Fluorescent probe for detecting thiophanate-methyl in tobacco and application of fluorescent probe

Abstract

The invention discloses a fluorescent probe for detecting thiophanate-methyl in tobacco and application of the fluorescent probe, the fluorescent probe is alpha-CuI-m-iah, and the fluorescent probe has the following properties: a PXRD measurement diffraction peak is not changed when the temperature is increased from 40 DEG C to 120 DEG C; the thermal stability temperature of a thermogravimetric analysis curve in a nitrogen atmosphere is 250 DEG C; after the PXRD spectrogram is soaked in DMSO and DMF for 24 hours, the PXRD spectrogram has no obvious change; the degree of crystallinity is not obviously changed in a 0.1 M HCl buffer solution, a 0.1 M NaOH buffer solution and a 0.01 M Tris-HCl buffer solution with the pH value of 3.0-12.0; when being excited at 366nm, the compound shows obvious yellow luminescence, and the maximum emission wavelength is 513nm; quenching luminescence is shown in DMSO (Dimethylsulfoxide); the AIE property is realized. The probe is a coordination polymer with new performance, and the coordination polymer has good thermal stability and chemical stability and also has AIE properties; the coordination polymer can be applied to detection of thiophanate-methyl.

Description

Technical Field

[0001] This application relates to the field of thiophanate-methyl detection technology, and in particular to a fluorescent probe for the detection of thiophanate-methyl in tobacco and its application. Background Technology

[0002] Coordination polymers (CPs) have been widely used in various fields such as optics, magnetism, catalysis, gas separation and storage, and biomarkers due to their diverse structures, high porosity, and large specific surface area. Among them, cuprous iodide-based coordination polymers are particularly attractive due to their high abundance, low cost, low toxicity, and excellent photoluminescence and optical tunability.

[0003] Due to the structural diversity, luminescent properties, and coordination modes of cuprous iodide (CuI) clusters, a large number of copper iodide complexes with diverse structures can be obtained after coordination with organic ligands. Coordination polymers (CPs) of copper iodide complexes with different topologies exhibit a variety of properties, such as stimuli-responsiveness, conductivity, catalytic activity, and magnetism. Furthermore, given the abundant and inexpensive availability of metallic copper in nature, CuI coordination polymers hold great promise for applications from both economic and environmental perspectives.

[0004] Aggregation-induced emission (AIE) refers to the phenomenon where fluorescence emission increases significantly when a fluorophore transitions from a monomeric to an aggregated state. AIE is primarily caused by restricted intramolecular motion (RIM). Therefore, substances exhibiting AIE properties can be used in numerous fields such as analytical detection, bioimaging, and light-emitting diodes. Most reported coordination polymers do not possess aggregation-induced emission (AIE) properties.

[0005] Existing coordination polymers (CPs) suffer from poor thermal and chemical stability, which hinders their widespread application in various fields, such as pesticide residue detection.

[0006] Thiophanate-methyl, also known as methyl thiophanate, is a potent systemic fungicide with low toxicity. It effectively controls diseases in various crops, including vegetables, flowers, fruit trees, and wheat, by acting as both a systemic and preventative agent against pathogens. It is primarily used to control tobacco powdery mildew, tomato leaf mold, and citrus anthracnose. Current methods for detecting thiophanate-methyl include chromatography and fluorescence reaction. However, existing chromatographic methods require significant equipment investment and complex sample pretreatment, reducing detection efficiency and making them unsuitable for rapid testing of large batches of samples. Currently, there are relatively few probes available for fluorescence reaction methods. For example, CN 202311336043.2 discloses a CuI-p-DPA for the fluorescent detection of thiophanate-methyl in tobacco and its application. The molecular formula of CuI-p-DPA is 2,7-bis(pyridin-4-)acridine cuprous iodide. This α-CuI-p-DPA has a specific reaction to thiophanate-methyl and possesses advantages such as good selectivity, high sensitivity, short response time, and strong anti-interference ability, making it suitable for detecting thiophanate-methyl residues in tobacco samples. However, this literature only discloses the structure of one thiophanate-methyl detection substance, resulting in a limited variety of existing fluorescent probes for thiophanate-methyl detection. Based on this, this application proposes a different structure for a fluorescent probe for thiophanate-methyl detection.

[0007] The information disclosed in the background section is intended only to enhance the understanding of the overall background of the invention and should not be construed as an admission or in any way implying that such information constitutes prior art known to those skilled in the art. Summary of the Invention

[0008] This application addresses the aforementioned technical problems by providing a fluorescent probe for the detection of thiophanate-methyl in tobacco and its application. It provides a fluorescent probe with another structure for detecting thiophanate-methyl in tobacco, which exhibits a wide linear range, low detection limit, short response time, and good recovery rate for thiophanate-methyl.

[0009] This application provides a fluorescent probe for the detection of thiophanate-methyl in tobacco, α-CuI-m-iah, with the following properties: PXRD diffraction peaks remain unchanged after heating from 40℃ to 120℃; the thermogravimetric analysis curve under nitrogen atmosphere shows a thermal stability temperature of 250℃; after soaking in DMSO and DMF for 24 hours, the PXRD spectrum shows no significant change; crystallinity shows no significant change in 0.1M HCl, 0.1M NaOH, and 0.01M Tris-HCl buffer solutions with pH 3.0–12.0; it exhibits obvious yellow emission upon excitation at 366nm, with a maximum emission wavelength of 513nm; it exhibits quenching luminescence in DMSO; it possesses AIE properties; its morphology in DMSO / EtOH mixed solvents with fe = 20%, 40%, 60%, and 90% gradually changes from amorphous flocculent to blocky, with the highest fluorescence intensity obtained at fe = 100%; the spectral data are: E m Greater than 470nm; Greater than 0.75%; k r / 10 7 Greater than 0.015s -1 .

[0010] The probe α-CuI-m-iah exhibits excellent stability. Upon binding with 0.1 mM thiophanate-methyl, the addition of thiophanate-methyl under 339 nm excitation resulted in a decrease in fluorescence intensity of the suspension, down by 81.69% compared to the original α-CuI-m-iah. Therefore, it can be used as a fluorescent probe for the detection of thiophanate-methyl. Since crystalline α-CuI-m-iah was not obtained, its chemical and structural formulas could not be determined. The following characterizes the substance using various properties. fe represents the volume ratio of EtOH in the mixed solvent.

