A method for detecting glyphosate in coffee by Cu-CDs / ABEI@Ag nanozyme catalytic chemiluminescence

Abstract

The application discloses a method for detecting glyphosate in coffee by Cu-CDs / ABEI@Ag nanozyme catalytic chemiluminescence, and belongs to the technical field of chemical analysis and detection. 2+ The application is characterized in that 4-aminoantipyrine is used as a precursor, Cu is used as a doping element, Cu-doped carbon dots (Cu-CDs) are synthesized by a microwave method, Cu-CDs and ABEI are used as reducing agents to synthesize Cu-CDs / ABEI@Ag nanozyme, the Cu-CDs / ABEI@Ag nanozyme has excellent peroxidase-like (POD-like) activity, promotes the decomposition of H2O2, and enhances a chemiluminescence signal; glyphosate can interact with the Cu-CDs / ABEI@Ag nanozyme to inhibit the POD-like activity of the nanozyme, so that the chemiluminescence intensity generated by the oxidation of ABEI is reduced; and based on this, the method is used for detecting glyphosate in coffee samples, and has the characteristics of high specificity, simple operation and high sensitivity.

Description

TECHNICAL FIELD

[0001] The application belongs to the technical field of chemical analysis and detection, and particularly relates to a method for detecting glyphosate residues in coffee by Cu-CDs / ABEI@Ag nanoenzyme catalytic chemiluminescence. BACKGROUND

[0002] Glyphosate (PMG), also known as N-(phosphonomethyl)glycine, is a non-selective and broad-spectrum insecticide of organic phosphorus pesticide that is currently used most widely. Glyphosate has become one of the most widely used herbicides due to its low production cost, high efficacy, and lower toxicity to mammals than other insecticides, which has caused environmental pollution. Recent studies have shown that the residues of glyphosate in the environment and agricultural products not only pose ecological and agricultural risks to humans and animals, but also are a serious public health problem. Therefore, many countries or organizations have stipulated the maximum residue limit (MRL) of glyphosate in agricultural products and food, and the WHO has stipulated that the maximum residue level in drinking water is 0.90 mg / L. At present, various analytical methods such as HPLC, LC-MS / MS, ELISA, electrochemistry, ion chromatography, chemiluminescence (CL) and colorimetry have been used to detect the content of glyphosate. Among them, colorimetry and CL sensors have attracted widespread attention due to their rapidness, simple equipment, high sensitivity, low cost and other advantages. However, colorimetry is greatly challenged when the matrix of the detection object has a deep color.

[0003] At present, the chemiluminescence (CL) technology based on nanoenzymes is widely used in the sensing field of glyphosate due to its high sensitivity and signal-to-noise ratio and less color interference of the sample itself, but it is still very important to improve the detection sensitivity and selectivity. SUMMARY

[0004] The application provides a method for detecting glyphosate residues in coffee by Cu-CDs / ABEI@Ag nanoenzyme catalytic chemiluminescence. 2+ Cu-CDs, Cu-CDs / ABEI@Ag nanoenzyme and Cu-CDs / ABEI@Ag nanoenzyme catalytic chemiluminescence detection of glyphosate residues in coffee - and 1O2, enhanced chemiluminescence (CL) signal. Since glyphosate (PMG) can interact with Cu-CDs / ABEI@Ag and inhibit the POD-like activity of nanozyme, thereby reducing the CL intensity generated by ABEI oxidation. Based on this high-sensitivity inhibition response of CL, a CL nanosensor for glyphosate detection was developed, with a detection limit as low as 3.87 μg / L. The established method was used for glyphosate detection in coffee samples, and due to the high anti-interference ability of the CL method, satisfactory standard addition recoveries were obtained, highlighting the application prospects of the sensor in on-site glyphosate monitoring.

