A fluorescent material and its application in rapid detection of fish anesthetic MS-222

Abstract

The application discloses a fluorescent material, Eu@UiO-66-(COOH)2 is a rare earth Eu doped functionalized metal organic framework material UiO-66-(COOH)2, the morphology is a spherical structure, the particle size is 40-50 nm, the fluorescent material can detect the fishing anesthetic MS-222 with high selectivity, the detection time is short, the accuracy and stability are high, and the fishing anesthetic MS-222 residual on-site screening monitoring can be used in the aquatic product circulation link.

Description

Technical Field

[0001] This invention relates to the field of anesthetic detection technology, and in particular to a fluorescent material and its application in the rapid detection of the fishing anesthetic MS-222. Background Technology

[0002] In the live transport of aquatic products, the legal and compliant use of fish anesthetics, such as ethyl m-aminobenzoate methanesulfonate (MS-222), can reduce fish metabolism and stress responses, thereby improving survival rates. However, excessive or illegal use can lead to residues of fish anesthetics in the fish, threatening consumer health. Therefore, establishing effective methods for monitoring fish anesthetic residues is crucial.

[0003] Currently, the mainstream detection methods for fish anesthetics include liquid chromatography-mass spectrometry (LC-MS) and gas chromatography-mass spectrometry (GC-MS). While these methods offer high accuracy, they generally have the following limitations:

[0004] 1) Cumbersome pretreatment: It requires complex organic solvent extraction, purification and concentration steps, which can take up to several hours;

[0005] 2) High dependence on equipment: It relies on large-scale precision instruments, resulting in high costs;

[0006] 3) Long testing cycle: Aquatic products such as fish and shrimp have a short circulation cycle, while existing gas chromatography-mass spectrometry and liquid chromatography-mass spectrometry methods usually take more than a day from sampling to issuing results. This cannot meet the timeliness requirements of testing in the transportation and sales process, nor can it meet the needs of rapid testing for on-site food safety supervision.

[0007] Therefore, the food safety regulatory industry urgently needs an MS-222 on-site screening technology that is easy to operate, fast and intuitive, low in cost, and highly selective. Summary of the Invention

[0008] In view of this, the purpose of this invention is to address the shortcomings of the prior art by providing a fluorescent material that can be used for rapid detection of the fishing anesthetic MS-222, exhibiting strong stability, high accuracy, and short detection time, and can be used for on-site screening of MS-222 residues in aquatic products during the circulation process.

[0009] To achieve the above objectives, the present invention adopts the following technical solution:

[0010] A fluorescent material, wherein the fluorescent material Eu@UiO-66-(COOH)2 is a rare earth Eu-doped functionalized metal-organic framework material UiO-66-(COOH)2 with a spherical structure and a particle size of 40-50 nm.

[0011] A process for preparing a fluorescent material includes the following steps:

[0012] Take UiO-66-(COOH)2, Eu(NO3)3⋅6H2O and an appropriate amount of deionized water, and react them in a hydrothermal reactor at 80~100℃ and 100~200rpm for 15~24 hours. The weight ratio of Eu(NO3)3⋅6H2O to UiO-66-(COOH)2 is 1:1-5.

[0013] After cooling to room temperature, the solid product is washed, collected, and dried to obtain the final product.

[0014] Furthermore, the UiO-66-(COOH)2 is prepared by the following steps:

[0015] Pyromellitic anhydride and ZrCl4 were added to a hydrothermal reactor containing an appropriate amount of deionized water. The reactor was fixed in an air-heated oven that could rotate around an axis and reacted at 80-100℃ and 100-200rpm for 15-24 hours. After cooling to room temperature, the initial product was obtained by washing. The weight ratio of pyromellitic anhydride to ZrCl4 was 1-2.5:1.

[0016] Take the initial product and add it into a hydrothermal reactor. Add 5-20 times the weight of deionized water and react at 80-100℃ and 100-200rpm for 10-20 hours. After cooling to room temperature, clean the product, collect the solid product, and dry it to obtain the final product.

[0017] Application of a fluorescent material in the rapid detection of the fishing anesthetic MS-222.

[0018] Furthermore, using fluorescent materials as the photosensitive substances for fluorescence sensing analysis, a standard curve for the fluorescence sensing analysis of the fishing anesthetic MS-222 was plotted, including the following steps:

[0019] Preparation of standard sensing solution: Weigh an appropriate amount of fluorescent material and add deionized water to prepare a standard fluorescent sensing solution with a concentration of H0.

[0020] Preparation of MS-222 standard solutions: Add different volumes of MS-222 standard stock solution to the standard fluorescence sensing solution to prepare n MS-222 standard solutions of different concentrations, n≥5, with a concentration range of 0μmol / L-2000μmol / L;

[0021] Fluorescence scans were performed on MS-222 standard solutions of different concentrations at excitation wavelengths of 240-280 nm, and the fluorescence intensity I of the characteristic fluorescence emission peak at 550-650 nm was recorded; the fluorescence intensity of the MS-222 standard solution with a concentration of 0 was recorded as I0.

[0022] With the concentration [Q] of the MS-222 standard solution as the X-axis and I0 / I as the Y-axis, a standard curve for MS-222 fluorescence sensing analysis was plotted, and the Stern-Volmer equation was obtained by fitting: I0 / I=1+K[Q].

[0023] Furthermore, the method for rapid detection of the fishing anesthetic MS-222 includes the following steps:

[0024] (1) Sampling; pretreatment of the product to be tested to obtain the test solution;

[0025] (2) Detection;

[0026] Weigh an appropriate amount of fluorescent material and add deionized water to prepare a standard fluorescent sensing solution with a concentration of H0.

[0027] The test solution is added to the solution of the fluorescent material, and an appropriate amount of deionized water is added to prepare the fluorescent test solution. The final concentration of the fluorescent material in the fluorescent test solution is H0.

[0028] At an excitation wavelength of 240-280 nm, the standard fluorescent sensing solution and the fluorescent test solution were subjected to spectral scanning, and the fluorescence intensities at 550-650 nm were recorded as I0 and I, respectively.

[0029] (3) Substitute I0 and I from step (2) into the Stern-Volmer equation I0 / I=1+K[Q] to obtain the concentration of the test solution.