[0011] Preferably, the linear detection range of α-CuI-m-iah for the fluorescence detection of thiophanate-methyl is 0.5 μM to 90 μM.

[0012] Preferably, the fluorescence detection limit of α-CuI-m-iah for thiophanate-methyl is 0.41 μM, i.e. 0.167 mg / kg; preferably, the fluorescence detection response time of α-CuI-m-iah for thiophanate-methyl is at least 10 min.

[0013] Another aspect of this application provides a method for preparing the aforementioned rapid detection fluorescent probe for thiophanate-methyl, comprising the following steps: grinding CuI and m-iah at room temperature, adding acetonitrile dropwise during the grinding process to obtain the fluorescent probe. Specifically, grinding for 5-10 minutes is sufficient to complete the preparation.

[0014] Preferably, when the molar ratio of CuI:m-iah is 1:1 to 5:1, the product obtained is α-CuI-m-iah.

[0015] Another aspect of this application provides a method for preparing the above-mentioned rapid detection fluorescent probe for methyl thiophanate, comprising the following steps: mixing and stirring m-iah solution with CuI solution until a white precipitate appears, followed by post-processing to obtain the fluorescent probe.

[0016] Preferably, when the molar ratio of CuI:m-iah is 1:1 to 5:1, the product obtained is α-CuI-m-iah.

[0017] Preferably, the post-treatment involves centrifuging to separate the precipitate, washing the precipitate multiple times with an organic solvent, and drying. Specifically, the organic solvents used for washing are ethanol and acetonitrile, and the product is washed with ethanol and acetonitrile separately. Specifically, the drying conditions are drying at 40°C under vacuum for 16 hours. Specifically, the reaction time is 4 hours, and the stirring speed is 680 rpm.

[0018] Preferably, the solvent used for the m-iah solution is ethanol; the solvent used for the CuI solution is acetonitrile.

[0019] Preferably, the reaction temperature is 20–40°C; more preferably, it is 25°C; preferably, the reaction system contains at least acetonitrile.

[0020] Another aspect of this application provides a fluorescent probe detection method for thiophanate-methyl, comprising the following steps: mixing the sample with the above-mentioned fluorescent probe solution to obtain a mixed solution, allowing it to stand for at least 10 minutes, using 339 nm as the excitation wavelength, recording and collecting the fluorescence spectrum in the range of 380–690 nm to observe whether the fluorescence intensity of the mixed solution decreases; if it decreases significantly, the sample contains thiophanate-methyl; if it does not decrease significantly, the sample does not contain thiophanate-methyl.

[0021] Preferably, the fluorescent probe is dissolved in DMF solution to obtain a fluorescent probe solution; the concentration of the fluorescent probe in the fluorescent probe solution is greater than or equal to 0.5 mg / mL.

[0022] Preferably, the sample pretreatment includes: after the sample is crushed into powder, it is added to a 4 mol / L hydrochloric acid solution, heated to 120°C under stirring, cooled to room temperature, centrifuged, and the supernatant is diluted and mixed with the fluorescent probe solution for detection.

[0023] Preferably, the sample is tobacco;

[0024] Preferably, the sample is a water sample.

[0025] This probe can be used for the detection of water samples, and the detection results show a good recovery rate of 94.51% to 109.87% in response to HPLC detection results.

[0026] The beneficial effects that this application can produce include:

[0027] 1) The fluorescent probe for the detection of thiophanate-methyl in tobacco provided in this application and its application, the probe is a new type of coordination polymer with good thermal stability and chemical stability, and also has AIE properties; the coordination polymer can be applied to the detection of thiophanate-methyl.

[0028] 2) The fluorescent probe for the detection of thiophanate-methyl in tobacco provided in this application and its application, the α-CuI-m-iah probe has a linear range of 0.5 μM to 90 μM for thiophanate-methyl, a low detection limit (0.41 μM, 0.167 mg / kg) and a response time of only 10 min. When applied to the detection of thiophanate-methyl in tobacco dust samples in DMF, a satisfactory recovery rate range (89.64% to 107.72%) was obtained. Attached Figure Description

[0029] Figure 1 A schematic diagram of the fluorescent probe structure for rapid detection of methyl thiophanate in at least one embodiment provided in this application; wherein A) is the synthesis reaction equation of α-CuI-m-iah and β-CuI-m-iah; B) is the coordination environment of Cu(I) ions; C) is the coordination mode of the ligand (m-iah); D) is the 1D chain of β-CuI-m-iah; E) is the copper iodide chain of β-CuI-m-iah, and the green arc is drawn by angle measurement, with distance (in...) (Unit: black) F) represents π…π interactions between adjacent ligands on the same chain; G) represents hydrogen bond interactions between adjacent ligands; β-CuI-m-iah is a one-dimensional structure observed from three directions: (H)a, (I, J)b, and (K)c. Atomic colors in the diagram: Cu - blue, N - dark blue, I - purple, O - red, C - gray;

[0030] Figure 2 PXRD spectra of ligand m-iah and coordination polymer β-CuI-m-iah in at least one embodiment provided in this application;

[0031] Figure 3 PXRD spectra of α-CuI-m-iah and β-CuI-m-iah in Examples 2-19 provided for this application; (A) PXRD spectra of powders synthesized with CuI and ligands in different proportions;

[0032] (B) PXRD spectra of ligands m-iah and CuI synthesized by solid-phase grinding in different proportions;

[0033] Figure 4PXRD spectra of the products obtained at different temperatures in at least one embodiment provided in this application; PXRD spectra of α-CuI-m-iah(A) and β-CuI-m-iah(B) synthesized at different temperatures; PXRD spectra of α-CuI-m-iah(C) and β-CuI-m-iah(D) synthesized in different solvents.

