Preparation and application of fluorescent hydrogel P (MAA-MMA-CAMA) based on coumarin acylhydrazone derivative

Abstract

A hydrophobic fluorescent monomer CAMA is designed based on a coumarin acylhydrazone derivative CA, the hydrophobic fluorescent monomer CAMA is introduced into a poly (methacrylic acid-methyl methacrylate) network through chemical polymerization, and the fluorescent hydrogel P (MAA-MMA-CAMA) is obtained through solvent exchange. The fluorescent hydrogel P (MAA-MMA-CAMA) not only has excellent optical properties, can realize specific recognition and quantitative detection of Cu < 2 + >, but also has good mechanical properties and anti-swelling properties, overcomes the defects of poor portability, limited application scenarios, fluorescent molecule leakage caused by physical doping and the like of a traditional solution phase probe, and has good application prospects. And the method has higher stability and high efficiency in practical application.

Description

Technical Field

[0001] The present invention belongs to the field of analytical chemistry, and particularly relates to the preparation and application of a fluorescent hydrogel P(MAA-MMA-CAMA) based on a coumarin acylhydrazone derivative. Background Art

[0002] Copper ions are the third most abundant transition metals in the human body and are also one of the indispensable trace elements. They combine with proteins to form ceruloplasmin and various enzymes, playing an important role in physiological processes such as energy generation, iron metabolism, and nerve conduction. However, excessive intake of Cu 2+ can cause liver damage, kidney dysfunction, and various neurodegenerative diseases, such as amyotrophic lateral sclerosis, Parkinson's disease, Alzheimer's disease, and prion disease. The main sources of copper ions in the human body include food intake and drinking water. According to the "Dietary Reference Intakes for Chinese Residents", the recommended daily intake of copper for adults is 0.8 mg, and the tolerable upper intake level is 8 mg. The World Health Organization (WHO) has set the allowable limit for Cu 2+ content in drinking water at 30 μM. However, with the development of agriculture, the extensive use of some copper-based fungicides such as Bordeaux mixture may lead to the accumulation of copper in the soil and then penetrate into groundwater, polluting the water source. Crops sprayed with Bordeaux mixture have copper residues, and long-term consumption may lead to excessive copper intake and cause health problems. Therefore, it is of great significance to research and develop sensitive and efficient technologies for detecting Cu 2+ in crops and drinking water.

[0003] Compared with traditional analytical methods (such as atomic absorption spectroscopy, inductively coupled plasma mass spectrometry, electrochemical analysis, surface plasmon resonance, surface-enhanced Raman spectroscopy), the fluorescence probe technology shows broad application prospects in the rapid detection of metal ions due to its high sensitivity, good selectivity, low detection cost, simple operation, small biological damage, and real-time dynamic monitoring. In practical applications, solid-state fluorescence sensing materials have more advantages in terms of portability, ease of operation, and applicability compared to small molecule fluorescence probes based on the solution phase. As a three-dimensional cross-linked polymer network material, hydrogel has high water content, good biocompatibility, and adjustable physical and chemical properties, providing an ideal carrier platform for fluorescence probes.

[0004] This patent designs a hydrophobic fluorescent monomer CAMA based on a coumarin acylhydrazone derivative CA, introduces it into the poly(methacrylic acid-methyl methacrylate) network through chemical polymerization, and obtains a fluorescent hydrogel P(MAA-MMA-CAMA) through solvent exchange. The performance of the fluorescent hydrogel probe is studied through a series of tests, and its selectivity and sensitivity to Cu 2+It has the advantages of high selectivity and high sensitivity in quantitative detection, strong anti-interference ability and more intuitive and convenient visual detection effect. At the same time, the introduction of hydrophobic fluorescent probes also significantly enhances the mechanical properties and anti-swelling properties of the hydrogel, greatly expanding its practical application value in the fields of environmental monitoring, food safety, etc. Summary of the Invention

[0005] The object of the present invention is to provide a preparation of a fluorescent hydrogel based on coumarin acylhydrazone derivative P(MAA-MMA-CAMA) and its application in the detection of Cu 2+ detection.

