon dots system changes from blue to blue-green and then to green.

[0060] With the fluorescence intensity at the peak as the ordinate and the TCs concentration as the abscissa, the linear equation is obtained as follows: Figure 4 As shown in the small and medium figures, they are as follows: F = -57.30C MC +3665.88, F=-54.71C TC +3561.24, F=-55.98C OTC +3849.01, F=-46.72C DOC +3916.90, F=-59.29C CTC +3991.36, linear correlation coefficient R 2 >0.976.

[0061] The sample solution to be tested and the Cordyceps militaris polysaccharide fluorescent carbon dot solution synthesized in Example 2 are mixed in a ratio of 1:1 (mg:mL), and after mixing evenly; the fluorescence intensity at the peak in the test solution is obtained by fluorescence spectroscopy under excitation at 366nm, and the content of tetracycline antibiotics in the sample solution to be tested can be calculated by comparing the linear equation obtained in the above steps.

[0062] The detection limit (LOD) of Cordyceps militaris polysaccharide fluorescent carbon dots for TCs was calculated using the equation LOD=3Sb / s, where Sb represents the standard error of 10 consecutive scans of the blank sample and s represents the slope of the calibration curve. The results showed that the detection limit of the above method was 3.5-7.3 nM.

[0063] 2. Detection of Tetracycline Antibiotics in Milk

[0064] Before the experiment, the milk samples were pre-treated, i.e., the protein precipitation procedure was performed: an equal volume (5 mL) of milk and 1% (v / v) trichloroacetic acid solution were thoroughly mixed, and then the mixed sample was ultrasonically treated at 40°C for 20 min and transferred to a centrifuge tube. After centrifugation at 12000 rpm for 8 min, the milk extraction supernatant was obtained. The pH was adjusted to neutral with sodium hydroxide aqueous solution.

[0065] Tetracycline antibiotics were added to the milk extraction supernatant, and a series of standard milk sample solutions containing different concentrations of TCs (0-50 μM) were prepared.

[0066] The above-mentioned standard milk sample solutions containing different concentrations of tetracycline antibiotics were taken respectively, and the Cordyceps militaris polysaccharide fluorescent carbon dots prepared in Example 2 were added thereto at a ratio of 1:1 (mg:mL); after mixing evenly, the solutions were incubated at room temperature for 30 minutes, the emission spectrum of the carbon dots was measured with an excitation intensity of 366 nm, and the color change of the carbon dots was observed under ultraviolet light.

[0067] The fluorescence intensity at the peak is used as the ordinate and the antibiotic concentration is used as the abscissa to obtain the relationship between the fluorescence intensity at the peak and the antibiotic concentration in the milk sample.

[0068] After the milk sample solution to be tested has been treated by the above sample pretreatment step, it is mixed with the Cordyceps militaris polysaccharide fluorescent carbon dot solution synthesized in Example 2 in a ratio of 1:1 (mg:mL) and mixed evenly; incubated at room temperature for 30 minutes, the color change of the carbon dots is observed under ultraviolet light, and the fluorescence spectrum is measured; and the content of tetracycline in the milk sample solution to be tested is calculated by referring to the linear equation obtained in the above steps.

[0069] The test results are shown in Table 1:

[0070] Table 1. Recovery results of spiked milk samples

[0071]

[0072]

[0073] As shown in Table 1, the Cordyceps militaris polysaccharide fluorescent carbon dots prepared by the present invention are used to detect tetracycline antibiotics in milk, which has good stability and high sensitivity.

[0074] 3. Detection mechanism of Cordyceps militaris polysaccharide fluorescent carbon dots

[0075] In order to clarify the mechanism of TC-induced quenching of the fluorescent carbon dots of Cordyceps militaris polysaccharides, the excitation spectrum (Ex of Zn, Cl-CDs) and emission spectrum (Em of Zn, Cl-CDs) of the fluorescent carbon dots of Cordyceps militaris polysaccharides and the UV-visible absorption spectrum of TC were detected. Figure 5 As shown, the TC absorption band at 357 nm partially overlaps with the excitation spectrum of the Cordyceps militaris polysaccharide fluorescent carbon dots. The results show that the TC-induced fluorescence quenching of the Cordyceps militaris polysaccharide fluorescent carbon dots is a stable quenching mechanism based on the inner filter effect (IFE).