[0005] The method for detecting glyphosate residues in coffee based on Cu-CDs / ABEI@Ag nanozyme catalytic chemiluminescence is as follows:

[0006] 1. Dissolve 0.70-1.0 g of 4-aminoantipyrine and 0.6-0.8 g of CuCl2·2H2O in 20-30 mL of deionized water, mix and dissolve by ultrasonic treatment, then mix the solution under microwave at 180℃ for 100-150 min. After cooling, centrifuge the reaction product, pass the supernatant through a 0.22 mm filter membrane, and vacuum dry to obtain copper-doped carbon dots Cu-CDs;

[0007] The microwave power is 1.0-1.2 kW, and the centrifugation is at 8000-10000 r / min for 5-15 min;

[0008] 2. Mix 50-100 μL of 0.2 mg / mL Cu-CDs solution, 200-400 μL of 1 mg / mL ABEI solution, 300-500 μL of 11 mg / mL AgNO3 solution, and 10-15 mL of deionized water, stir at room temperature for 20-30 min, and vacuum dry to obtain Cu-CDs / ABEI@Ag nanozyme;

[0009] 3. Mix Cu-CDs / ABEI@Ag nanozyme solution, ABEI solution, H2O2, and different concentrations of glyphosate solution, add pH 11 Na2CO3-NaHCO3 buffer solution, mix well, and then use an enzyme marker to measure the chemiluminescence intensity and calculate ΔI CL , to determine the linear relationship between glyphosate concentration and ΔI CL , and obtain the regression equation, where ΔI CL = I-I0, I and I0 are the chemiluminescence intensities of the reaction system without adding glyphosate and with adding glyphosate, respectively;

[0010] The concentration of the Cu-CDs / ABEI@Ag nanoscale enzyme solution is 2 mg / mL, the added amount is 10-20 μL; the concentration of the ABEI solution is 10 mmol / L, the added amount is 40-60 μL; the concentration of the H2O2 solution is 50 mmol / L, the added amount is 20-40 μL; the concentration of the pH 11 Na2CO3-NaHCO3 buffer solution is 0.1 mol / L, the added amount is 50-100 μL;

[0011] 4. The chemiluminescence intensity of the sample to be tested is determined according to the method of step 3, and ΔI is calculated CL , and the concentration of glyphosate in the sample to be tested is obtained by substituting the regression equation.

[0012] The present application has the advantages that:

[0013] 1. The present application uses 4-aminoantipyrine as a precursor, Cu 2+ as a doping element, and synthesizes copper-doped carbon dots (Cu-CDs) by a microwave method. Cu-CDs and ABEI are used as reducing agents to synthesize Cu-CDs / ABEI@Ag nanoscale enzymes. The Cu-CDs / ABEI@Ag nanoscale enzymes have excellent peroxidase-like (POD-like) activity, promote the decomposition of H2O2 to generate ·OH, ·O2 - and 1 O2, ABEI is incorporated into the catalyst, which increases the concentration of the CL reagent in the catalyst structure and reduces the spatial distance between the catalyst and ABEI, thereby enhancing the CL signal. Based on the interaction between glyphosate and Cu-CDs / ABEI@Ag, the present application establishes a new method for chemiluminescence detection of glyphosate;

[0014] 2. The method for chemiluminescence detection of glyphosate established by the present application has a wide linear range of 6.67-333.33 μg / L and a low detection limit of 3.87 μg / L. The method is used for the detection of glyphosate in coffee samples, and the glyphosate in the coffee samples is successfully detected by the standard addition method, with a recovery rate of 95.86-111.13%. The detection system provided by the present application has high accuracy.

[0015] In summary, the method of the present application has the characteristics of strong specificity, simple operation and high sensitivity. BRIEF DESCRIPTION OF DRAWINGS

[0016] Figure 1 TEM (a) and HRTEM (b) images of the Cu-CDs / ABEI@Ag nanoscale enzyme in Example 1;

[0017] Figure 2 is an FTIR graph of the Cu-CDs / ABEI@Ag nanoscale enzyme in Example 1;

[0018] Figure 3 shows the Cu 2p (a) and Ag 3d (b) XPS images of the Cu-CDs / ABEI@Ag nanozyme in Example 1;

[0019] Figure 4 shows the UV-Vis and fluorescence spectra of TMB, ABTS, and OPD oxidized by Cu-CDs / ABEI@Ag+H2O2 in Example 1.