[0030] Furthermore, the pretreatment involves pretreating the product to be tested before sampling. For aquaculture water, the pretreatment method is as follows: take an appropriate amount of aquaculture water and pass it through a 0.22μm water-based filter membrane to obtain the filtrate as the aquaculture water test solution.

[0031] The pretreatment method for aquatic products is as follows: the aquatic products are crushed and pulped using a meat grinder, the homogenized sample is weighed into a centrifuge tube, MgSO4, NaCl and acetonitrile are added, vortexed and centrifuged, and the weight ratio of the homogenized sample to MgSO4, NaCl and acetonitrile is 5:1-3:0.5-2:10-15.

[0032] Take an appropriate amount of supernatant into a purification tube, vortex, centrifuge, and filter through a 0.22μm organic membrane to obtain the filtrate as the test solution for aquatic products.

[0033] Furthermore, in step (2), H0 is 100 μg / mL.

[0034] Furthermore, in step (2), the excitation wavelength is 255 nm, and the fluorescence emission peak at 613 nm is used as the indicator peak.

[0035] The beneficial effects of this invention are:

[0036] 1. This invention develops a fluorescent material Eu@UiO-66-(COOH)2, which is a rare-earth europium (Eu)-doped functionalized metal-organic framework material with a spherical structure and a particle size of 40-50 nm. After doping, Eu... 3+ It exists in a highly dispersed state in the channels or defect sites of Eu@UiO-66-(COOH)2, without forming a crystalline europium oxide or carboxylate impurity phase, and the relative atomic content of Eu doping is 13.44 wt%.

[0037] The applicant's research found that the fluorescent material synthesized in this application, when formulated into a sensing liquid, exhibits very strong fluorescence intensity near 591 nm and 613 nm under an excitation wavelength of 220 nm-280 nm. MS-222 shows a significant fluorescence quenching response to the Eu-MOF sensing liquid. Therefore, Eu-MOF sensing can be used for MS-222 fluorescence detection.

[0038] Since aquatic products sold in the market and their aquaculture water may contain the fish anesthetic MS-222, on-site testing is required. Existing gas chromatography-mass spectrometry and liquid chromatography-mass spectrometry methods are difficult to meet the rapid testing needs of on-site food safety supervision. Therefore, fluorescence detection based on Eu-MOF sensing can open up new directions for on-site detection of MS-222.

[0039] 2. The Eu@UiO-66-(COOH)2 synthesized in this application can selectively detect MS-222, exhibits strong stability, and resists interference. The principle is as follows: under acidic conditions, the amino groups on the benzene ring of the fish anesthetic MS-222 undergo a condensation reaction with the numerous exposed carboxyl groups (-COOH) in the fluorescent material Eu@UiO-66-(COOH)2, effectively drawing the MS-222 molecule closer to the Eu@UiO-66-(COOH)2 ring. 3+ Near the luminescent center, the amino group acts as an electron donor, undergoing a photoinduced electron transfer process with the excited Eu@UiO-66-(COOH)2, efficiently quenching the characteristic fluorescence of Eu@UiO-66-(COOH)2, resulting in a significant decrease in fluorescence intensity and enabling selective detection by MS-222.

[0040] Using the application and testing methods of this application, after simple pretreatment of aquaculture water or aquatic products, the sample is added to the fluorescent sensing liquid, enabling rapid qualitative and quantitative detection of MS-222 in aquatic products and their aquaculture water within 60 seconds.

[0041] The linear range of the fluorescence sensor analysis for detecting MS-222 is 7.65-1530.87 μmol / L, the limit of detection is 7.65 μmol / L, and the limit of quantitation is 22.95 μmol / L. It has high accuracy and strong stability, and can be used for on-site screening and monitoring of MS-222 residues in aquatic products during the circulation process.

[0042] 3. This application innovates the synthesis method of fluorescent material Eu@UiO-66-(COOH)2. The traditional method uses pyromellitic acid as raw material and synthesizes Eu@UiO-66-(COOH)2 by magnetic stirring and reflux in a flask. However, it has problems such as high cost of pyromellitic acid, difficulty in precise temperature control, and easy escape of solvent during reflux (reaching the boiling point of water, 100℃).

[0043] This application uses pyromellitic anhydride instead of pyromellitic acid as a raw material. It undergoes in-situ hydrolysis in high-temperature water to produce pyromellitic acid, achieving the same effect while reducing raw material costs by 30%. Furthermore, it employs a fully sealed, precisely temperature-controlled externally stirred PTFE hydrothermal reactor instead of a traditional magnetically stirred flask reflux reactor. The synthesis process, lasting up to 24 hours, can be completed with a single button setting. The overall stirring effect of the hydrothermal reactor rotating within a constant-temperature oven is significantly superior to magnetic stirring. The entire process requires no supervision, has no solvent evaporation, and multiple hydrothermal reactors can be placed simultaneously on the oven's rotating shaft, enabling large batch synthesis and allowing for the same batch synthesis with different raw material ratios, saving time, effort, and worry. The synthesis method is green and environmentally friendly, requiring no toxic reagents and facilitating widespread application. Attached Figure Description

[0044] Figure 1 This is a schematic diagram of the synthesis method of Eu@UiO-66-(COOH)2;

[0045] Figure 2 This is a flowchart of the detection method of the present invention;

[0046] Figure 3 Infrared spectra of UiO-66-(COOH)2 and Eu@UiO-66-(COOH)2;

[0047] Figure 4 X-ray diffraction patterns of UiO-66-(COOH)2 and Eu@UiO-66-(COOH)2;

[0048] Figure 5 The SEM and EDS spectra of UiO-66-(COOH)2 are shown.

[0049] Figure 6 SEM and EDS spectra of Eu@UiO-66-(COOH)2;

[0050] Figure 7 TEM image of Eu@UiO-66-(COOH)2;

[0051] Figure 8 The fluorescence emission characteristic peak of the Eu-MOF sensing fluid at an excitation wavelength of 255 nm is shown.

[0052] Figure 9 The fluorescence emission spectra of 100 μg / mLEu-MOF sensing fluid under different excitation wavelengths are shown.

