Rapid detection method for fluazinam based on L-cys-Cu NCs fluorescence sensing system

Abstract

The invention discloses a method for rapidly detecting fluazinam on the basis of an L-cys-Cu NCs fluorescent sensing system. The L-cys-Cu NCs fluorescent nano material is successfully obtained by taking L-cysteine and copper sulfate as raw materials and adopting a simple and rapid one-step synthesis method. The L-cys-Cu NCs synthesized by the invention realizes trace detection of fluazinam under the optimal condition, and when the concentration of fluazinam is between 0.05 mu M and 25 mu M, the detection result has a good linear relationship, and the detection limit reaches 1.4 nM. Meanwhile, the fluorescent sensing system is successfully used for detecting fluazinam in a real sample, and experimental results show that the fluorescent system has good potential application value in fluazinam monitoring.

Description

Technical Field

[0001] This invention belongs to the field of detection technology, specifically relating to a rapid detection method for fluazinam based on an L-cys-Cu NCs fluorescence sensing system. Background Technology

[0002] Fluazinam is a benzylpyridinium pesticide widely used in agriculture to control late blight. However, fluazinam is difficult to remove and has a long-lasting effect on crops. Fluazinam residues can cause varying degrees of dermatitis, asthma, and other diseases in humans, seriously impacting human health. Furthermore, excessive fluazinam may directly affect the expression of tyrosine hydroxylase. Therefore, monitoring the concentration of fluazinam in the environment is of paramount importance. Existing technologies disclose many analytical methods such as gas chromatography (GC), high-performance liquid chromatography (HPLC), gas chromatography-mass spectrometry (GC-MS), and liquid chromatography-tandem mass spectrometry (LC-MS) for analyzing and monitoring fluazinam. These methods offer sensitive and effective measurements, but also suffer from drawbacks such as expensive equipment, complex operation, and high levels of expertise required. These limitations restrict their further application. Therefore, developing a simple, rapid, and sensitive method for detecting fluazinam is of great significance.

[0003] Existing technologies disclose several methods for the fluorescence detection of fluazinam. For example, a FRET-based fluorescence sensing platform is used to detect fluazinam with a detection limit of 2.3 nM; alternatively, existing technologies disclose the synthesis of N,S-doped carbon quantum dots (N,S-CQDs) for the quantitative analysis of fluazinam, achieving a detection limit as low as 10 nM through the internal filtration effect (IFE). While these methods achieve satisfactory sensitivity in detecting fluazinam, the FRET-based fluorescence sensing platform uses cadmium telluride quantum dots (CdTe) with low toxicity, which can cause secondary pollution, and the synthesis process of N,S-CQDs requires high temperatures and is time-consuming. These drawbacks significantly reduce their practicality. Summary of the Invention

[0004] In view of the above-mentioned shortcomings of the existing technology, the purpose of this invention is to provide a rapid detection method for fluazinam based on the L-cys-Cu NCs fluorescence sensing system, so as to solve the problems of the existing fluorescent detection methods for fluazinam, which have the problems of the detection substance being toxic and causing secondary pollution, and the detection substance requiring high temperature and long time to synthesize.

[0005] Problems include low electrode integration and inability to be reused.

[0006] To solve the above-mentioned technical problems, the present invention adopts the following technical solution:

[0007] A rapid detection method for fluazinam based on an L-cys-Cu NCs fluorescence sensing system, characterized by the following specific steps:

[0008] Step 1: Dilute L-cys-Cu NCs copper nanoclusters with HEPES buffer solution to obtain a mixed solution; add fluazinam standard solutions of different concentrations to the mixed solution, mix thoroughly, and react at room temperature for 50-100 seconds to obtain the test solution; wherein, the volume ratio of L-cys-Cu NCs copper nanoclusters to HEPES buffer solution is 1:(65-75); the concentration of HEPES buffer solution is 0.1 mol / L; the pH value of the test solution is not lower than 10;

[0009] Step 2: Under the conditions of excitation wavelength of 365 nm and emission wavelength of 497 nm, the fluorescence intensity of the test solution after the reaction corresponding to different concentrations of fluazinam standard solution was measured; at the same time, a blank control group was set up without the addition of fluazinam and replaced with an equal volume of HEPES buffer solution.