[0034] Figure 5 Infrared spectra of the ligand m-iah and coordination polymers α-CuI-m-iah and β-CuI-m-iah in at least one embodiment provided in this application; A) The excitation wavelength is 3500–500 cm⁻¹ -1 The excitation wavelength of B) is 2000–500 cm⁻¹ -1 ;

[0035] Figure 6 PXRD patterns and thermogravimetric curves of at least one embodiment provided in this application; wherein (A) α-CuI-m-iah and (B) β-CuI-m-iah PXRD patterns at different temperatures;

[0036] (C) shows the thermogravimetric curves of the ligand, α-CuI-m-iah, and β-CuI-m-iah;

[0037] Figure 7 PXRD spectra in at least one embodiment provided in this application; wherein (A) PXRD spectra of α-CuI-m-iah after soaking in different solvents for 24 hours; (B) PXRD spectra of β-CuI-m-iah after soaking in different solvents for 24 hours; (C) PXRD spectra of α-CuI-m-iah and (D) PXRD spectra of β-CuI-m-iah after soaking in aqueous solutions at different pH values;

[0038] Figure 8 Excitation spectra of CuI-m-iah and ligands in at least one embodiment provided in this application; wherein (A) solid UV-Vis absorption spectrum; (B) fluorescence excitation emission spectrum;

[0039] Figure 9 The experimental results of the luminescence properties of CuI-m-iah in at least one embodiment provided in this application are as follows: (A) Visible light; (C) Photograph of α-CuI-m-iah under ultraviolet light in different solvents; (B) Visible light; (D) Photograph of β-CuI-m-iah under ultraviolet light in different solvents; (E) Fluorescence emission spectra of α-CuI-m-iah (excitation wavelength Ex is 350 nm); (F) Fluorescence emission spectra of β-CuI-m-iah (excitation wavelength Ex is 330 nm) in different solvents. Figure 91-H2O; 2-DMSO; 3-MeCN; 4-DMF; 5-MeOH; 10-EtOH; 7-acetone; 8-THF; 9-EtOAc; 10-DCM in A, B, C, and D.

[0040] Figure 10 The images show CuI-m-iah coordination polymers from at least one embodiment provided in this application under natural light and 365nm ultraviolet light irradiation, containing different amounts of ethanol; wherein α-CuI-m-iah under different components of DMSO / EtOH is shown in (A) visible light and (C) ultraviolet light; β-CuI-m-iah under different components of DMSO / EtOH is shown in (B) visible light and (D) ultraviolet light, demonstrating solvent-induced aggregation characteristics; Figure 10 f for each sample in A, B, C, and D c (%) are 10, 20, 30, 40, 50, 60, 70, 80, 90, 100.

[0041] Figure 11 Fluorescence spectra of DMSO / EtOH mixed solutions with different ethanol components in at least one embodiment provided in this application; wherein (A) the emission spectrum of α-CuI-m-iah in DMSO / EtOH mixtures with different ethanol components; (B) the variation of aggregate emission intensity under different ethanol components; (C) the emission spectrum of β-CuI-m-iah in DMSO / EtOH mixtures with different ethanol components; (D) the variation of aggregate emission intensity under different ethanol components;

[0042] Figure 12 Fluorescence lifetime diagrams of CuI-m-iah in at least one embodiment provided in this application; wherein (A) fluorescence lifetimes of complexes assembled with CuI and m-iah in different proportions;

[0043] (B) UV-Vis absorption spectra of powders synthesized with CuI and ligands in different proportions; (C) Emission spectrum of CuI-m-iah; (D) Image of powders synthesized with CuI and ligands in different proportions under 365nm UV light.

[0044] Figure 13 The detection results of α-CuI-m-iah against thiophanate-methyl (TM) in at least one embodiment provided in this application, wherein (A) fluorescence emission spectra of α-CuI-m-iah solution before and after the addition of thiophanate-methyl (excitation wavelength Ex is 339 nm); (B) the detection results after the addition of thiophanate-methyl.

[0045] (10 -4M) Relationship between fluorescence intensity and time in α-CuI-m-iah (0.5 mg / mL) solution (excitation wavelength Ex: 339 nm); (C) Bar graph of fluorescence intensity after adding thiophanate-methyl or other pesticides to α-CuI-m-iah solution (excitation wavelength Ex: 339 nm); 1-Blank; 2-Thiophanate-methyl; 3-Fluoride; 4-Carbendazim; 5-Acetamiprid; 6-Triazole; 7-Methamphetamine; 8-Iprodione; 9-Triadimefon; 10-Imidacloprid; (D) Anti-interference experiment of α-CuI-m-iah detection of thiophanate-methyl 1-Blank; 2-Iprodione; 3-Triadimefon; 4-Acetamiprid; 5-Methamphetamine; 6-Triazole; 7-Na + ;8-NO3 - 9-K + ;10-Cl - ;

[0046] Figure 14 The quantitative detection potential results of α-CuI-m-iah in thiophanate-methyl in at least one embodiment provided in this application, wherein (A) fluorescence emission spectra of α-CuI-m-iah solution with different concentrations of thiophanate-methyl (0.5 μM to 90 μM) (excitation wavelength Ex is 339 nm);

[0047] (B) Linear relationship between fluorescence intensity of α-CuI-m-iah solution and concentration of thiophanate-methyl. Detailed Implementation

[0048] The present invention will now be described in further detail with reference to the accompanying drawings and embodiments, but this does not limit the present invention in any way. Any modifications or improvements made based on the teachings of the present invention shall fall within the protection scope of the present invention.

[0049] Example

[0050] Unless otherwise specified, all materials and instruments used in the following embodiments were obtained through commercial channels; and all detection methods used are existing methods unless otherwise specified.

[0051] The sources of materials, reagents, and instruments used in the following embodiments are as follows:

[0052] CuI and 3-pyridinecarboxylhydrazide were purchased from Anaiji Chemical Reagent Co., Ltd. Dimethyl sulfoxide (DMSO), acetonitrile (MeCN), N,N-dimethylformamide (DMF), methanol (MeOH), ethanol (EtOH), acetone (Acetone), tetrahydrofuran (THF), ethyl acetate (EtOAc), and dichloromethane (DCM) were purchased from Chengdu Kelong Chemical Co., Ltd. All reagents used were purchased commercially and were of analytical grade, without any purification procedures.