[0053] Figure 10 Fluorescence emission spectra of Eu-MOF sensing solutions at different concentrations;

[0054] Figure 11 Fluorescence quenching of Eu-MOF sensing fluid by different concentrations of MS-222;

[0055] Figure 12 Calibration curves for fluorescence sensing analysis of MS-222;

[0056] Figure 13 The fluorescence response of Eu-MOF sensing liquids with different added chemicals under 254nm ultraviolet light irradiation;

[0057] Figure 14 The effect of different chemical substances on the intensity of the fluorescence characteristic peak at 591 nm of Eu-MOF sensing fluid;

[0058] Figure 15 The effect of different chemical substances on the intensity of the fluorescence characteristic peak at 613 nm of Eu-MOF sensing fluid;

[0059] Figure 16 Fluorescence spectra of different chemical substances in Eu-MOF sensing fluid;

[0060] Figure 17 The fluorescence intensity changes of the Eu-MOF sensing solution within different pH ranges;

[0061] Figure 18 The fluorescence intensity change of the Eu-MOF sensing fluid over 10 days;

[0062] Figure 19 The fluorescence emission spectrum of the positive sample;

[0063] Figure 20 The total ion chromatogram of the positive sample is obtained by liquid chromatography-mass spectrometry. Detailed Implementation

[0064] The present invention will be further described below with reference to the accompanying drawings and embodiments, wherein Eu-MOF is an abbreviation for Eu@UiO-66-(COOH)2 and MOF is an abbreviation for UiO-66-(COOH)2, and the water used in the following specific embodiments and application tests is deionized water.

[0065] Example 1

[0066] A fluorescent material, Eu@UiO-66-(COOH)2, is a rare-earth europium (Eu)-doped functionalized metal-organic framework material with a spherical morphology and a particle size of 50 nm. Its synthesis method is described in [link to original text]. Figure 1 As shown, it includes the following steps:

[0067] (1) Synthesis of metal-organic framework material UiO-66-(COOH)2

[0068] 4.3 g of pyromellitic anhydride and 2.3 g of ZrCl4 were added to a 100 mL polytetrafluoroethylene hydrothermal reactor containing 50 mL of deionized water. The reactor was fixed in an air-heated oven (externally stirred hydrothermal reactor) that could rotate around its axis, and reacted at 100 °C and 120 rpm for 24 hours. After naturally cooling to room temperature, the product was transferred to a centrifuge tube and centrifuged at 10,000 rpm for 10 min. The supernatant was discarded, and 40 mL of deionized water was added to the centrifuge tube. The mixture was shaken well and centrifuged again at 10,000 rpm for 10 min. This washing process was repeated three times to obtain the initial product.

[0069] Add 5g of the initial product to a polytetrafluoroethylene hydrothermal reactor, add 50mL of deionized water, and react again at 100℃ and 120 rpm for 16 hours. After naturally cooling to room temperature, transfer the product to a centrifuge tube, centrifuge at 10000rpm for 10min, discard the supernatant, add 40ml of acetone, shake well, and centrifuge at 10000rpm for 10min. Repeat this washing process three times. Collect the solid product and vacuum dry at 60℃ for 12h to obtain a white powder product UiO-66-(COOH)2.

[0070] (2) Synthesis of rare earth europium (Eu) doped functionalized fluorescent material Eu@UiO-66-(COOH)2

[0071] Take 0.5g of the synthesized UiO-66-(COOH)2 white powder, 0.223g of Eu(NO3)3·6H2O, and 50mL of deionized water, and add them together into a 100mL polytetrafluoroethylene hydrothermal reactor. React at 100℃ and 120 rpm for 24 hours.

[0072] After cooling to room temperature, the product was transferred to a centrifuge tube and centrifuged at 10,000 rpm for 10 min. The supernatant was discarded, and 40 mL of deionized water was added to the centrifuge tube. The mixture was shaken well and centrifuged at 10,000 rpm for 10 min again. This washing process was repeated three times. Then, 40 mL of acetone was added, the mixture was shaken well, and centrifuged at 10,000 rpm for 10 min. This washing process was repeated three times. The solid product was collected and dried under vacuum at 60 °C for 10 h to obtain a white powder product Eu@UiO-66-(COOH)2.

[0073] The synthesized fluorescent material Eu@UiO-66-(COOH)2 was used as the photosensitizer for rapid detection of the fishing anesthetic MS-222. The specific method is described in the following steps:

[0074] (1) Sampling: Sampling requires pretreatment of the samples. In this embodiment, two samples are used: aquaculture water and aquatic products.

[0075] 1) The pretreatment method for aquaculture water is as follows: take an appropriate amount of aquaculture water and filter it through a 0.22μm water system filter membrane to obtain the filtrate as the test solution of aquaculture water;

[0076] 2) The pretreatment method for aquatic products is as follows: The aquatic products are pulverized and pulped using a meat grinder. 5g of the homogenized sample is weighed into a centrifuge tube, and 2g of MgSO4, 1g of NaCl, and 10mL of acetonitrile are added. The mixture is vortexed for 5 minutes and then centrifuged at 8000 rpm for 5 minutes. 5mL of the supernatant is taken and added to a purification tube (containing 500mg MgSO4, 40mg C...). 18 Vortex in 20 mg PSA and 20 mg GCB for 1 min, centrifuge at 8000 rpm for 5 min, filter 3 mL of supernatant through a 0.22 μm organic membrane, and obtain the filtrate as the test solution for aquatic products;

[0077] (2) Testing:

[0078] 1) Preparation of standard sensing solution: Eu@UiO-66-(COOH)2 is added to deionized water to prepare a fluorescent sensing solution (0.5 mg / mL Eu@UiO-66-(COOH)2 aqueous solution);

[0079] Take 400 μL of fluorescence sensing solution (0.5 mg / mL Eu@UiO-66-(COOH)2 aqueous solution), add 1600 μL of deionized water, mix well, and record it as the standard fluorescence sensing solution with a concentration H0 of 100 μg / mL;