[0010] Step 3: Calculate the relative fluorescence intensity, plot a standard curve with fluazinam concentration on the x-axis and relative fluorescence intensity on the y-axis, and obtain the linear regression equation;

[0011] Step 4: Process the sample to be tested to obtain the sample extract, perform fluorescence detection as in Step 2, calculate the relative fluorescence intensity of the sample to be tested, and compare it with the standard curve to obtain the concentration of fluazinam in the sample to be tested.

[0012] Preferably, L-cys-Cu NCs copper nanoclusters are obtained through the following steps:

[0013] Anhydrous copper sulfate and L-cysteine ​​were dissolved in deionized water and stirred continuously before adding NaOH aqueous solution. Stirring was continued for 1 to 3 hours at room temperature. Finally, the supernatant was extracted by centrifugation and stored at 4°C for later use.

[0014] The concentration ratio of anhydrous copper sulfate to L-cysteine ​​is (1~2):(1~2); the concentration of NaOH aqueous solution is 1~3 mol / mL.

[0015] Preferably, in step 2, the concentration of the fluazinam standard solution is 0.05 μmol / L to 25 μmol / L.

[0016] Preferably, the detection method has a detection limit of ≥1.4 nmol / L for fluazinam.

[0017] Preferably, the rapid detection method is used for the detection of fluazinam in vegetables and fruits.

[0018] Compared with the prior art, the present invention has the following beneficial effects:

[0019] 1. This invention employs a strategy of modifying copper nanoclusters with L-cysteine, achieving a perfect unity between material preparation and functionalization. This one-step synthesis method is not only simple to operate and has mild reaction conditions, but also successfully constructs fluorescent nanomaterials with excellent optical properties. At the molecular structure level, L-cysteine ​​is firmly modified on the surface of copper nanoclusters through sulfur-copper covalent bonds, forming a stable three-dimensional structure and endowing the material with excellent water solubility and biocompatibility. This biomolecular modification strategy cleverly avoids the toxicity problem of traditional heavy metal nanomaterials, fundamentally solving the risk of secondary pollution during the detection process; at the same time, the negative charge characteristics and rich functional group distribution on the material surface not only enhance its stability in complex media, but also lay the foundation for subsequent specific interactions with target molecules.

[0020] 2. In the application of copper nanoclusters to detect fluazinam, this invention unexpectedly discovered that although copper nanoclusters and fluazinam molecules carry the same type of charge on their surfaces, theoretically they should be unable to interact effectively due to electrostatic repulsion. However, this system can achieve highly sensitive recognition of the target molecule through a unique internal filtration effect mechanism. In practical applications, the detection method exhibits excellent selectivity, accurately identifying the target molecule even in complex environments where multiple pesticides and ions coexist, unaffected by interference from other substances. Simultaneously, it demonstrates excellent detection accuracy and stability in agricultural product sample testing, with highly reliable recovery rates. Compared to traditional expensive instrumental analysis methods, the technical operation process of the detection method described in this invention is significantly simplified, the detection time is greatly reduced, and it can be mastered without professional training, providing a practical tool that is easily promoted and popularized for food safety monitoring and environmental health assessment. Attached Figure Description

[0021] Figure 1 This is a schematic diagram illustrating the synthesis of L-cys-Cu NCs and the fluorescence detection principle of fluazinam.

[0022] Figure 2 The images show the TEM and size distribution of L-cys-Cu NCs; A is the TEM image, and B is the size distribution image.

[0023] Figure 3 The Fourier transform infrared spectrum of L-cys-Cu NCs is shown.

[0024] Figure 4 A is the X-ray photoelectron spectrum of L-cys-Cu NCs; A is the full-scan XPS spectrum, and B is the high-resolution XPS spectrum of Cu 2p.

[0025] Figure 5The diagram shows the optical properties of L-cys-Cu NCs; in the inset, photographs of L-cys-Cu NCs obtained under visible light (1) and ultraviolet light (2) are shown.

[0026] Figure 6 In A, a is the fluorescence excitation spectrum of L-cys-Cu NCs, b is the fluorescence emission spectrum of L-cys-Cu NCs, and c is the UV-Vis absorption spectrum of fluazinam; B is the fluorescence decay curve of L-cys-Cu NCs before and after the addition of fluazinam; C is the Zeta potential of L-cys-Cu NCs.