[0053] Instruments and Equipment: AVANCED RX400 nuclear magnetic resonance spectrometer (Bruker, Germany); APEX-IICCD single-crystal X-ray diffractometer (Bruker, Germany); 6540Q-TOF high-resolution mass spectrometer (Agilent Technologies, USA); F-4700 fluorescence spectrophotometer (Hitachi, Japan); Fluorolog-3 transient / steady-state fluorescence spectrometer (Horiba, Japan); 1260infinity II high-performance liquid chromatograph (Agilent Technologies, USA); Smartlab SE powder X-ray diffractometer (Rigaku, Japan); Thermo Nicolet 10 Fourier transform infrared spectrometer (Olympus, Japan); UV-3600 ultraviolet-visible-near-infrared spectrophotometer (Shimadzu, Japan).

[0054] LABRAMHR EVO Raman spectrometer (Horiba Corporation, Japan); Mettler-Toledo 1600LF thermogravimetric analyzer (Mettler Toledo Corporation, Switzerland); JEM 2100 transmission electron microscope (JEOL Corporation, Japan); K-Alpha X-ray photoelectron spectrometer (Thermal Instruments, USA).

[0055] Example 1: Synthesis of coordination polymers α-CuI-m-iah and β-CuI-m-iah

[0056] 1. Preparation of coordination polymer α-CuI-m-iah: The ligand m-iah (28 mg, 0.2 mmol) was dissolved in 3 mL of ethanol and added dropwise to a solution of CuI (194.4 mg, 1.0 mmol) dissolved in acetonitrile (15 mL) at 25 °C. The mixture was stirred at 680 rpm for 4 hours. A white precipitate (α-CuI-m-iah) gradually appeared. The precipitate was separated by centrifugation, and the powder was washed five times each with ethanol and acetonitrile. The powder was then dried in a vacuum drying oven at 40 °C for 16 hours to obtain pure α-CuI-m-iah with a yield of 46.5%. To improve the dispersibility of the coordination polymer, α-CuI-m-iah solid (340 mg) was ground in a ball mill with acetonitrile (6 mL) for 12 hours.

[0057] 2. Preparation of coordination polymer β-CuI-m-iah: The ligand m-iah (28 mg, 0.2 mmol) was dissolved in 3 mL of ethanol and added dropwise to a solution of CuI (7.8 mg, 0.04 mmol) dissolved in acetonitrile (0.6 mL) at 25 °C. The mixture was stirred at 680 rpm for 4 hours. A white precipitate gradually appeared. The precipitate was separated by centrifugation. The centrifuged powder was washed five times each with ethanol and acetonitrile, and then dried in a vacuum drying oven at 40 °C for 16 hours to obtain pure β-CuI-m-iah, with a yield of 24.3%.

[0058] The reaction equation is as follows Figure 1 As shown in Figure A.

[0059] 3. Single Crystal Cultivation of β-CuI-m-iah Coordination Polymer: β-CuI-m-iah single crystals were prepared by diffusion method at room temperature. 8 mg of the ligand was dissolved in 1 mL of ethanol and placed at the bottom of a test tube. 4 mL of acetonitrile solution was slowly added as a buffer layer. Finally, 5 mg / mL cuprous iodide acetonitrile solution was slowly added to the top layer. After one day, single crystals were obtained under static conditions.

[0060] Performance analysis of the product obtained in Example 1:

[0061] 1. Single-crystal structure analysis of β-CuI-m-iah:

[0062] Needle-like crystals of CuI and 3-pyridinecarboxylhydrazine (m-iah) (molar ratio 1:2) were prepared at room temperature via diffusion. Single-crystal X-ray diffraction studies showed that the β-CuI-m-iah crystals belonged to the orthorhombic crystal system of space group Pna21. Crystallographic data showed that the chemical formula of β-CuI-m-iah is [C6H7CuIN3O]. n Other data are detailed in Table 1. Each asymmetric structural unit contains one m-iah ligand and half of a [Cu2I2] cluster. The Cu(I) center presents only one coordination environment, that is, each Cu(I) atom is coordinated with three bridging iodine atoms and one pyridine N atom of a ligand in a four-coordinate mode, and the local coordination geometry is a tetrahedral structure. Figure 1 B). Single-crystal structure analysis shows that β-CuI-m-iah consists of a one-dimensional chain along the c-axis, which is composed of double-stranded (Cu2I2) chains. n The structure is composed of a band, supported on both sides by a 3-pyridinecarboxylhydrazine ligand via Cu-N coordination bonds. Figure 1 D). β-CuI-m-iah exhibits a symmetrical rhombic [Cu2I2] metallic center with equal Cu···Cu distances. Very close to twice the van der Waals radius of Cu This indicates the presence of significant copper-philic interactions within the [Cu₂I₂] secondary building blocks. Furthermore, the coordination mode of the m-iah ligand is monodentate coordination (…). Figure 1 C), only the nitrogen atom of the pyridine ring is bonded to a single Cu(I) atom, arranged in two directions along the CuI chain. The Cu-N and Cu-I bond lengths are respectively... and The I-Cu-I bond angle is 99.88–115.99°. Figure 1 E). In the CuI 1D chain structure, intermolecular forces exist between the organic ligands, and the distance from the center of mass to the center of mass is... There are π…π stacking interactions between the pyridine rings. Furthermore, as… Figure 1 As shown in F and G, β-CuI-m-iah exhibits hydrogen bonding interactions (N…H) between the terminal N of the hydrazide group in the m-iah ligand and the H of the adjacent ligand forming the next chain. This plays a crucial role in the structural stability. Comparison of the experimentally measured powder X-ray diffraction (PXRD) pattern of β-CuI-m-iah powder with the simulated PXRD pattern showed that the main diffraction peaks were completely identical, indicating that the synthesized β-CuI-m-iah is a pure phase. Figure 2 ).

[0063] Table 1 Crystal structure data of β-CuI-m-iah

[0064]

[0065] 2. FT-IR characterization of ligand m-iah and coordination polymers α-CuI-m-iah and β-CuI-m-iah

[0066] To investigate the coordination interaction between the metal and the ligand, Fourier transform infrared spectroscopy (FT-IR) was used to characterize the ligand m-iah and the coordination polymers α-CuI-m-iah and β-CuI-m-iah.