[0080] 2) Preparation of aquaculture water sensing solution: Take 400 μL of fluorescence sensing solution (0.5 mg / mL Eu@UiO-66-(COOH)2 aqueous solution), add 500 μL of aquaculture water test solution, add 1100 μL of deionized water, mix well, and record the concentration of the aquaculture water sensing solution as [Q]. a ;

[0081] 3) Preparation of aquatic product sensing solution: Take 400 μL of fluorescence sensing solution (0.5 mg / mL Eu@UiO-66-(COOH)2 aqueous solution), add 300 μL of aquatic product test solution, add 1300 μL of deionized water, mix well, and record the concentration of the aquatic product sensing solution as [Q]. b ;

[0082] 4) Fluorescence spectroscopy scanning: At an excitation wavelength of 240-280 nm (255 nm in this example), fluorescence spectroscopy scanning was performed sequentially on the standard fluorescence sensing solution, aquaculture water sensing solution, and aquatic product sensing solution. The fluorescence intensity of the characteristic fluorescence emission peak at 550-650 nm (613 nm in this example) was recorded as I0, I... a and I b ;

[0083] (3) Plot the standard curve: accurately prepare MS-222 standard stock solution with a concentration of 10 mg / mL, and add different volumes of MS-222 standard stock solution to 2 mL of Eu-MOF sensing solution (100 μg / mL) to prepare 13 MS-222 standard solutions with different concentrations, ranging from 0 μmol / L to 2000 μmol / L.

[0084] The final concentrations of MS-222 at 13 concentration gradients were 0, 2, 5, 10, 20, 50, 100, 150, 200, 250, 300, 350, and 400 μg / mL, equivalent to 0, 7.65, 19.14, 38.27, 76.54, 191.36, 382.72, 574.07, 765.43, 956.79, 1148.15, 1339.51, and 1530.87 μmol / L, respectively. Fluorescence scanning at an excitation wavelength of 255 nm recorded the characteristic fluorescence emission peak intensities of the mixed solutions at 591 nm and 613 nm as I0, I1, I2, I3, I4, I5, I6, I7, I8, I9, and I... 10 I 11 I 12 , where I0 is the fluorescence intensity when the concentration of the MS-222 standard solution is 0.

[0085] See results Figure 11 As shown, with increasing MS-222 concentration, the fluorescence emission characteristic peak intensities of the mixed solution at 591 nm and 613 nm gradually decreased. Under 254 nm UV irradiation, the red fluorescence of the mixed solution gradually weakened with increasing MS-222 concentration, consistent with the experimental results showing a decrease in fluorescence emission characteristic peak intensity, indicating that MS-222 has a significant fluorescence quenching response to the Eu-MOF sensing fluid.

[0086] The fluorescence intensity ratios I0 / I at 591 nm and 613 nm both showed a good linear relationship with the MS-222 concentration [Q]. The linear fitting equation for the fluorescence intensity ratio at 591 nm and the MS-222 molar concentration was I... 0(591nm) / I 591nm =1.03792+0.0023×[Q](R 2=0.993), the linear fitting equation for the fluorescence intensity ratio at 613 nm and the molar concentration of MS-222 is I. 0(613nm) / I 613nm =1.03321+0.0027×[Q](R 2 =0.997), such as Figure 12 As shown, the fluorescence quenching effect is quantitatively described by the Stern-Volmer equation, and the calculated fluorescence quenching constant is approximately 2700 L / mol, which is considered effective quenching.

[0087] The linear range of the MS-222 fluorescence sensor analysis is 2-400 μg / mL (equivalent to 7.65-1530.87 μmol / L), the method detection limit is 2 μg / mL (equivalent to 7.65 μmol / L), and the method quantification limit is 6 μg / mL (equivalent to 22.95 μmol / L). The fluorescence emission peak at 613 nm is due to Eu... 3+ The fluorescence emission peak at 613 nm is the typical fluorescence characteristic peak, with the highest intensity and better peak shape. It is less likely to be affected by other surrounding peaks. Therefore, in subsequent quantitative detection experiments, the fluorescence emission peak at 613 nm was used as the indicator peak.

[0088] Therefore, this application obtains the Stern-Volmer equation I0 / I=1+K[Q] through fitting, specifically using the fluorescence emission peak at 613nm as the indicator peak:

[0089] I0 / I = 1.03321 + 0.0027 × [Q] (R) 2 =0.997)

[0090] (4) Take the I0 and I measured in step (2) a and I b (where I) a and I b The fluorescence intensity I measured at 613 nm in the substitution equation was substituted into the fitted Stern-Volmer equation to calculate [Q]. a and [Q] b .

[0091] Example 2

[0092] The difference between Example 2 and Example 1 is that:

[0093] (1) Synthesis of metal-organic framework material UiO-66-(COOH)2

[0094] Add 2g of pyromellitic anhydride and 2g of ZrCl4 to a polytetrafluoroethylene hydrothermal reactor containing 50mL of deionized water. Fix the reactor in an air-heated oven (externally stirred hydrothermal reactor) that can rotate around its axis, and react at 80℃ and 100rpm for 15 hours. After naturally cooling to room temperature, transfer the product to a centrifuge tube, centrifuge at 10000rpm for 10min, discard the supernatant, add 40mL of deionized water to the centrifuge tube, shake well, and centrifuge again at 10000rpm for 10min. Repeat this washing process three times to obtain the initial product.

[0095] Take 3g of the initial product and add it to a polytetrafluoroethylene hydrothermal reactor. Add 30mL of deionized water and react at 80℃ and 100rpm for 10 hours. After naturally cooling to room temperature, transfer the product to a centrifuge tube and centrifuge at 10000rpm for 10min. Discard the supernatant, add 20mL of acetone, shake well, and centrifuge at 10000rpm for 10min. Repeat this washing process three times. Collect the solid product and vacuum dry it at 60℃ for 12h to obtain a white powder product UiO-66-(COOH)2.