[0076] The present invention uses Cordyceps militaris polysaccharide as a carbon source, adds ZnCl2 and water, and adopts a one-step hydrothermal method to prepare blue luminescent zinc and chlorine co-doped carbon quantum dots (Zn, Cl-CDs). After the carbon quantum dots act on tetracycline antibiotics (TCs), based on the internal filter effect (IFE), the blue fluorescence of the carbon quantum dots is gradually quenched, and the characteristic green fluorescence of doped zinc and chlorine elements is gradually enhanced, resulting in a change in the fluorescence signal based on the type and content of tetracycline antibiotics. The fluorescent carbon dots have a recognition space specifically matched with the TCs molecule. After identifying TCs, the blue fluorescence emission spectrum of the fluorescent carbon dots at 434nm is gradually quenched, and red shifts to varying degrees occur due to different types of TCs. Highly selective visual on-site detection and differentiation of TCs molecules can be achieved.

[0077] 4. Detection of tetracycline antibiotics by combining Cordyceps militaris polysaccharide fluorescent carbon dots with smartphones

[0078] 4.0 mg of the Cordyceps militaris polysaccharide fluorescent carbon dots prepared in Example 2 were added to 4 mL of an aqueous solution of tetracycline antibiotics of the same concentration (100 μM) and mixed evenly.

[0079] Then the mixed solution was transferred to a cuvette, and a color scanning application (APP: colorimeter) was downloaded on a smartphone through the app store to photograph the color of the carbon dots in the standard solution of the same tetracycline antibiotic concentration under ultraviolet light, and the obtained fluorescence color was digitized and output to obtain the RGB value; the principal component analysis was performed using the obtained experimental data to obtain a PCA graph, as shown in Figure 6 shown.

[0080] Figure 6 a shows that at this concentration, the fluorescence colors of tetracycline antibiotics are significantly different, and the type of tetracycline antibiotics can be determined by visually identifying the fluorescence color. Figure 6 b shows that the change of fluorescence color of tetracycline antibiotics with concentration is a gradual process, and the change is obvious within 0 to 50 μM.

[0081] The Cordyceps militaris polysaccharide fluorescent carbon dots prepared in Example 2 were mixed with the sample solution to be tested (containing TC, MC, OTC, respectively, with a concentration of 100 μM) in a ratio of 1: 1 (mg: mL), and mixed evenly; the fluorescent color of the gel balls in the solution to be tested under ultraviolet light was photographed using a color scanning application of a smart phone, and digitized and output to obtain RGB values; a PCA graph was obtained after principal component analysis. The result was compared with the PCA graph of a 100 μM tetracycline antibiotic standard solution, and the type of antibiotic was accurately determined according to the location of the data. At the same time, the fluorescent color of the solution to be tested under 365 nm ultraviolet irradiation was observed with the naked eye and the type of antibiotic was successfully determined.

[0082] 5. Effects of different ions and amino acids on the mixed system of Cordyceps militaris polysaccharide fluorescent carbon dots

[0083] The Cordyceps militaris polysaccharide fluorescent carbon dot prepared by 4.0mg embodiment 2 is added to 4mL tetracycline aqueous solution (100μM), and 2000μL of different ions and amino acid aqueous solution (concentration 50μM) are added respectively, and the Cordyceps militaris polysaccharide fluorescent carbon dot mixed system without adding any ions and amino acids is used as a control (Blank).Then the mixed solution is transferred to a cuvette, and the emission intensity is measured with an excitation wavelength of 366nm, and the emission spectrum of the mixture is recorded. The slit width of emission and excitation is 10nm.Record experimental data calculates F / F0, and makes a bar graph.

[0084] The test results are as follows Figure 7 As shown:

[0085] Depend on Figure 7 It can be seen that ions and amino acids have little effect on the fluorescence intensity of the samples.

[0086] 6. Photocatalytic degradation of tetracycline antibiotics

[0087] 4.0 mg of the Cordyceps militaris polysaccharide fluorescent carbon dots prepared in Example 2 were added to 4 mL of the same concentration (100 μM) of each tetracycline antibiotic aqueous solution and mixed evenly. The mixture was placed in a dark environment and kept away from light for 24 h. The mixed solution was then transferred to a watch glass, which was then placed under a xenon lamp for uniform illumination for 24 min, and the ultraviolet spectrum of the solution was recorded in real time, as shown in FIG. Figure 8 shown.

[0088] Combined with the standard curve of tetracycline antibiotic concentration and absorbance, the concentration of tetracycline antibiotics is indicated by the absorbance at the peak around 370nm, and the concentration of tetracycline antibiotics c is obtained in real time. t , the initial concentration is set to c0;

[0089] The experimental data obtained were sorted and plotted as ln c t / c0 as the ordinate and time as the abscissa, and obtain the time and ln c t / c0 linear equation, such as Fig. 9 shown.