[0067] FT-IR spectrum ( Figure 5 The display shows that it is located at 710cm. -1 835cm -1 and 886cm -1 The out-of-plane bending vibration absorption peak of pyridine and the 1340 cm⁻¹ -1 The absorption peak of the stretching vibration of the pyridine ring C=C is weakened. Compared with the ligand m-iah, α-CuI-m-iah and β-CuI-m-iah show a stronger absorption peak at 1550 cm⁻¹. -1 The absorption peaks of the pyridine C=N stretching vibration at 1000 cm⁻¹ were weakened and shifted to 1510 cm⁻¹. -1 and 1530cm -1This indicates that the pyridine nitrogen atom of m-iah is coordinated with Cu(I).

[0068] 3. Temperature-dependent PXRD of coordination polymers α-CuI-m-iah and β-CuI-m-iah

[0069] To investigate the thermal stability of the products, two coordination polymers were subjected to temperature-dependent PXRD and thermogravimetric analysis (TGA). The powders were heated from 40°C to 120°C and PXRD measurements were performed. The PXRD results showed that the diffraction peaks did not change (e.g., ...). Figure 6 A and B) indicate that the crystal structure remains intact below 120℃.

[0070] 4. Thermogravimetric (TGA) analysis of ligand m-iah and coordination polymers α-CuI-m-iah and β-CuI-m-iah

[0071] Thermogravimetric analysis curves under nitrogen atmosphere are as follows: Figure 6 C shows that α-CuI-m-iah begins to show mass loss at 250℃, indicating that the coordination polymer is stable in this temperature range. The weight loss between 250℃ and 490℃ can be attributed to the collapse of the coordination polymer backbone, while the weight loss between 490℃ and 800℃ is due to the loss of organic ligands. The final residues are metal oxides and ligand residues.

[0072] Before 200℃, β-CuI-m-iah loses approximately 0.63% weight due to solvent evaporation. A first weight loss step occurs between 200℃ and 495℃, followed by a second weight loss step after 495℃ due to ligand carbonization, indicating that the β-CuI-m-iah framework can be stably retained up to 200℃. Therefore, both coordination polymers exhibit good thermal stability. However, α-CuI-m-iah is more stable than β-CuI-m-iah, and the different decomposition temperatures may be due to the different crystal structures of these two coordination polymers.

[0073] 5. Chemical stability study of coordination polymers α-CuI-m-iah and β-CuI-m-iah

[0074] To investigate the chemical stability of the CuI-m-iah coordination polymers, the two coordination polymers were immersed in various solvents for 24 hours, including acetone, N,N-dimethylacetamide, ethanol, ethyl acetate, water, acetonitrile, methanol, tetrahydrofuran, N,N-dimethylformamide, and dimethyl sulfoxide. After 48 hours, they were centrifuged and dried, and their PXRD spectra were measured.

[0075] from Figure 7As shown in A and B, the PXRD pattern of α-CuI-m-iah does not change significantly, while the crystallinity of β-CuI-m-iah decreases in DMSO and DMF, and the peak shape changes in acetone solvent. The PXRD patterns are completely consistent after immersion in other solvents. This indicates that both coordination polymers have good solvent stability.

[0076] In addition, the CuI-m-iah coordination polymer powder was dispersed in Tris-HCl buffer solutions of different pH values, soaked for 24 hours, centrifuged, dried, and PXRD was measured. The results are as follows: Figure 7 As shown in C and D, the crystallinity of α-CuI-m-iah did not change significantly in 0.1M strong acid (HCl) and strong base (NaOH) and in 0.01M Tris-HCl buffer solution with pH 3.0–12.0. The crystallinity of β-CuI-m-iah was slightly lower in 0.1M strong alkaline environment, but good under other conditions. This further confirms that the two coordination polymers have good structural stability.

[0077] The above results indicate that the coordination polymer CuI-m-iah possesses good thermal and chemical stability, which provides a solid foundation for its future applications.

[0078] 6. Luminescent properties of CuI-m-iah

[0079] The solid-state excitation, emission, and UV-Vis absorption spectra of m-iah, α-CuI-m-iah, and β-CuI-m-iah at room temperature were tested. Figure 8 As shown in Figure A, the absorption band of ligand m-iah in the 200 nm–280 nm range can be attributed to π→π* electronic transitions. After forming a coordination polymer with CuI, the absorption peaks all red-shift, and the red-shift of α-CuI-m-iah (peak value 360 ​​nm) is greater than that of β-CuI-m-iah (peak value 310 nm), which may be related to their different crystal environments. Figure 8 As shown in Figure B, the m-iah ligand exhibits virtually no fluorescence under 365 nm excitation.

[0080] α-CuI-m-iah exhibits a distinct yellow emission upon excitation at 366 nm, with a maximum emission wavelength of 513 nm. β-CuI-m-iah, on the other hand, shows a maximum emission wavelength of 456 nm upon excitation at 365 nm. The fluorescence emission of both α-CuI-m-iah and β-CuI-m-iah is likely due to the combined effect of the short Cu…Cu distance within the Cu₂I₂ secondary building blocks and the metal-to-ligand charge transfer (MLCT).

[0081] The luminescence properties of CuI-m-iah in solution were investigated. 0.5 mg of CuI-m-iah was dispersed in 1 mL of different solvents, and its luminescence under natural light was recorded. Figure 9 A and B) and 365nm ultraviolet light ( Figure 9 The images were taken under C and D irradiation, and the fluorescence emission spectra of CuI-m-iah suspensions dispersed in different solvents were measured.

[0082] The results are as follows Figure 9 As shown in E and F, α-CuI-m-iah exhibits strong luminescence intensity in most solvents, such as EtOAc and EtOH, while showing quenched luminescence in DMSO. With increasing solution polarity, the fluorescence emission wavelength does not change significantly, with the maximum emission wavelength remaining at 513 nm, indicating that solvent polarity has little effect on the luminescence of α-CuI-m-iah. β-CuI-m-iah exhibits strong luminescence intensity in MeOH, while showing quenched luminescence in DMF and DMSO.

[0083] Depend on Figure 9 It is known that CuI-m-iah dissolves in DMSO, but is dispersed in most other solvents. Therefore, CuI-m-iah emits light in unsuitable solvents but does not emit light in good solvents.

[0084] 7. Solubility and aggregation state of coordination polymers:

[0085] DMSO was chosen as a good solvent and EtOH as a poor solvent to study its aggregation process. Figure 10 Images of CuI-m-iah coordination polymers under natural light and 365 nm UV irradiation containing different amounts of ethanol are shown. The results show that the emission intensity gradually increases with increasing content of the undesirable solvent ethanol.