[0020] Figure 5 This is a steady-state kinetic diagram of the Cu-CDs / ABEI@Ag nanozyme in Example 1 when H2O2 is a variable.

[0021] Figure 6 This is a steady-state kinetic diagram of the Cu-CDs / ABEI@Ag nanozyme in Example 1 when TMB is a variable;

[0022] Figure 7 The graph shows the chemiluminescence performance results of ABEI+H2O2+Cu-CDs / ABEI@Ag, ABEI+Cu-CDs / ABEI@Ag, and ABEI+H2O2+PMG+Cu-CDs / ABEI@Ag in Example 1.

[0023] Figure 8 Fourier transform infrared spectra of Cu-CDs / ABEI@Ag nanozyme, Cu-CDs / ABEI@Ag+PMG, and PMG+Cu-CDs / ABEI@Ag after water washing in Example 1;

[0024] Figure 9 The UV-Vis spectra of PMG, PMG+Cu-CDs / ABEI@Ag, and H2O2+PMG+Cu-CDs / ABEI@Ag in Example 1 are shown below.

[0025] Figure 10 This is a free radical capture diagram of the H2O2+ABEI+Cu-CDs / ABEI@Ag system in Example 1;

[0026] Figure 11 This is a free radical capture diagram of the ABEI+H2O2+PMG+Cu-CDs / ABEI@Ag system in Example 1;

[0027] Figure 12 Figure a shows the CL spectrum and linear regression equation of PMG in Example 1. Figure a is the chemiluminescence intensity, and Figure b is the standard curve.

[0028] Figure 13 The results show the effects of major interfering substances and organophosphorus pesticides in coffee on the PMG detection system.

[0029] Figure 14This study illustrates the effect of different coffee samples on the chemiluminescence signal of the H2O2+ABEI+Cu-CDs / ABEI@Ag system in Example 1. Detailed Implementation

[0030] The following examples further illustrate the content of the present invention, but these examples do not limit the scope of protection of the present invention. Unless otherwise specified, the methods in the examples are conventional methods, and unless otherwise specified, the reagents used are conventional commercial reagents or reagents prepared according to conventional methods.

[0031] Example 1: Determination of glyphosate in coffee samples

[0032] 1. Preparation of Cu-CDs

[0033] 0.8 g of 4-aminoantipyrine and 0.7 g of CuCl2·2H2O were dissolved in 25 mL of deionized water. After ultrasonic treatment to mix and dissolve, the mixture was reacted in a microwave at 1000 W and 180 °C for 120 min. After the reaction product was cooled, it was centrifuged at 8000 r / min for 15 min. The supernatant was filtered through a 0.22 mm filter membrane and dried under vacuum at 60 °C for 12 h to obtain copper-doped carbon dots Cu-CDs.

[0034] 2. Preparation of Cu-CDs / ABEI@Ag nanozymes

[0035] 80 μL of 0.2 mg / mL Cu-CDs and 300 μL of 1 mg / mL ABEI solution were added to a mixed solution of 400 μL of 11 mg / mL AgNO3 solution and 12 mL of deionized water. The mixture was stirred at room temperature for 25 min and then dried under vacuum at 60 °C for 12 h to obtain Cu-CDs / ABEI@Ag nanozyme.