[0096] (2) Synthesis of rare earth europium (Eu) doped functionalized fluorescent material Eu@UiO-66-(COOH)2

[0097] Take 0.5g of the synthesized UiO-66-(COOH)2 white powder, 0.5g of Eu(NO3)3⋅6H2O, and 50mL of deionized water, and add them together to a 100mL polytetrafluoroethylene hydrothermal reactor. React at 80℃ and 100rpm for 15 hours. After cooling to room temperature, transfer the product to a centrifuge tube and centrifuge at 10000rpm for 10min. Discard the supernatant, add 40mL of deionized water to the centrifuge tube, shake well, and centrifuge at 10000rpm for 10min again. Repeat this washing process three times. Add 20mL of acetone, shake well, and centrifuge at 10000rpm for 10min. Repeat this washing process three times. Collect the solid product and dry it under vacuum at 60℃ for 10h to obtain the white powder product Eu@UiO-66-(COOH)2.

[0098] Example 3

[0099] The difference between Example 3 and Example 1 is that:

[0100] (1) Synthesis of metal-organic framework material UiO-66-(COOH)2

[0101] 4.0 g of pyromellitic anhydride and 2.0 g of ZrCl4 were added to a polytetrafluoroethylene hydrothermal reactor containing 50 mL of deionized water. The reactor was fixed in an air-heated oven (externally stirred hydrothermal reactor) that could rotate around its axis, and reacted at 100 °C and 200 rpm for 20 hours. After naturally cooling to room temperature, the product was transferred to a centrifuge tube and centrifuged at 10,000 rpm for 10 min. The supernatant was discarded, and 50 mL of deionized water was added to the centrifuge tube. The mixture was shaken well and centrifuged again at 10,000 rpm for 10 min. This washing process was repeated three times to obtain the initial product.

[0102] Take 3g of the initial product and add it to a polytetrafluoroethylene hydrothermal reactor. Add 30mL of deionized water and react at 100℃ and 200rpm for 20 hours. After naturally cooling to room temperature, transfer the product to a centrifuge tube and centrifuge at 10000rpm for 10min. Discard the supernatant, add 50mL of acetone, shake well, and centrifuge at 10000rpm for 10min. Repeat this washing process three times. Collect the solid product and vacuum dry it at 60℃ for 12h to obtain a white powder product UiO-66-(COOH)2.

[0103] (2) Synthesis of rare earth europium (Eu) doped functionalized fluorescent material Eu@UiO-66-(COOH)2

[0104] Take 0.4g of the synthesized UiO-66-(COOH)2 white powder, 0.1g of Eu(NO3)3⋅6H2O, and 50mL of deionized water, and add them together to a 100mL polytetrafluoroethylene hydrothermal reactor. React at 90℃ and 150rpm for 20 hours. After cooling to room temperature, transfer the product to a centrifuge tube and centrifuge at 10000rpm for 10min. Discard the supernatant, add 40mL of deionized water to the centrifuge tube, shake well, and centrifuge at 10000rpm for 10min again. Repeat this washing process three times. Add 30mL of acetone, shake well, and centrifuge at 10000rpm for 10min. Repeat this washing process three times. Collect the solid product and dry it under vacuum at 60℃ for 10h to obtain the white powder product Eu@UiO-66-(COOH)2.

[0105] Structural characterization and application testing

[0106] The fluorescent material used in the following characterization is the product prepared in Example 1.

[0107] 1. Structural characterization of the fluorescent material Eu@UiO-66-(COOH)2

[0108] (1) Infrared spectroscopy analysis

[0109] The surface groups of UiO-66-(COOH)2 and Eu@UiO-66-(COOH)2 were characterized by Fourier transform infrared spectroscopy (FT-IR), such as... Figure 3 As shown. The peak at 1700 cm⁻¹ can be attributed to the stretching vibration of C=O in the uncoordinated carboxyl group (–COOH) (typical range 1720–1700 cm⁻¹), indicating that a small amount of uncoordinated carboxylic acid groups still exist in both materials. The characteristic antisymmetric and symmetric stretching vibration peaks of the carboxylic acid groups are located near 1570 cm⁻¹ and 1390 cm⁻¹, respectively. In the low wavenumber fingerprint region (typically 1390–400 cm⁻¹), the stretching vibrations of the Zr-O bond are located near 550 cm⁻¹ and 656 cm⁻¹, while the stretching vibrations of the Eu-O bond are located in the 450–500 cm⁻¹ region, indicating that Eu… 3+ It successfully coordinated with the carboxylic acid group.

[0110] (2) X-ray diffraction analysis

[0111] The crystal structure and phase purity of UiO-66-(COOH)2 and Eu@UiO-66-(COOH)2 were studied using X-ray diffraction patterns, such as... Figure 4 As shown, the diffraction peak positions of UiO-66-(COOH)2 are consistent with the reported topological structures of the UiO-66 series, indicating the successful synthesis of MOF materials with good crystallinity. Strong diffraction peaks appear in the low-angle region, with 2θ peaks at 7.5°, 8.8°, and 11.6°, which are attributed to the typical UiO-66 crystal planes (111), (200), and (220), respectively. In the mid-to-high-angle region (2θ = 15°–35°), a set of relatively broad and low-intensity diffuse peaks (such as those at approximately 18°, 20°, 25°, 28°, and 30°) appear, corresponding to short-range ordered scattering from higher-index crystal planes and pore structures. After doping with Eu³⁺, the diffraction peak positions of Eu@UiO-66-(COOH)2 almost completely overlap with those of UiO-66-(COOH)2 (peak position difference ≤0.1°), with no obvious position shift or new peak generation. The possible reasons are: (a) Eu³⁺ did not enter the Zr-oxo cluster nodes, but was located in the channels or defect sites in the form of ion exchange; (b) the doping amount of Eu³⁺ was low, and the resulting lattice changes were insufficient to be detected by conventional XRD; (c) the flexibility or defects of the coordinating carboxyl groups (–COOH) compensated for the difference in ionic radius.

[0112] Furthermore, the full width at half maximum (FWHM) of the diffraction peaks of UiO-66-(COOH)2 and Eu@UiO-66-(COOH)2 are basically consistent (e.g., the peak width at 8.8° is approximately 0.25°), indicating that Eu 3+Doping did not significantly alter the grain size. The average grain size was estimated to be approximately 40-50 nm according to the Scherrer formula. Throughout the 2θ range, Eu@UiO-66-(COOH)2 did not exhibit any diffraction peaks that could be attributed to Eu2O3 (characteristic peaks approximately 28.5°, 33.1°) or Eu(COOH)3 (characteristic peaks approximately 10.2°, 21.5°), indicating that the introduction of Eu³⁺ did not disrupt the long-range order of the UiO-66-(COOH)2 framework. 3+ It exists in a highly dispersed state in the channels or defect sites of Eu@UiO-66-(COOH)2, without forming a crystalline europium oxide or carboxylate impurity phase.