[0086] Fluorescence spectra of DMSO / EtOH mixed solutions with different ethanol components were measured. Figure 11 In EtOH at low volume ratios, α-CuI-m-iah fluorescence is weak or non-fluorescent.

[0087] When the volume fraction of EtOH (fe, where fe represents the volume ratio of EtOH) increases from 30% to 70%, the fluorescence intensity of α-CuI-m-iah increases rapidly and reaches its maximum value at 70%. Its fluorescence intensity is about 49 times stronger than that of pure DMSO (fe = 0%). The significant enhancement in emission intensity proves that α-CuI-m-iah has AIE properties.

[0088] Furthermore, with increasing ethanol content, the emission wavelength of α-CuI-m-iah redshifts ( Figure 11A) The emission wavelength shifted from 476 nm to 511 nm. As can be seen from the above analysis of the solvent effect of α-CuI-m-iah, both the fluorescence intensity and wavelength are related to the degree of aggregation of CuI-m-iah.

[0089] β-CuI-m-iah exhibits similar phenomena to α-CuI-m-iah; when the EtOH content is less than 50%, the fluorescence intensity is very low, and it basically does not fluoresce. Figure 11 C). As the EtOH content increases, the AIE effect in the system becomes more pronounced, and the fluorescence intensity gradually increases. The highest fluorescence intensity is obtained when fe is 100%, which is 41.4 times higher than when fe is 0% (pure DMSO solution). Figure 11 D).

[0090] Examples 2-19: Synthesis of coordination polymers α-CuI-m-iah and β-CuI-m-iah using different molar ratios of CuI to m-iah.

[0091] The synthesis method differs from that in Example 1 in that the molar ratio of CuI to m-iah is as follows:

[0092] Example number CuI:m-iah molar ratio Example number 2 1:5 11 3 1:4 12 4 1:3 13 5 1:2 14 6 1:1 15 7 2:1 16 8 3:1 17 9 4:1 18 10 5:1 19

[0093] The products obtained in Examples 2-10 were powders; Examples 11-19 were obtained by solid-phase grinding synthesis. The solid-phase grinding synthesis method used was to add CuI and m-iah in proportion to a mortar and grind for 6 minutes. During the grinding process, 1-2 drops of acetonitrile were added to promote the synthesis of coordination polymers.

[0094] In the above embodiments, the molar ratio of metal to ligand was adjusted from 1:5 to 5:1, and the nine synthesized products were subjected to PXRD analysis. The PXRD results of the substances obtained in Examples 2-10 are shown below. Figure 3 As shown in Figure A, the PXRD results indicate that when the ratio of CuI to m-iah is in the range of 1:5 to 1:2, a crystalline coordination polymer is obtained, named β-CuI-m-iah.

[0095] When the molar ratio of CuI to m-iah is in the range of 1:1 to 5:1, another peak shape appears, resulting in a coordination polymer with another crystal form, named α-CuI-m-iah.

[0096] The PXRD patterns of the products obtained in Examples 11-19 are as follows: Figure 3 As shown in B, the main diffraction peaks of the powder samples synthesized under various ratios are consistent with those of the samples synthesized by the solution method. This method is simple and relatively time-consuming, indicating that CuI-m-iah can be synthesized on a large scale using the solid-phase grinding synthesis method.

[0097] Performance testing and analysis:

[0098] 1. The products obtained in Examples 2-10 are a group of coordination polymer powders with tunable optical properties. Under 365 nm ultraviolet light irradiation, their emission color changes from blue to yellow with increasing CuI ratio (Figure 12D). The fluorescence lifetime, UV-Vis absorption spectrum, and fluorescence emission spectrum of coordination polymers with different ratios were tested. Figure 12 As shown in Figure C, with the increase of CuI ratio, its fluorescence emission wavelength redshifts from 456 nm to 513 nm. PXRD results also indicate that... Figure 1 A) The peak shape of the coordination polymer changed, indicating that as the CuI ratio increased, the product gradually transformed from the β-CuI-m-iah phase to the α-CuI-m-iah phase.

[0099] The spectral detection results are shown in the table below.

[0100] Table: Spectral data of CuI-m-iah

[0101]

[0102]

[0103] As can be seen from the spectral data in the table above, with the increase of CuI ratio, the product gradually transforms from the β-CuI-m-iah phase to the α-CuI-m-iah phase.

[0104] Examples 20-27: Synthesis of coordination polymers α-CuI-m-iah and β-CuI-m-iah at different reaction temperatures

[0105] The differences between the synthesized coordination polymers α-CuI-m-iah and β-CuI-m-iah and those in Example 1 are as follows: The reaction temperatures are shown in the table below:

[0106]

[0107]

[0108] Depend on Figure 4 Results A and B show that the diffraction peaks of the obtained products did not change, indicating that temperature has little effect on the structure of the products.

[0109] Examples 28-31: Synthesis of coordination polymers α-CuI-m-iah and β-CuI-m-iah using different reaction solvents

[0110] The differences between the synthesized coordination polymers α-CuI-m-iah and β-CuI-m-iah and those in Example 1 are as follows: The reaction temperatures are shown in the table below:

[0111]

[0112] As shown in the table above, all components are soluble in acetonitrile solution. The synthesized products were then mixed with a solution (ethanol and acetonitrile), and the results are as follows: Figure 4 As shown in C and D, the diffraction peaks remained unchanged, indicating that CuI-m-iah can also be prepared in a pure acetonitrile system.

[0113] The properties of the products obtained in Examples 2 to 31 are similar to the test results in Example 1, and will not be repeated here.

[0114] Example 32: Study on the detection performance of α-CuI-m-iah against thiophanate-methyl (TM)

[0115] 1. Fluorescence Properties Study: 0.1 mM thiophanate-methyl was added to a DMF solution of α-CuI-m-iah (0.5 mg / mL), and after 30 minutes, its fluorescence emission spectrum was measured as follows: Figure 13 A. Under 339 nm excitation, the addition of thiophanate-methyl resulted in a decrease in the fluorescence intensity of the suspension, down by 81.69% compared to the original α-CuI-m-iah. Based on this phenomenon, we attempted to use α-CuI-m-iah to identify and detect thiophanate-methyl. To obtain a highly sensitive fluorescence sensor for detecting thiophanate-methyl, we optimized the detection conditions.