[0036] 3. Characterization of Cu-CDs / ABEI@Ag nanozymes

[0037] The morphology of Cu-CDs / ABEI@Ag nanozymes was characterized by TEM, and the results are as follows: Figure 1 Cu-CDs / ABEI@Ag nanozymes are quasi-spherical particles with good dispersibility. High-resolution TEM (HRTEM) of Cu-CDs / ABEI@Ag nanozymes clearly revealed a lattice spacing of 0.22 nm, corresponding to the (110) crystal plane of graphite carbon; the FTIR spectrum of Cu-CDs / ABEI@Ag nanozymes ( Figure 2 The display shows 3460cm -1 3042cm -1 1644cm -1 and 1382cm -1 The characteristic peaks at 603 cm⁻¹ are attributed to OH, CH, C=O, and C=C bonds, respectively.-1 The characteristic peak at this location is attributed to the stretching vibration of Cu-O; the high-resolution XPS image of Cu is shown below. Figure 3 a. The high-resolution Cu 2p spectrum shows two peaks at 953.15 eV and 933.14 eV, corresponding to Cu 2p... 1 / 2 and Cu 2p 3 / 2 The results showed that the Cu doped in Cu-CDs / ABEI@Ag mainly existed in the Cu(I) state; the high-resolution Ag 3d spectrum ( Figure 3 b) Displays 367.9 (Ag3d) 5 / 2 ) and 373.9 eV (Ag 3d 3 / 2 The two peaks can be attributed to Ag(0), therefore, it is speculated that both Cu-CDs and ABEI can reduce Ag(I) to Ag(0) and further aggregate in situ onto the Ag nucleus. Furthermore, Cu-CDs and ABEI also act as protective agents, attaching to the Ag nucleus surface via Ag-N bonds. Finally, through coordination and electrostatic interactions between Cu-CDs and Cu-O and Cu-N bonds, Cu... 2+ Further coating on the surface yields Cu-CDs / ABEI@Ag nanozymes.

[0038] 4. Evaluation of peroxidase activity of Cu-CDs / ABEI@Ag nanozymes

[0039] To verify whether Cu-CDs / ABEI@Ag nanozymes have peroxidase-like activity, three compounds, TMB, ABTS, and OPD, were selected as catalytic substrates. 100 μL each of 5 mmol / L TMB, ABTS, and OPD were added to 100 μL of 50 mmol / L H2O2, 100 μL of 2 μg / mL Cu-CDs / ABEI@Ag nanozyme, and 100 μL of 0.1 mol / L pH 4.0 NaAc-HAc buffer solution, respectively. After shaking and standing for 10 min, the absorbance of oxTMB and oxABTS was measured at wavelengths of 654 nm and 415 nm, respectively. The fluorescence intensity of DAP, the oxidation product of OPD, was measured at an excitation wavelength of 420 nm and a wavelength of 564 nm.

[0040] The results are as follows Figure 4 As shown, in the presence of H2O2, Cu-CDs / ABEI@Ag nanozymes catalyze the oxidation of colorless TMB, ABTS, and OPD into colored products (oxTMB and oxABTS) and fluorescent products (DAP), respectively. Among the three substrates, the degree of oxidation is in the order of OPD > TMB > ABTS. ABTS is hardly oxidized, which may be due to the negative charge of Cu-CDs / ABEI@Ag nanozymes, which repels the negatively charged ABTS.

[0041] Furthermore, the POD-like catalytic activity of Cu-CDs / ABEI@Ag nanozymes was investigated using steady-state kinetics tests. The Michaelis constants of Cu-CDs / ABEI@Ag nanozymes were determined with TMB and H2O2 as substrates, respectively. K m and reaction rate constant V max , usually, lower K m This implies a high affinity between the enzyme and the substrate; where the TMB concentration is 0.125 mM when H2O2 is a variable; and the H2O2 concentration is 0.25 mM when TMB is a variable.

[0042] The results are as follows Figure 5 , Figure 6 As shown, Cu-CDs / ABEI@Ag nanozymes exhibit resistance to H2O2 and TMB. K m The values ​​were 3.248 and 0.882 mM, respectively. V max The values ​​are 4.297 × 10 -8 Ms -1 and 1.108×10 -8 Ms -1 Compared to horseradish peroxidase (HRP), Cu-CDs / ABEI@Ag nanozymes have comparable performance. K m and V max This indicates that Cu-CDs / ABEI@Ag nanozymes are POD-ike nanozymes with good binding affinity to substrates (Table 1).