[0113] (3) Electron microscopy-energy dispersive spectroscopy analysis

[0114] The morphology and elemental composition of UiO-66-(COOH)2 and Eu@UiO-66-(COOH)2 were characterized by field emission scanning electron microscopy combined with energy dispersive spectroscopy. Figure 5 , 6 As shown, both UiO-66-(COOH)2 and Eu@UiO-66-(COOH)2 exhibit a spherical structure with a particle size of approximately 50 nm, which is consistent with the grain size estimated by X-ray diffraction analysis. The elemental chemical distribution and composition of UiO-66-(COOH)2 and Eu@UiO-66-(COOH)2 were analyzed by energy-dispersive X-ray spectroscopy (EDS). The EDS spectra show that UiO-66-(COOH)2 is mainly composed of carbon (C), oxygen (O), and zirconium (Zr), while Eu@UiO-66-(COOH)2 is mainly composed of carbon (C), oxygen (O), zirconium (Zr), and europium (Eu). (Comparison) Figure 5 and Figure 6 Apart from the C, O, and Zr peaks from the UiO-66-(COOH)2 framework, Eu@UiO-66-(COOH)2 exhibited clearly visible Eu characteristic peaks, confirming the presence of Eu. Semi-quantitative analysis showed that the relative atomic content of Eu in Eu@UiO-66-(COOH)2 was 13.44 wt%, and no other impurity elements were detected.

[0115] The morphology of Eu@UiO-66-(COOH)2 was further characterized using transmission electron microscopy. The transmission electron microscopy results are as follows: Figure 7 The results were consistent with those obtained by scanning electron microscopy (SEM), confirming that Eu@UiO-66-(COOH)2 has a spherical structure.

[0116] 2. Fluorescence performance of the fluorescent material Eu@UiO-66-(COOH)2

[0117] (1) Capturing the fluorescence characteristic peaks of Eu-MOF sensing liquid

[0118] Prepare UiO-66-(COOH)2 aqueous solution (hereinafter referred to as "MOF solution") and Eu@UiO-66-(COOH)2 aqueous solution (hereinafter referred to as "Eu-MOF sensing solution") with a concentration of 0.5 mg / mL respectively.

[0119] Take 400 μL of 0.5 mg / mL Eu-MOF sensing solution, add 1600 μL of deionized water, mix well, and measure the fluorescence spectrum of the Eu-MOF sensing solution at an excitation wavelength of 255 nm. Similarly, measure the fluorescence spectrum of the MOF solution. Simultaneously, irradiate with a 254 nm UV lamp and observe the fluorescence emission phenomena of the MOF solution and Eu-MOF sensing solution. Figure 8 As shown.

[0120] At an excitation wavelength of 255 nm, the Eu-MOF sensing fluid exhibits two distinct fluorescence characteristic peaks in the 550 nm-750 nm range, with the peak positions located near 591 nm and 613 nm, respectively. This is attributed to the presence of Eu... 3+ Ionic 5 D0→ 7 F1 5 D0→ 7 The F2 transition confirms the successful synthesis of Eu@UiO-66-(COOH)2. The highest fluorescence emission peak intensity is observed at 613 nm, which is the characteristic emission peak of Eu. 3+ The typical fluorescence emission characteristic peaks are observed. Under 254 nm ultraviolet light irradiation, the MOF solution does not emit red fluorescence, while the Eu-MOF sensing solution emits strong red fluorescence.

[0121] (2) Determine the optimal excitation wavelength of the Eu-MOF sensing fluid

[0122] The fluorescence emission peak intensity of the Eu-MOF sensing solution was investigated at different excitation wavelengths (220 nm to 280 nm) to determine the optimal excitation wavelength. The Eu-MOF sensing solution exhibited red fluorescence emission independent of the excitation wavelength. When the excitation wavelength was 255 nm, the fluorescence emission peak intensity of the Eu-MOF sensing solution was highest near 591 nm and 613 nm; therefore, the optimal excitation wavelength was determined to be 255 nm. Figure 9 As shown.

[0123] (3) Determine the optimal concentration of Eu-MOF sensing fluid.

[0124] A 0.5 mg / mL Eu-MOF sensing solution was prepared, and then diluted to prepare Eu-MOF sensing solutions with concentrations of 10, 20, 50, 80, 100, 120, 150, 200, 250, 300, 350, and 400 μg / mL. Fluorescence scanning was performed at an excitation wavelength of 255 nm, and the intensity changes of the characteristic fluorescence emission peaks at 592 nm and 613 nm were recorded to determine the optimal concentration of the Eu-MOF sensing solution for subsequent experiments. Within the range of 10-400 μg / mL, the fluorescence intensity of the Eu-MOF sensing solution initially increased and then decreased with increasing concentration, with the 100 μg / mL Eu-MOF sensing solution exhibiting the highest fluorescence peak intensity. Figure 10 As shown. Therefore, the concentration of the Eu-MOF sensing solution used in the experiment was 100 μg / mL.

[0125] (4) Specificity test

[0126] To investigate the selectivity and anti-interference ability of the Eu-MOF sensing fluid against MS-222, 13 chemical interfering substances and MS-222 standard solution of equal mass concentration were simultaneously added to the Eu-MOF sensing fluid. After mixing, fluorescence scanning was performed at an excitation wavelength of 255 nm, and the corresponding fluorescence emission spectra were recorded. Figure 13-16 As shown.