[0027] Figure 7 In the middle: A is the fluorescence emission spectrum of L-cys-Cu NCs after adding different concentrations of fluazinam; B is the fitted scatter plot of the fluorescence intensity change (∆F) of L-cys-CuNCs with the concentration of fluazinam, and the inset is the linear relationship graph.

[0028] Figure 8 In the diagram: A represents the effect of pH value on the change in fluorescence intensity (∆F); B represents the effect of response time on the change in fluorescence intensity (∆F).

[0029] Figure 9 The change in fluorescence emission intensity (ΔF) of L-cys-Cu NCs caused by fluazinam and other pesticides and cations.

[0030] Figure 10 In the figure: A represents the stability of the L-cys-Cu NCs sensing system in the detection of fluazinam, and B represents the reproducibility of the L-cys-Cu NCs sensing system in the detection of fluazinam. Detailed Implementation

[0031] The present invention will be further described below with reference to the accompanying drawings and embodiments. It should be understood that the following embodiments are for illustrative purposes only and are not intended to limit the scope of the invention.

[0032] Example 1: Preparation and characterization of L-cys-Cu NCs copper nanoclusters (i.e., L-cys-Cu NCs)

[0033] (1) Preparation

[0034] 5.0 mM anhydrous copper sulfate and 5.0 mM L-cysteine ​​were dissolved in 20 mL of deionized water and stirred vigorously for 5 min. Then, 0.5 mL of 1 M NaOH aqueous solution was added, and stirring was continued for 2 h at room temperature. Finally, the supernatant was collected after centrifugation at 10000 rpm for 20 min and stored in a refrigerator at 4 °C to obtain the L-cys-Cu NCs nanoclusters.

[0035] The size of the prepared L-cys-Cu NCs was obtained under transmission electron microscopy (TEM), surface state data of L-cys-Cu NCs were obtained using X-ray photoelectron spectroscopy (XPS), chemical functional groups of L-cys-Cu NCs were analyzed using Fourier transform infrared spectroscopy (FT-IR), data of optimal excitation and emission wavelengths were acquired using a fluorescence spectrophotometer, and ultraviolet absorption spectra were obtained by scanning with UV-vis. Fluorescence lifetime tester and Zeta potential tester can measure its fluorescence lifetime data and surface charge.

[0036] (2) Characterization

[0037] ① TEM characterization of L-cys-Cu NCs

[0038] The morphology and size of the synthesized L-cys-Cu NCs were confirmed from typical TEM images ( Figure 2 The results indicate that L-cys-CuNCs have good dispersibility (uniform distribution). The size distribution image of L-cys-CuNCs shows that their average size is approximately 2.4 nm.

[0039] ②FT-IR characterization of L-cys-Cu NCs

[0040] Surface group information of L-cys-Cu NCs was obtained from FT-IR spectroscopy. Figure 3 The presence of a distinct absorption band at 3441 cm⁻¹, attributed to the stretching vibration of -NH₂, indicates the abundance of amino groups on the surface of L-cys-Cu NCs. The two absorption peaks at 1631 cm⁻¹ and 1393 cm⁻¹ are likely due to the stretching vibration of the COO- group. Compared to the FT-IR of free L-cysteine, the -SH band at 2551 cm⁻¹ almost completely disappears in L-cys-Cu NCs, indicating the formation of a Cu-S covalent bond between L-cysteine ​​and copper sulfate, thus demonstrating the successful synthesis of L-cys-Cu NCs.

[0041] ③ X-ray photoelectron spectroscopy (XPS) characterization of L-cys-Cu NCs

[0042] The chemical composition of L-cys-Cu NCs was further identified using X-ray photoelectron spectroscopy (XPS). Figure 4 As can be seen, the XPS full spectrum of L-cys-Cu NCs shows six characteristic peaks: C 1s, N 1s, O 1s, S 2p, Cu 2p, and Na 1s. (For example...) Figure 4As shown in (B), the characteristic peaks of Cu at 951.3 eV and 931.7 eV are Cu 2p1 / 2 and Cu 2p3 / 2, respectively. Furthermore, there is no characteristic peak near 942 eV, indicating that the peaks of L-cys-Cu NCs do not contain Cu(II). All these experimental data analyses demonstrate that L-cysteine ​​was successfully modified onto Cu NCs.