[0043] Table 1 Comparison of Michaelis constants

[0044] .

[0045] 5. Chemiluminescent properties of Cu-CDs / ABEI@Ag nanozymes

[0046] The chemiluminescence performance of the prepared Cu-CDs / ABEI@Ag nanozyme was investigated using a chemiluminescence-enabled microplate reader. In a 98-well microplate, 50 μL of 10 mmol / L ABEI solution, 20 μL of 50 mmol / L H2O2, 10 μL of 2 mg / mL Cu-CDs / ABEI@Ag nanozyme, 50 μL of 100 μg / L glyphosate solution, and 20 μL of 0.1 mol / L pH 11 Na2CO3-NaHCO3 buffer solution were added and mixed. The CL intensity of the reaction system was measured at 420 nm. A reaction system without H2O2 and glyphosate was also included as a control.

[0047] The results are as follows Figure 7 As shown, the maximum emission wavelength of this reaction system is 420 nm, consistent with the previously reported CL spectrum of ABEI, indicating that the luminescent agent is N-(aminobutyl)-N-(ethylphthalimide) (ox-ABEI). The results show that the Cu-CDs / ABEI@Ag nanozyme can catalyze the luminescence of ABEI regardless of the presence or absence of H2O2, but the reaction system exhibits a stronger CL peak in the presence of H2O2. However, the chemiluminescence intensity of the ABEI / H2O2-Cu-CDs / ABEI@Ag system significantly decreased after the addition of PMG, demonstrating that PMG suppressed the chemiluminescence intensity of the system.

[0048] 6. Chemiluminescent detection mechanism of glyphosate by Cu-CDs / ABEI@Ag nanozymes

[0049] To achieve detection, the mechanism by which glyphosate inhibits the catalytic activity of Cu-CDs / ABEI@Ag nanozymes was investigated, and the interaction between Cu-CDs / ABEI@Ag nanozymes and glyphosate was studied by FTIR spectroscopy. Figure 8 The figures show the FTIR spectra of Cu-CDs / ABEI@Ag and Cu-CDs / ABEI@Ag+PMG, with the values ​​displayed in the 847–1033 cm⁻¹ range. -1 In the spectral band, the signal intensity increases with the addition of PMG, particularly at 918 cm⁻¹ in the FTIR spectrum of Cu-CDs / ABEI@Ag⁺ PMG. -1 and 949cm -1 The band represents the stretching vibration of the CCNC bond, 763 cm⁻¹. -1 and 847cm -1The enhanced Cu-O stretching vibration peak at the chromatogram indicates an interaction between PMG and Cu-CDs. However, after washing the Cu-CDs / ABEI@Ag+PMG sample with ultrapure water, the CCNC stretching vibration in the FTIR of the washed Cu-CDs / ABEI@Ag+PMG was weakened, indicating that PMG had partially disappeared from the Cu-CDs / ABEI@Ag surface. Furthermore, previous studies have verified the ability of PMG to chelate with Cu(II) via phosphate, amino, and carboxyl groups. Figure 9 The UV-Vis absorption spectra of Cu-CDs / ABEI@Ag, PMG, and Cu-CDs / ABEI@Ag+PMG systems are presented. Compared with the PMG system alone, the Cu-CDs / ABEI@Ag+PMG system shows an enhanced absorption peak at 267 nm, indicating an interaction between Cu-CDs / ABEI@Ag and PMG.