[0127] Because some interfering substances are insoluble in water, methanol, which is infinitely miscible with water, was chosen as the solvent to prepare some of the interfering chemicals. These chemicals include NO3. - Cl - SO4 2- BrO3 - Na + K + The following chemicals were used: methanol (JC), furazolidone (AOZ), furazolidone (AMOZ), florfenicol (FBNKA), ribavirin (LBWL), rimantadine (JGYA), and eugenol (DXF). Simultaneously, their fluorescence emission was observed under 254 nm UV light irradiation. The results showed that these chemicals, at the same concentration, did not interfere with the detection of MS-222 by the Eu-MOF sensing liquid. The Eu-MOF sensing liquid of this application exhibits specific selectivity and anti-interference performance for MS-222.

[0128] Application Trial

[0129] (1) Spiked recovery test

[0130] The MS-222 fluorescence sensing method in aquaculture water and fish and shrimp substrates was validated using the standard addition method, with the Eu-MOF sensing solution concentration being 0.5 mg / mL.

[0131] 1) Aquaculture substrate

[0132] Weigh 5g (accurate to 0.01g) of negative aquaculture water samples into 10mL stoppered centrifuge tubes, add 0, 10, 50, and 150μL of 10mg / mL MS-222 standard solution respectively (equivalent to MS-222 concentrations of 0, 76.54, 382.72, and 1148.15μmol / L after spiked), mix well, centrifuge at 8000 rpm for 5 minutes, collect the supernatant, and filter through a 0.22μm aqueous filter membrane.

[0133] ① This fluorescence sensing analysis method: Take 500 μL of filtrate, add 400 μL of Eu-MOF sensing solution, add 1100 μL of water, mix well, and scan the fluorescence under 255 nm excitation light to record the fluorescence intensity I at 613 nm.

[0134] Take 400 μL of Eu-MOF sensing solution, add 1600 μL of water, mix well, and scan under 255 nm excitation light. Record the fluorescence intensity at 613 nm as I0. Substitute the I0 / I ratio into the linear equation I0 / I=1.03321+0.0027×[Q] (R2=0.997) to calculate the MS-222 concentration [Q] in the aquaculture water samples at each spiking level.

[0135] ② Reference method (liquid chromatography-mass spectrometry): Pipette 500 μL of the above filtrate into a centrifuge tube, dilute to 10 mL, mix well, then pipette 10 μL of the sample solution into a sample vial, add 990 μL of 50% acetonitrile solution, mix well, and then perform the test. The MS-222 standard curve for liquid chromatography-mass spectrometry uses concentrations of 5, 10, 20, 50, 100, 200, and 400 ng / mL. The MS-222 concentration in the aquaculture water samples at each spiked level was measured.

[0136] For each addition level, three parallel independent trials were conducted, and the mean and relative deviation were calculated.

[0137] 2) Fish and shrimp substrate

[0138] Weigh four 5g (accurate to 0.01g) negative samples of fish or shrimp and place them in 50mL polypropylene centrifuge tubes. Add 0, 20, 100, and 300 μL of 10mg / mL MS-222 standard solution (equivalent to MS-222 concentrations of 0, 76.54, 382.72, and 1148.15 μmol / L after spiked treatment, respectively). Add 2g MgSO4, 1g NaCl, and 10mL acetonitrile. Vortex for 5 min and centrifuge at 8000rpm for 5 min. Take 5mL of the supernatant and transfer it to a purification tube (containing 500mg MgSO4 and 40mg C). 18Vortex in a solution of 20 mg PSA and 20 mg GCB for 1 min, centrifuge at 8000 rpm for 5 min, and filter 3 mL of the supernatant through a 0.22 μm organic membrane.

[0139] ① This fluorescence sensing analysis method: Take 300 μL of filtrate, add 400 μL of Eu-MOF sensing solution, add 1300 μL of water, mix well, and scan under 255 nm excitation light to record the fluorescence intensity I at 613 nm. Take 400 μL of Eu-MOF sensing solution, add 1600 μL of water, mix well, and scan under 255 nm excitation light to record the fluorescence intensity I0 at 613 nm. Substitute the I0 / I ratio into the linear equation I0 / I=1.03321+0.0027×[Q] (R2=0.997) to calculate the MS-222 concentration in fish or shrimp samples at each spiking level.

[0140] ② Reference method (liquid chromatography-mass spectrometry): Take 500 μL of filtrate into a centrifuge tube, dilute to 10 mL, mix well, then pipette 10 μL of sample solution into a sample vial, add 990 μL of 50% acetonitrile water, mix well, and then perform the test. The MS-222 standard curve for liquid chromatography-mass spectrometry uses concentrations of 5, 10, 20, 50, 100, 200, and 400 ng / mL. The MS-222 concentration in fish or shrimp samples at each spiked level was measured.

[0141] For each addition level, three parallel independent trials were conducted, and the mean and relative deviation were calculated.

[0142] The recoveries of MS-222 in aquaculture water, fish meat, and shrimp meat were 94.8%–106.0%, 89.5%–103.7%, and 87.1%–103.7%, respectively. The relative standard deviations (RSDs) in all three matrices were less than 10%, meeting the requirements of GB 5009.295-2023 "General Rules for Validation of Chemical Analysis Methods in National Food Safety Standards" (see Table 1 for details). Compared with the results of the reference method (liquid chromatography-mass spectrometry), this method can effectively detect the approximate concentration of MS-222, indicating that it is suitable for MS-222 detection.

[0143] Table 1 Spiked Recovery Test

[0144]

[0145] (2) Stability test

[0146] ① Acid-base stability

[0147] The pH stability of the Eu-MOF sensing solution was investigated by monitoring its fluorescence response at different pH values ​​(pH was adjusted by adding 0.05M hydrochloric acid or sodium hydroxide). Within a pH range from 1 to 9, the fluorescence intensity of the Eu-MOF sensing solution at 613 nm initially increased and then decreased with increasing pH. The fluorescence intensity was highest at pH 4. Figure 17 As shown. Since the pH of 100 μg / mEu-MOF sensing solution is approximately 4.3, Eu-MOF sensing solution was used directly in subsequent experiments without pH adjustment. Relatively speaking, Eu-MOF sensing solution exhibits better stability in acidic environments, while alkaline environments affect the fluorescence peak intensity of Eu-MOF sensing solution.