[0043] ④ Characterization of optical properties of L-cys-Cu NCs

[0044] The optical, morphological, and chemical properties of the synthesized L-cys-Cu NCs fluorescent nanoparticles were investigated using a fluorescence spectrophotometer, transmission electron microscope, and UV-Vis microscope. The optical properties of L-cys-CuNCs were studied using a fluorescence spectrophotometer. Figure 5 Figures show the fluorescence excitation (b), fluorescence emission (c), and UV-Vis absorption (a) spectra of L-cys-Cu NCs, respectively. The insets show photographs of L-cys-Cu NCs obtained under visible light (1) and ultraviolet light (2). The optimal emission wavelength of L-cys-Cu NCs under 365 nm excitation is 497 nm. Furthermore, aqueous solutions of L-cys-Cu NCs exhibit bright green fluorescence under 365 nm ultraviolet light irradiation.

[0045] This paper explores the analysis and detection of fluazinam using the L-cys-Cu NCs fluorescence detection method constructed in this experiment and further elucidates its fluorescence quenching mechanism. Firstly, the optical properties of L-cys-Cu NCs and fluazinam were investigated. (The text then abruptly shifts to a different topic:) Figure 6 It can be seen that fluazinam has a relatively obvious UV-Vis absorption peak at 352 nm, while the excitation wavelength of L-cys-Cu NCs happens to be at 365 nm. The results indicate that the UV-Vis absorption band of fluazinam significantly overlaps with the fluorescence excitation spectrum of L-cys-Cu NCs, causing the fluorescence intensity of some L-cys-Cu NCs to be absorbed and quenched by fluazinam. This provides a prerequisite for the IFE principle. This invention further investigated the change in fluorescence lifetime of L-cys-Cu NCs before and after the addition of fluazinam. Figure 6As shown in Figure B, the fluorescence lifetime of L-cys-Cu NCs was 3.45 ns before the addition of fluazinam, and 2.97 ns after the addition, showing no significant change. These conclusions can be used to distinguish between IFE and FRET, where the absorber competitively absorbs the excitation and emission light of the fluorophore without affecting its luminescence properties. Therefore, since the fluorescence lifetime of the fluorophore remains unchanged before and after the introduction of the absorber, it can be inferred that IFE has occurred. Furthermore, the Zeta potential of L-cys-Cu NCs was investigated, as shown... Figure 6 As shown in (C), its surface charge is -17.4 mV, which repels the similarly negatively charged fluazinam. In summary, L-cys-Cu NCs and fluazinam carry the same surface charge, and their mutual repulsion results in a large distance between them, with no charge transfer. Therefore, it can be inferred that the principle of the fluorescence method used in this invention to detect fluazinam is based on the quenching of the fluorescence intensity of L-cys-Cu NCs caused by IFE. Figure 1 The reaction mechanism of IFE is explained in detail.

[0046] Example 2: Determination of Fluazinam Standard Solution

[0047] 40 µL of L-cys-Cu NCs was diluted in 2.93 mL of HEPES (0.1 M) buffer solution and then added to a 5 mL centrifuge tube and mixed thoroughly. 30 µL of standard solutions of fluazinam at different concentrations were added to the mixture and shaken thoroughly. After the reaction was complete at room temperature for 60 s, the fluorescence intensity of L-cys-Cu NCs at 497 nm was measured using a fluorescence spectrophotometer with an excitation wavelength of 365 nm.

[0048] When a series of different concentrations of fluazinam were added to an aqueous solution of L-cys-Cu NCs, the fluorescence intensity of L-cys-Cu NCs gradually decreased as the concentration of fluazinam increased. Figure 7 The results show that a linear calibration curve (F0-F1) can be established when the concentration of fluazinam is in the range of 0.05 μM to 25 μM, exhibiting a reliable linear relationship. The calibration equation can be expressed as F0-F1 = 14.00 c + 62.07 (c: μM), with a correlation coefficient R = 0.9953. Based on the principle of S / N = 3 (S is the standard deviation of 11 blank samples, and N is the slope of the calibration line), the lowest detection limit of the constructed fluazinam detection system is calculated to be 1.4 nM. Compared with other fluorescence sensing platforms for fluazinam detection in Table 1, the L-cys-Cu NCs synthesized in this invention have a lower detection limit for fluazinam.