[0116] 2. Detection response time: 100.0 μM thiophanate-methyl was added to a suspension of α-CuI-m-iah (0.5 mg / mL), and the change in fluorescence intensity of the suspension over time was recorded. The results are as follows: Figure 13 As shown in Figure B, the fluorescence intensity of the suspension decreased rapidly 2 minutes after the addition of thiophanate-methyl. The fluorescence intensity stabilized and remained balanced after approximately 10 minutes, indicating that the α-CuI-m-iah detection method for thiophanate-methyl has a short response time and can perform rapid and efficient detection in practical applications.

[0117] 3. Verification of the selectivity of α-CuI-m-iah for thiophanate-methyl: Take 1 ml of standard solutions of thiophanate-methyl, flubendiamide, carbendazim, acetamiprid, triazole, metalaxyl, iprodione, triadimefon, and acetamiprid at the same concentration (100.0 μM) and add them to 0.5 mg / mL of α-CuI-m-iah suspension for selectivity testing. A blank control was used.

[0118] Experimental results are as follows Figure 13 As shown in Figure C, it can be observed that other pesticide molecules have little or no effect on α-CuI-m-iah, while only thiophanate-methyl has a significant quenching effect on the luminescence of α-CuI-m-iah. This indicates that the α-CuI-m-iah coordination polymer can achieve selective recognition of thiophanate-methyl with high specificity.

[0119] 4. Evaluation of the effect of interfering substances on the sensing performance of α-CuI-m-iah: 1 ml of aqueous solutions of different interfering substances (iprodione, triadimefon, acetamiprid, metalaxyl, triazole, thiophanate-methyl, carbendazim) at a concentration of 1 mM were added to 0.5 mg / mL α-CuI-m-iah dispersions. Then, 0.1 mM thiophanate-methyl was added to the above solutions, and the luminescence intensity of α-CuI-m-iah was measured when the two substances coexisted. The results are as follows: Figure 13 As shown in Figure D, the fluorescence intensity change of α-CuI-m-iah at 469 nm is similar to that of the solution with only thiophanate-methyl added.

[0120] This indicates that the quenching effect of thiophanate-methyl on α-CuI-m-iah is not affected by coexisting components, confirming that α-CuI-m-iah has good anti-interference ability in the presence of interfering substances.

[0121] 5. To further explore the quantitative detection potential of α-CuI-m-iah in thiophanate-methyl, the relationship between thiophanate-methyl concentration and the luminescence intensity of α-CuI-m-iah was investigated. 20 μL of thiophanate-methyl standard solutions of different concentrations were taken respectively, see [reference missing]. Figure 14 The concentrations of 0 μM, 10 μM, 20 μM, 30 μM, 40 μM, 50 μM, 60 μM, 70 μM, 80 μM, and 90 μM were added to a suspension of α-CuI-m-iah (0.5 mg / mL, 1 mL). After reacting for 20 minutes, the fluorescence spectra of the methyl thiophanate solutions at each concentration were recorded to study the analytical performance of the α-CuI-m-iah sensor under optimal experimental conditions.

[0122] The results are as follows Figure 14 As shown, the fluorescence intensity of the α-CuI-m-iah suspension gradually decreased with increasing thiophanate-methyl concentration. Adding thiophanate-methyl solution within the concentration range of 0.5 μM to 90 μM resulted in a linear decrease in fluorescence, with a relatively wide linear range. The linear regression equation was I = 379.40 - 3.85C, with a correlation coefficient R² = 0.9960, and the limit of detection (LOD = 3σ / k) was 0.41 μM, or 0.167 mg / kg (where σ is the standard deviation of the blank and k is the slope of the calibration curve). Conclusion: The coordination polymer α-CuI-m-iah can be used for quantitative detection of thiophanate-methyl in samples within the concentration range of 0.5 μM to 600 μM.

[0123] Example 32: Detection of α-CuI-m-iah in samples containing methyl thiophanate

[0124] 1. Pretreatment of tobacco leaf samples

[0125] Weigh 2.0 g of tobacco powder sample, add 20 mL of hydrochloric acid solution (4 mol / L), and extract magnetically for 1 h in an oil bath at 120 °C. After cooling to room temperature, centrifuge and collect the supernatant for detection. To avoid the acidity of the tobacco extract affecting the fluorescence properties of the probe, dilute the supernatant 10 times before adding it to the probe solution.

[0126] 2. Determination of thiophanate-methyl

[0127] Configure the following steps:

[0128] Treatment 1: 50 μL of 10 mg / mL α-CuI-m-iah solution + 5 μL of 50 μM thiophanate-methyl standard solution + 945 μL of supernatant diluted 10 times;

[0129] Treatment 2: 50 μL of 10 mg / mL α-CuI-m-iah solution + 6 μL (60 μM) thiophanate-methyl standard solution + 944 μL of supernatant diluted 10 times;

[0130] Treatment 3: 50 μL of 10 mg / mL α-CuI-m-iah solution + 9 μL (90 μM) thiophanate-methyl standard solution + 941 μL of supernatant diluted 10 times;

[0131] The mixtures from each treatment were vortexed until homogeneous and incubated at room temperature for 10 min. Fluorescence spectra were recorded and collected in the range of 380–690 nm using 339 nm as the excitation wavelength. The fluorescence intensity at 469 nm was linearly fitted to the concentration of thiophanate-methyl when different concentrations of thiophanate-methyl were added to the probe solution.

[0132] 3. Calculation of pesticide residues

[0133] Calculate the residue (R, mg / kg) of thiophanate-methyl on a dry basis according to formula (1):

[0134]

[0135] Where: C—concentration of thiophanate-methyl calculated from the standard working curve, μmol / L; V—volume of extract, mL; M—molar mass of thiophanate-methyl, g / mol; m—mass of the sample to be tested, g.

[0136] 4. This method demonstrates good sensitivity and selectivity for detecting thiophanate-methyl in DMF. Further exploration is needed to demonstrate its applicability in DMF from tobacco extracts using the standard spiking method. The original mass concentration of the actual sample was first determined by HPLC, and then thiophanate-methyl was detected using this fluorescence detection method. A spiked recovery experiment was also conducted.