[0148] ② Fluorescence stability

[0149] The fluorescence stability of Eu-MOF sensing fluid in aqueous solution was studied by monitoring the fluorescence intensity at different time points. Figure 18 As shown, the fluorescence intensity of the Eu-MOF sensing liquid did not change significantly within 10 days, indicating that the fluorescence emission of the Eu-MOF sensing liquid has high stability.

[0150] (3) Actual sample application test

[0151] The MS-222 fluorescence sensing method was used to analyze the actual samples.

[0152] MS-222 was extracted and purified from the aquaculture water samples, fish and shrimp samples according to the sample pretreatment method in the spiked recovery test. Concentration or dilution was performed as needed. The MS-222 content was then determined using fluorescence sensing and liquid chromatography-mass spectrometry, respectively. The results are shown in Table 2. Figures 19-20 As shown. The United States, the United Kingdom, Canada and other countries have approved MS-222 for use in edible fish, with a recommended concentration of 5-80 mg / L.

[0153] Table 2 Actual Sample Detection Results

[0154]

[0155] The results show that the detection method of this application is very close to the detection results of liquid chromatography-mass spectrometry, indicating that the detection method of this application has high stability and accuracy and is suitable for the detection of MS-222 in actual field samples.

[0156] Finally, it should be noted that the above embodiments are only used to illustrate the technical solutions of the present invention and are not intended to limit it. Any other modifications or equivalent substitutions made by those skilled in the art to the technical solutions of the present invention, as long as they do not depart from the spirit and scope of the technical solutions of the present invention, should be covered within the scope of the claims of the present invention.

Claims

1. A fluorescent material, characterized in that, The fluorescent material Eu@UiO-66-(COOH)2 is a rare earth Eu-doped functionalized metal-organic framework material UiO-66-(COOH)2 with a spherical structure and a particle size of 40-50 nm.

2. A preparation process for the fluorescent material according to claim 1, characterized in that, It is prepared by the following steps: Take UiO-66-(COOH)2, Eu(NO3)3⋅6H2O and an appropriate amount of deionized water, and react them in a hydrothermal reactor at 80~100℃ and 100~200rpm for 15~24 hours. The weight ratio of Eu(NO3)3⋅6H2O to UiO-66-(COOH)2 is 1:1-5. After cooling to room temperature, the solid product is washed, collected, and dried to obtain the final product.

3. The preparation process according to claim 2, characterized in that, The UiO-66-(COOH)2 is prepared by the following steps: Pyromellitic anhydride and ZrCl4 were added to a hydrothermal reactor containing an appropriate amount of deionized water. The reactor was fixed in an air-heated oven that could rotate around an axis and reacted at 80-100℃ and 100-200rpm for 15-24 hours. After cooling to room temperature, the initial product was obtained by washing. The weight ratio of pyromellitic anhydride to ZrCl4 was 1-2.5:

1. Take the initial product and add it into a hydrothermal reactor. Add 5-20 times the weight of deionized water and react at 80-100℃ and 100-200rpm for 10-20 hours. After cooling to room temperature, clean the product, collect the solid product, and dry it to obtain the final product.

4. The application of the fluorescent material of claim 1 in the rapid detection of the fishing anesthetic MS-222.

5. The application according to claim 4, characterized in that: Using fluorescent materials as the photosensitizers in fluorescence sensing analysis, a standard curve for the fluorescence sensing analysis of the fishing anesthetic MS-222 was plotted, including the following steps: Preparation of standard sensing solution: Weigh an appropriate amount of fluorescent material and add deionized water to prepare a standard fluorescent sensing solution with a concentration of H0. Preparation of MS-222 standard solutions: Add different volumes of MS-222 standard stock solution to the standard fluorescence sensing solution to prepare n MS-222 standard solutions of different concentrations, n≥5, with a concentration range of 0μmol / L-2000μmol / L; Fluorescence scans were performed on MS-222 standard solutions of different concentrations at excitation wavelengths of 240-280 nm, and the fluorescence intensity I of the characteristic fluorescence emission peak at 550-650 nm was recorded; the fluorescence intensity of the MS-222 standard solution with a concentration of 0 was recorded as I0. With the concentration [Q] of the MS-222 standard solution as the X-axis and I0 / I as the Y-axis, a standard curve for MS-222 fluorescence sensing analysis was plotted, and the Stern-Volmer equation was obtained by fitting: I0 / I=1+K[Q].

6. The application according to claim 5, characterized in that, The method for rapid detection of the fishing anesthetic MS-222 includes the following steps: (1) Sampling; pretreatment of the product to be tested to obtain the test solution; (2) Detection; Weigh an appropriate amount of fluorescent material and add deionized water to prepare a standard fluorescent sensing solution with a concentration of H0. The test solution is added to the solution of the fluorescent material, and an appropriate amount of deionized water is added to prepare the fluorescent test solution. The final concentration of the fluorescent material in the fluorescent test solution is H0. At an excitation wavelength of 240-280 nm, the standard fluorescent sensing solution and the fluorescent test solution were subjected to spectral scanning, and the fluorescence intensities at 550-650 nm were recorded as I0 and I, respectively. (3) Substitute I0 and I from step (2) into the Stern-Volmer equation I0 / I=1+K[Q] to obtain the concentration of the test solution.

7. The application according to claim 6, characterized in that, The pretreatment refers to taking a sample after pretreating the product to be tested. The pretreatment method for aquaculture water is as follows: take an appropriate amount of aquaculture water and pass it through a 0.22μm water-based filter membrane to obtain the filtrate as the aquaculture water test solution. The pretreatment method for aquatic products is as follows: the aquatic products are crushed and pulped using a meat grinder, the homogenized sample is weighed into a centrifuge tube, MgSO4, NaCl and acetonitrile are added, vortexed and centrifuged, and the weight ratio of the homogenized sample to MgSO4, NaCl and acetonitrile is 5:1-3:0.5-2:10-15. Take an appropriate amount of supernatant into a purification tube, vortex, centrifuge, and filter through a 0.22μm organic membrane to obtain the filtrate as the test solution for aquatic products.

8. The application according to claim 6, characterized in that, In step (2), H0 is 100 μg / mL.

9. The application according to claim 6, characterized in that, In step (2), the excitation wavelength is 255nm, and the fluorescence emission peak at 613nm is used as the indicator peak.

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