[0049] Table 1

[0050]

[0051] Example 3: Optimization of the detection method

[0052] Based on Example 2, adjustments were made, the difference being that the concentration of the fluazinam standard solution was 0.1 μM, the reaction time and pH value were adjusted, and the effects of different reaction times and pH values ​​on the detection effect were studied.

[0053] like Figure 8 As shown in Figure A, this invention investigated the effect of different pH values ​​of the HEPES buffer solution on the experiment. ΔF (ΔF represents the difference in fluorescence change with and without fluazinam) gradually increased with pH from 5 to 10, and remained almost constant above pH 10. This may be because the carboxyl and amine groups on the surface of L-cys-Cu NCs tend to protonate in acidic environments, thus reducing fluorescence intensity at acidic pH, while increasing fluorescence intensity under alkaline conditions due to the increased deprotonation concentration. Therefore, pH=10 is the optimal choice for determining fluazinam. Furthermore, this invention also considered the effect of reaction time on the experiment. Figure 8 As shown in Figure B, ΔF (ΔF = F0 - F1) describes the fluorescence change of L-cys-Cu NCs aqueous solution in the presence and absence of fluazinam. The ΔF value gradually increases with increasing reaction time. When the reaction time is 60 s, the reaction system basically reaches a relative equilibrium state, and ΔF reaches its maximum value. Therefore, a reaction time of 60 s between L-cys-Cu NCs and fluazinam is optimal, and no further reaction time is required.

[0054] Example 4: Selectivity of the detection method

[0055] Add 40 µL of L-cys-Cu NCs to 2.93 mL of HEPES (0.1 M) buffer solution and dilute to an appropriate concentration. Take 2.97 mL of the buffer-diluted L-cys-Cu NCs into a 5 mL centrifuge tube. Then add 30 µL of standard solutions of fluazinam at different concentrations and other anti-interference substances, including common cations: K, Na, NH4, Ca, Al, Zn, Mg, and some other pesticides, such as: cyanazine, glyphosate, acetamiprid, methamidophos, triadimefon, trichlorfon, dimethoate, and fenitrothion, to the above solution and shake thoroughly. After the reaction is complete at room temperature for 60 s, record the fluorescence intensity change of L-cys-Cu NCs at an emission wavelength of 497 nm.

[0056] To investigate the selectivity of L-cys-Cu NCs for the detection of fluazinam, this invention analyzed the influence of several potential interfering substances on the experiment, including positively charged ions such as K, Na, NH4, Ca, Al, Zn, and Mg, and other pesticides such as cyanazine, glyphosate, acetamiprid, methamidophos, triadimefon, trichlorfon, dimethoate, and fenitrothion. The final concentration of fluazinam was 0.1 μM, while the concentrations of other pesticides and cations were 1 μM and 2 μM, respectively. Figure 9 It can be seen that, under optimal conditions, these interfering substances, with concentrations 10 or 20 times higher than fluazinam, did not produce significant fluorescence changes in L-cys-Cu NCs in the reaction system. Only after the addition of fluazinam did the fluorescence intensity of L-cys-Cu NCs undergo significant quenching. Therefore, the L-cys-Cu NCs fluorescence method proposed in this invention has good anti-interference ability for the detection of fluazinam and can be used for the quantitative detection of fluazinam.

[0057] Example 5: Detection of actual samples

[0058] This study used Chinese cabbage and pear as real samples to verify the selectivity and feasibility of the L-cys-Cu NCs fluorescence method for the analysis and detection of fluazinam. Both the Chinese cabbage and pear were purchased from a local supermarket in Chongqing. Approximately 2.5 g of each vegetable and pear were chopped and dissolved in 10 mL of ethanol with a certain concentration of fluazinam. After sonication for 20 min, the liquid was centrifuged at 8000 rpm for 10 min to remove insoluble substances. The supernatant was collected and spiked with L-cys-Cu NCs to detect fluazinam.

[0059] To evaluate the feasibility and practicality of the L-cys-Cu NCs fluorescence method for the determination of fluazinam, pears and cabbage were used as actual samples for spiked recovery experiments. Following the experimental procedures in Example 2, a series of specific concentrations (1, 5, 10 ppm) of fluazinam were added to pears and cabbage, and the analysis was performed using the same method as before. As shown in Table 2, the recoveries ranged from 98.92% to 104.63%, and the relative standard deviations (RSDs) for each sample were less than 5%. These experimental results demonstrate that the L-cys-Cu NCs fluorescence analysis method established in this invention has good practicality for the analysis and detection of fluazinam in actual samples.