[0090] The results of photocatalytic degradation of tetracycline antibiotics are shown in Table 2:

[0091] Table 2 Degradation rate of tetracycline antibiotics

[0092]

[0093]

[0094] From the above table and picture results, we can see that Cordyceps militaris polysaccharide fluorescent carbon dots can achieve rapid degradation of tetracycline antibiotics within 24 minutes, and its degradation rate conforms to pseudo-first-order kinetics, with a correlation coefficient R 2 >0.986.

[0095] 7. Degradation of Tetracycline Antibiotics in Milk

[0096] Before the experiment, the milk was pretreated, i.e., the protein precipitation procedure: an equal volume (5 mL) of milk and a 1% (v / v) trichloroacetic acid solution were completely mixed, and then the mixed sample was ultrasonically treated at 40°C for 20 min and transferred to a centrifuge tube. After centrifugation at 12000 rpm for 8 min, the milk extract supernatant was obtained. The pH was adjusted to neutral with an aqueous sodium hydroxide solution. Each tetracycline antibiotic was added to the above-mentioned milk extract supernatant, and a series of standard milk sample solutions containing different concentrations of TCs (0-100 μM) were prepared. Then, the Cordyceps militaris polysaccharide fluorescent carbon dots prepared in Example 2 were added thereto at a ratio of 1:1 (mg:mL); after mixing evenly; incubated at room temperature for 30 min, and the ultraviolet spectrum was measured. The concentration of tetracycline antibiotics is indicated by the absorbance at the peak around 370 nm, and the concentration c of tetracycline antibiotics is obtained in real time. t , the initial concentration is set to c0.

[0097] The obtained experimental data were arranged and plotted, with absorbance as the ordinate and antibiotic concentration as the abscissa, to obtain a linear equation of antibiotic concentration and absorbance.

[0098] Then, the milk sample solution to be tested was treated by the above sample pretreatment step, and then mixed with the Cordyceps militaris polysaccharide fluorescent carbon dots prepared in Example 2 in a ratio of 1:1 (mg:mL), and after mixing evenly, incubated at room temperature for 30 minutes, the mixed solution was transferred to a watch glass, and then placed in a xenon lamp for uniform illumination for 24 minutes, and the ultraviolet spectrum of the solution was recorded. The degradation rate of tetracycline antibiotics was calculated in combination with the above standard curve.

[0099] The test results are shown in Table 3:

[0100] Table 3 Degradation rate of tetracycline antibiotics in milk samples

[0101]

[0102]

[0103] As shown in Table 3, the Cordyceps militaris polysaccharide fluorescent carbon dots have an obvious degradation effect on TCs in milk samples.

[0104] 8. Degradation mechanism of Cordyceps militaris polysaccharide fluorescent carbon dots

[0105] In order to study the degradation mechanism of tetracycline antibiotics by fluorescent carbon dots of Cordyceps militaris polysaccharide, tert-butyl alcohol (TBA), benzoquinone (BQ), furfuryl alcohol (FFA) and ammonium oxalate (AO) were added as ·OH, O 2- · 1 O2 and H + The effects of different free radicals on the degradation rate of tetracycline antibiotics were studied using quenchers. The specific steps are as follows:

[0106] The Cordyceps militaris polysaccharide fluorescent carbon dots prepared by 4.0mg embodiment 2 are added to the tetracycline antibiotic aqueous solution of 4mL same concentration (100μM), and mixed. The type of tetracycline antibiotics is the same, and tert-butyl alcohol (TBA), benzoquinone (BQ), ammonium oxalate (AO), furfuryl alcohol (FFA) are added respectively, and the reagent concentration added is 3.2μM, and the solution without adding any quencher is used as a control group. The resulting solution is placed in a dark environment dark treatment 24h. The solution is transferred to a watch dish, and then it is placed in a xenon lamp for uniform illumination, and the illumination time is 24min, and the ultraviolet spectrum of the solution is recorded in real time. In conjunction with the standard curve of tetracycline antibiotics concentration and absorbance, the concentration of tetracycline antibiotics is indicated with the absorbance at the peak value of about 370nm, and the degradation rate after illumination is calculated.