[0137] Following the procedures described in steps 1-3 above, different concentrations (40 μM, 60 μM, 80 μM) of thiophanate-methyl standard solution were added to an α-CuI-m-iah solution containing the supernatant from the smoke extract to obtain the test solution. The test solution was then subjected to fluorescence detection, and fluorescence spectra and intensities were collected. The experiment was repeated five times at each concentration. The same test solution was then analyzed using HPLC.

[0138] Table 1 shows the experimental results of α-CuI-m-iah detection of thiophanate-methyl in tobacco dust (n=5).

[0139]

[0140] Calculate the relative standard deviation (RSD) and the corresponding recovery rates. As shown in the table above, the RSDs range from 1.2% to 7.6%, and the recoveries range from 94.51% to 109.87%.

[0141] Example 33

[0142] The difference from Example 2 is that the grinding time is 5 minutes;

[0143] Example 34

[0144] The difference from Example 2 is that the grinding time is 10 minutes;

[0145] in conclusion

[0146] Replacing the detection medium with DMF solvent revealed that α-CuI-m-iah can also be used as a fluorescent probe for the determination of thiophanate-methyl. The α-CuI-m-iah probe exhibits a wide linear range (0.5 μM–90 μM), low detection limit (0.41 μM, 0.167 mg / kg), and short response time (10 min). Further mechanistic studies indicate that, based on the strong thiophilicity of Cu(I), thiophanate-methyl competes with the ligand for coordination, leading to the release of some ligands and thus causing fluorescence quenching in the solution. Applying this probe to the detection of thiophanate-methyl in tobacco samples within DMF yielded satisfactory recoveries (89.64%–107.72%).

[0147] Although the present invention has been described in detail with reference to the foregoing embodiments, those skilled in the art can still modify the technical solutions described in the foregoing embodiments or make equivalent substitutions for some of the technical features. Any modifications, equivalent substitutions, improvements, etc., made within the spirit and principles of the present invention should be included within the protection scope of the present invention.

Claims

1. A fluorescent probe for detecting thiophanate-methyl in tobacco, characterized in that, It is α-CuI-m-iah and has the following properties: PXRD measurements showed that the diffraction peaks did not change when heated from 40°C to 120°C. The thermal stability temperature of the thermogravimetric analysis curve under nitrogen atmosphere is 250℃. After soaking in DMSO and DMF for 24 hours, the PXRD spectrum showed no significant changes. The crystallinity did not change significantly in 0.1M HCl, 0.1M NaOH, and 0.01M Tris-HCl buffer solutions with pH 3.0–12.0; It exhibits a distinct yellow emission when excited at 366 nm, with a maximum emission wavelength of 513 nm; It exhibits quenching luminescence in DMSO; It possesses AIE properties; The morphology of the sample in DMSO / EtOH mixed solvent with a concentration of 20%, 40%, 60%, and 90% gradually changed from amorphous flocculent to blocky, and the highest fluorescence intensity was obtained when the concentration of atomized flocculent was 100%. The spectral data is: E m Greater than 470nm; Greater than 0.75%; k r / 10 7 Greater than 0.015s -1 .

2. The fluorescent probe for detecting thiophanate-methyl in tobacco according to claim 1, characterized in that, The linear detection range of α-CuI-m-iah for methyl thiophanate is 0.5 μM to 90 μM.

3. The fluorescent probe for detecting thiophanate-methyl in tobacco according to claim 1 and its application, characterized in that, The detection limit of α-CuI-m-iah for the fluorescence detection of thiophanate-methyl was 0.41 μM.

4. The fluorescent probe for detecting thiophanate-methyl in tobacco according to claim 1 and its application, characterized in that, The fluorescence detection response time of α-CuI-m-iah to thiophanate-methyl is at least 10 min.

5. A method for preparing a fluorescent probe for detecting thiophanate-methyl in tobacco as described in any one of claims 1 to 4, characterized in that, The process includes the following steps: grinding CuI and m-iah at room temperature, adding acetonitrile dropwise during the grinding process to obtain a fluorescent probe; Preferably, when the molar ratio of CuI:m-iah is 1:1 to 5:1, the product obtained is α-CuI-m-iah.

6. A method for preparing a fluorescent probe for the detection of thiophanate-methyl in tobacco as described in any one of claims 1 to 4, characterized in that, Includes the following steps: The m-iah solution and CuI solution were mixed and stirred until a white precipitate appeared. The fluorescent probe was obtained after post-processing. Preferably, when the molar ratio of CuI:m-iah is 1:1 to 5:1, the product obtained is α-CuI-m-iah.

7. The preparation method according to claim 6, characterized in that, The post-treatment involved centrifuging to separate the precipitate, washing the precipitate multiple times with organic solvents, and drying it. Preferably, the solvent used for the m-iah solution is ethanol; the solvent used for the CuI solution is acetonitrile.

8. The preparation method according to claim 6, characterized in that, The post-treatment involves centrifugation to separate the precipitate; the reaction temperature is 20–40°C; preferably, the reaction system contains at least acetonitrile.

9. A fluorescent probe detection method for thiophanate-methyl in tobacco, characterized in that, The steps include: mixing the sample with the above fluorescent probe solution to obtain a mixed solution, letting it stand for at least 10 minutes, using 339 nm as the excitation wavelength, recording and collecting the fluorescence spectrum in the range of 380–690 nm to observe whether the fluorescence intensity of the mixed solution decreases. If it decreases significantly, the sample contains thiophanate-methyl; if it does not decrease significantly, the sample does not contain thiophanate-methyl.

10. The detection method according to claim 7, characterized in that, The fluorescent probe is dissolved in DMF solution to obtain a fluorescent probe solution; Preferably, the concentration of the fluorescent probe in the fluorescent probe solution is greater than or equal to 0.5 mg / mL; Preferably, the sample pretreatment includes: after the sample is crushed into powder, it is added to a 4 mol / L hydrochloric acid solution, heated to 120°C under stirring, cooled to room temperature, centrifuged, and the supernatant is diluted and mixed with the fluorescent probe solution for detection; Preferably, the sample is tobacco; Preferably, the sample is a water sample.

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