[0060] Table 2. Determination of fluazinam-spiked samples by the L-cys-Cu NCs sensing system

[0061]

[0062] Example 6: Stability and reproducibility of L-cys-Cu NCs sensing system for fluazinam detection

[0063] To assess the stability and reproducibility of this fluorescence sensing system, this invention evaluates its stability for fluazinam detection by long-term monitoring of the fluorescence intensity of L-cys-Cu NCs and assesses its reproducibility by studying the differences in fluorescence intensity changes between different batches and between different parallel samples within the same batch. The L-cys-Cu NCs were placed in a 4 ℃ refrigerator, and their fluorescence intensity was measured daily. The results are as follows: Figure 10 As shown in Figure A, the fluorescence intensity of L-cys-Cu NCs began to change significantly after 7 days of storage, decreasing by 32.38%. This indicates that the fluorescence sensing system constructed in this invention has only moderate stability, and its stability needs further improvement. On the other hand, the reproducibility results of this sensing system for the detection of fluazinam are as follows... Figure 10 As shown in Figure B, the maximum relative standard deviations (RSDs) for the detection of fluazinam by the L-cys-Cu NCs sensing system in ten different synthetic batches and five parallel experiments of the same synthetic batch were 2.80% and 7.76%, respectively. These experimental results indicate that the L-cys-Cu NCs fluorescence sensing system exhibits good reproducibility in the detection of fluazinam and possesses potential application value.

[0064] Finally, it should be noted that the above embodiments are only used to illustrate the technical solutions of the present invention and not to limit the technical solutions. Those skilled in the art should understand that any modifications or equivalent substitutions to the technical solutions of the present invention without departing from the spirit and scope of the present invention should be covered within the scope of the claims of the present invention.

Claims

1. A rapid detection method for fluazinam based on an L-cys-Cu NCs fluorescence sensing system, characterized in that, The specific steps are as follows: Step 1: Dilute L-cys-Cu NCs copper nanoclusters with HEPES buffer solution to obtain a mixed solution; add fluazinam standard solutions of different concentrations to the mixed solution, mix thoroughly, and react at room temperature for 50-100 seconds to obtain the test solution; wherein, the volume ratio of L-cys-Cu NCs copper nanoclusters to HEPES buffer solution is 1:(65-75); the concentration of HEPES buffer solution is 0.1 mol / L; the pH value of the test solution is not lower than 10; Step 2: Under the conditions of excitation wavelength of 365 nm and emission wavelength of 497 nm, the fluorescence intensity of the test solution after the reaction corresponding to different concentrations of fluazinam standard solution was measured; at the same time, a blank control group was set up without the addition of fluazinam and replaced with an equal volume of HEPES buffer solution. Step 3: Calculate the relative fluorescence intensity, plot a standard curve with fluazinam concentration on the x-axis and relative fluorescence intensity on the y-axis, and obtain the linear regression equation; Step 4: Process the sample to be tested to obtain the sample extract, perform fluorescence detection as in Step 2, calculate the relative fluorescence intensity of the sample to be tested, and compare it with the standard curve to obtain the concentration of fluazinam in the sample to be tested.

2. The rapid detection method according to claim 1, characterized in that, L-cys-Cu NCs copper nanoclusters were obtained through the following steps: Anhydrous copper sulfate and L-cysteine ​​were dissolved in deionized water and stirred continuously before adding NaOH aqueous solution. Stirring was continued for 1 to 3 hours at room temperature. Finally, the supernatant was extracted by centrifugation and stored at 4°C for later use. The concentration ratio of anhydrous copper sulfate to L-cysteine ​​is (1~2):(1~2); the concentration of NaOH aqueous solution is 1~3 mol / mL.

3. The rapid detection method according to claim 1, characterized in that, In step 2, the concentration of the fluazinam standard solution is 0.05 μmol / L to 25 μmol / L.

4. The rapid detection method according to claim 1, characterized in that, The detection limit of the method for fluazinam is ≥1.4 nmol / L.

5. The rapid detection method according to claim 1, characterized in that, The rapid detection method is used to detect fluazinam in vegetables and fruits.

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