[0107] The test results are as follows Fig.10 As shown:

[0108] Depend on Fig.10 It can be seen that the mechanism of degradation of TCs by Cordyceps militaris polysaccharide fluorescent carbon dots is ·OH, O 2-· 1 O2 and H + The result of a combination of factors, but the main factor is 1 O2. The extra positive charge inside the carbon dots leads to more unpaired electrons, promoting singlet oxygen ( 1 O2) formation. 1 The long lifetime of O2 is beneficial for the degradation of dilute solutions, thus explaining the high degradation rate of C. militaris polysaccharide fluorescent carbon dots.

[0109] The above is only a preferred embodiment of the present invention, and does not limit the present invention in other forms. Any technician familiar with the profession may use the above disclosed technical content to change or modify it into an equivalent embodiment with equivalent changes. However, any simple modification, equivalent change and modification made to the above embodiment according to the technical essence of the present invention without departing from the technical solution of the present invention still belongs to the protection scope of the technical solution of the present invention.

Claims

1. A Cordyceps militaris polysaccharide fluorescent carbon dot for detecting and degrading tetracycline antibiotics, characterized in that: The fluorescent carbon dots are blue fluorescent quantum dots which use Cordyceps militaris polysaccharide as a carbon source and are co-doped with zinc and chlorine.

2. The Cordyceps militaris polysaccharide fluorescent carbon dots according to claim 1, characterized in that: The Cordyceps militaris polysaccharide fluorescent carbon dots are prepared by the following method: The Cordyceps militaris polysaccharide and zinc chloride are dissolved in water, and then subjected to a high temperature reaction; after the reaction is completed, the mixture is filtered, dialyzed, and dried to obtain the Cordyceps militaris polysaccharide fluorescent carbon dots.

3. The Cordyceps militaris polysaccharide fluorescent carbon dots according to claim 2, characterized in that: The raw materials are selected from the following parts by weight: 0.1-0.5 parts of Cordyceps militaris polysaccharide, 0.14-0.8 parts of zinc chloride, and 5-15 parts of water.

4. The Cordyceps militaris polysaccharide fluorescent carbon dots according to claim 2, characterized in that: The conditions of the high temperature reaction are selected from: hydrothermal reaction at 160-240° C. for 8-14 hours.

5. Use of the Cordyceps militaris polysaccharide fluorescent carbon dots according to any one of claims 1 to 4 in detecting and / or degrading tetracycline antibiotics.

6. The use according to claim 5, characterized in that: The tetracycline antibiotic is selected from one or more of minocycline, tetracycline, chlortetracycline, oxytetracycline and doxycycline.

7. A method for detecting tetracycline antibiotics, characterized in that: The steps include: The Cordyceps militaris polysaccharide fluorescent carbon dots according to any one of claims 1 to 4 are mixed with tetracycline antibiotic standard solutions of different concentration gradients, and then the emission intensity thereof is measured at an excitation wavelength of 366 nm, and the fluorescence intensity at the peak is recorded; a graph is drawn with the fluorescence intensity at the peak as the ordinate and the tetracycline antibiotic concentration as the abscissa to obtain a linear equation; Then, the test sample solution is mixed with the Cordyceps militaris polysaccharide fluorescent carbon dots, and the emission intensity is measured at an excitation wavelength of 366 nm. The fluorescence intensity at the peak of the test solution is recorded and substituted into the linear equation to obtain the concentration of tetracycline antibiotics in the test sample solution.

8. A detection method for distinguishing tetracycline antibiotics, characterized in that: The steps include: The Cordyceps militaris polysaccharide fluorescent carbon dots according to any one of claims 1 to 4 are mixed with aqueous solutions of various tetracycline antibiotics of the same concentration, and then the color of each carbon dot solution under ultraviolet light is photographed using a colorimeter APP on a smart phone, and the obtained fluorescent color is digitized and output to obtain an RGB value; principal component analysis (PCA) is performed using the obtained experimental data to obtain a PCA graph; and the difference in fluorescent color is analyzed using PCA; The sample solution to be tested was mixed with the Cordyceps militaris polysaccharide fluorescent carbon dots, and then the color of the carbon dots in each solution under ultraviolet light was photographed using the colorimeter APP on a smartphone, and the obtained fluorescent color was digitized and output to obtain the RGB value; The types of antibiotics in the sample solution were obtained after principal component analysis.

9. A method for degrading tetracycline antibiotics, characterized in that: The steps include: The Cordyceps militaris polysaccharide fluorescent carbon dots according to any one of claims 1 to 4 are mixed with a sample solution containing tetracycline antibiotics, and then subjected to light treatment to achieve degradation of the tetracycline antibiotics.

10. The degradation method according to claim 9, characterized in that: Xenon lamp was used for illumination.

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