[0050] Subsequently, free radical scavenging experiments were conducted to determine the types of reactive oxygen species (ROS). Three scavengers, namely isopropanol (IPA), tempol, and tryptophan, were used to capture and eliminate ·OH and ·O2, respectively. - and 1 O2, the reaction system is the same as in step 5; the results are as follows. Figure 10 and Figure 11 As shown, compared with the control group, the addition of the three free radical scavengers led to varying degrees of decrease in the CL intensity of the system, regardless of the presence or absence of PMG. This indicates that the three ROS are the main active substances in the reaction system. However, when PMG is present, the capture efficiency of IPA decreases from 99.8% to 98.6%, indicating that PMG can reduce the content of ∙OH. Combined with the changes in UV-vis spectra, it is speculated that the inhibition mechanism of PMG on the system is due to PMG occupying the catalytic active sites of Cu-CDs / ABEI@Ag, interfering with the interaction between H2O2 and Cu-CDs / ABEI@Ag, thereby reducing its free radicals, especially hydroxyl radicals.

[0051] 7. Preparation of PMG chemiluminescence intensity working curve

[0052] Add 20 μL of 2 mg / mL Cu-CDs / ABEI@Ag nanozyme solution, 50 μL of 10 mmol / L ABEI solution, 20 μL of 50 mmol / L H2O2 solution, and 50 μL of PMG solution (concentration range: 6.67-333 μg / L) to a 98-well ELISA plate. Add 100 μL of 0.1 mol / L pH 11 Na2CO3-NaHCO3 buffer solution, mix well, and use an ELISA reader to measure the chemiluminescence intensity and calculate Ig. CL Plotting PMG concentration on the x-axis, I CLUsing the vertical axis as the ordinate, a standard curve was plotted, and the regression equation, correlation coefficient, relative standard deviation, linear range, etc., were obtained (see Table 2). The results are shown below. Figure 12 ;

[0053] Table 2. Linear equation, correlation coefficient, relative standard deviation, and linear range

[0054] ;

[0055] 8. Method specificity investigation

[0056] The specificity and anti-interference ability of CL sensors are key factors affecting sensor performance. Since PMG is commonly used for weed control in coffee or crops, common metal ions (K+) are selected. + Mg 2+ Mn 2+ Zn 2+ The specificity and anti-interference ability of the reaction system of this invention were evaluated by using the main components of coffee (caffeine, citric acid, casein, tannic acid, caffeic acid, sucrose, glucose) and organophosphorus pesticides (methyl parathion, glyphosate, parathion, profenofos, paraoxon, glufosinate). The reaction system was the same as in step 7, except that the interfering substances added to the reaction system were: citric acid and tannic acid at 13 mg / L, caffeic acid at 146.7 mg / L, sucrose and glucose at 5 mg / L, glyphosate at 80 mg / L, glycine at 45 mg / L, and other substances at 500 mg / L. The results are as follows: Figure 13 As shown, no significant changes in CL were observed in the interfering substances compared to the blank, indicating that Cu-CDs / ABEI@Ag nanozyme as a CL sensor has good specificity for the quantitative determination of PMG.

[0057] 9. Determination of PMG in coffee samples

[0058] (1) Interference of sample matrix on CL sensor detection

[0059] To explore the feasibility of establishing a CL probe for application in actual coffee samples, four coffee samples were directly dissolved in water without pretreatment, and the luminescence of the system in the four coffee extracts was examined. Results are shown below. Figure 14 The system emitted light normally in all four types of coffee, providing support for the practical application of the probe.

[0060] (2) Coffee sample processing: Weigh 1.00g of sample powder into an Erlenmeyer flask, add 20 mL of 1mol / L NaOH deionized water solution, extract by sonication for 20 min, then centrifuge at 8000 rpm for 5 min, take 3 mL of supernatant and mix with 0.4g ZnSO4, add 300µL NaOH solution (1mol / L), vortex mix for 60 s, centrifuge at 6000 rpm for 5 min, collect the supernatant, filter through a 0.22 µm aqueous phase filter membrane, collect the filtrate and store at 4 ℃ for later use, to obtain the sample extract.

[0061] (3) Coffee sample determination

[0062] Prepare the reaction system according to the method in step 7, except that 100 μL of sample extract solution is used instead of the PMG standard solution. Measure the chemiluminescence intensity and calculate Ig. CL Substituting these values ​​into the regression equation of step 7, the results showed that PMG was not detected in any of the sample extracts.

[0063] (4) Recovery and precision experiments: Three different concentrations of PMG standard solution were added to the coffee sample; each concentration was measured in parallel three times, the spiked recovery rate was calculated, and the relative standard deviation (RSD) was calculated. The results are shown in Table 3. The spiked recovery rate of PMG was 95.86% to 111.13%, and the RSD was 1.07% to 3.76%. This method has good accuracy and precision.

[0064] Table 3. Detection of glyphosate in coffee samples (n=3)

[0065]

[0066] The glufosinate determination method established in this invention has the advantages of fewer processing steps, speed and simplicity, no need for large-scale instruments and equipment and professional operators required for the liquid chromatography-mass spectrometry method in GB 23200.108-2018, short processing time, low processing cost and simple operation, and has strong advantages in actual detection.

Claims

1. A method for the chemiluminescent detection of glyphosate in coffee catalyzed by Cu-CDs / ABEI@Ag nanozymes, characterized in that, The steps are as follows: (1) Dissolve 0.70-1.0g of 4-aminoantipyrine and 0.6-0.8g of CuCl2·2H2O in deionized water, mix and dissolve by ultrasonication, and react the mixture in microwave at 180℃ for 100-150min. After cooling the reaction product, centrifuge, filter the supernatant through a 0.22mm filter membrane, and dry under vacuum to obtain copper-doped carbon dots Cu-CDs; (2) Mix 50-100 μL of 0.2 mg / mL copper-doped carbon dot Cu-CDs solution, 200-400 μL of 1 mg / mL N-(4-aminobutyl)-N-ethyl isoluminol solution, 300-500 μL of 11 mg / mL AgNO3 solution, and 10-15 mL of deionized water. Stir at room temperature for 20-30 min and then vacuum dry to obtain Cu-CDs / ABEI@Ag nanozyme. (3) Mix Cu-CDs / ABEI@Ag nanozyme solution, N-(4-aminobutyl)-N-ethyl isoluminol solution, H2O2 solution and glyphosate solutions of different concentrations, add pH 11 Na2CO3-NaHCO3 buffer solution, mix well, and use an enzyme-linked immunosorbent assay (ELISA) reader to measure the chemiluminescence intensity and calculate ΔI. CL Determine the relationship between glyphosate concentration and ΔI CL The linear relationship is obtained to derive the regression equation, where ΔI CL = I-I0, where I and I0 are the chemiluminescence intensities of the reaction systems without and with glyphosate, respectively; (4) Measure the chemiluminescence intensity of the sample to be tested according to the method in step (3), and calculate ΔI. CL Substitute the glyphosate concentration into the regression equation to obtain the concentration of glyphosate in the sample to be tested.

2. The method for detecting glyphosate in coffee using Cu-CDs / ABEI@Ag nanozyme catalysis and chemiluminescence as described in claim 1, characterized in that: The concentration of Cu-CDs / ABEI@Ag nanozyme solution is 2 mg / mL, and the addition amount is 10-20 μL; the concentration of N-(4-aminobutyl)-N-ethyl isoluminol solution is 10 mmol / L, and the addition amount is 40-60 μL; the concentration of H2O2 solution is 50 mmol / L, and the addition amount is 20-40 μL; the concentration of pH 11 Na2CO3-NaHCO3 buffer solution is 0.1 mol / L, and the addition amount is 50-100 μL.

3. The method for detecting glyphosate in coffee using Cu-CDs / ABEI@Ag nanozyme catalysis and chemiluminescence as described in claim 1, characterized in that: The microwave power is 1.0-1.2kW, and the centrifugation is performed at 8000-10000r / min for 5-15min.

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