[0006] The technical solution to achieve the object of the present invention is as follows:

[0007] The preparation and application of a fluorescent hydrogel P(MAA-MMA-CAMA) based on coumarin acylhydrazone derivative, and the structure of this material is as follows:

[0008]

[0009] The preparation method of a fluorescent hydrogel based on coumarin acylhydrazone derivative in the present invention includes the following steps:

[0010] Dissolve coumarin acylhydrazone derivative CA (0.76 g) and 0.4 ml of triethylamine in 10 ml of N-methylpyrrolidone, heat and stir at 50 °C for 9 hours under N2 atmosphere. After the reaction, pour the mixture into pure water to obtain a precipitate, filter, wash and dry to obtain an orange fluorescent monomer CAMA; dissolve 232 mg of methacrylic acid, 30 mg of methyl methacrylate, 4.6 mg of fluorescent monomer CAMA and 12 mg of crosslinking agent polyethylene glycol diacrylate PEGDA in 1 ml of dimethyl sulfoxide, then add 3 mg of AIBN and mix evenly to obtain a precursor solution. Place it at 65 °C for polymerization for 6 h to obtain an organic gel; wash the unreacted monomers in the gel with DMSO and then soak the organic gel in a large amount of pure water for 48 h, changing the water every 6 h, and obtain a fluorescent hydrogel P(MAA-MMA-CAMA) based on coumarin acylhydrazone derivative through solvent exchange.

[0011] The fluorescent hydrogel based on coumarin acylhydrazone derivative described in the present invention can be used to detect Cu in the aqueous phase 2 + .

[0012] Compared with the prior art, the significant advantages of the present invention are: (1) The present invention synthesizes a fluorescent hydrogel based on coumarin acylhydrazone derivative, which has high sensitivity to Cu 2+It has the advantages of high selectivity and high sensitivity in quantitative detection, strong anti-interference ability and more intuitive and convenient visual detection effect. (2) The raw materials selected in the present invention have low cost, the synthesis method is simple, and the reaction conditions are mild. (3) By introducing a hydrophobic group, the mechanical properties and anti-swelling properties of the hydrogel are significantly enhanced, expanding its practical application value in the fields of environmental monitoring, food safety, etc. BRIEF DESCRIPTION OF THE DRAWINGS

[0013] Figure 1 Based on the coumarin acylhydrazone derivative CA 1 1H NMR spectrum (CDCl3, 400 MHz)

[0014] Figure 2 Based on the coumarin acylhydrazone derivative CA 13 13C NMR spectrum (CDCl3, 101 MHz)

[0015] Figure 3 For the fluorescent monomer CAMA 1 1H NMR spectrum (CDCl3, 400 MHz)

[0016] Figure 4 For the fluorescent monomer CAMA 13 13C NMR spectrum (CDCl3, 101 MHz)

[0017] Figure 5 Infrared spectra of CAMA, P(MAA-MMA) and P(MAA-MMA-CAMA)

[0018] Figure 6 (A) Fluorescence spectra of the hydrogel P(MAA-MMA-CAMA) after being soaked in different metal ions. (B) Selectivity and competition experiments of the hydrogel P(MAA-MMA-CAMA) with different metal ions, and the column height represents the fluorescence intensity at 525 nm. (C) Images of the hydrogel P(MAA-MMA-CAMA) under ultraviolet light and natural light after being soaked in different metal ions.

[0019] Figure 7 For the hydrogel P(MAA-MMA-CAMA) after being soaked in different concentrations of Cu 2+ aqueous solution: (A) Fluorescence spectra; (B) Linear fitting curve of the fluorescence intensity at a wavelength of 525 nm vs. the concentration of Cu 2+ ; (C) Benesi-Hildebrand fitting curve for Cu 2+ ; (D) Images under ultraviolet light and natural light.

[0020] Figure 8(A) Swelling rate curves of three hydrogel samples in water as a function of time. (B) Stress-strain curves of the hydrogel samples before and after swelling (the solid line represents before swelling, and the dashed line represents after 7 days of swelling)

[0021] Figure 9 Determination of Cu in actual samples using the fluorescent hydrogel P(MAA-MMA-CAMA) 2+ Concentration Detailed implementation manners

[0022] Example 1 Preparation and structural characterization of a fluorescent hydrogel P(MAA-MMA-CAMA) based on coumarin acylhydrazone derivatives

[0023] Example 2 Detection ability of the fluorescent hydrogel P(MAA-MMA-CAMA) for Cu 2+ of

[0024] Example 3 Mechanical properties and anti-swelling properties of the fluorescent hydrogel P(MAA-MMA-CAMA)

[0025] Example 4 Application of the fluorescent hydrogel P(MAA-MMA-CAMA) in actual samples

[0026] The present invention will be further described in detail below with reference to the accompanying drawings and specific examples.

[0027] Example 1

[0028] Preparation and structural characterization of a fluorescent hydrogel P(MAA-MMA-CAMA) based on coumarin acylhydrazone derivatives

[0029] Dissolve coumarin acylhydrazone derivative CA (0.76 g) and 0.4 ml of triethylamine in 10 ml of N-methylpyrrolidone, heat and stir at 50 °C for 9 hours under a N2 atmosphere. After the reaction, pour the mixture into pure water to obtain a precipitate, which is filtered, washed, and dried to obtain an orange fluorescent monomer CAMA; dissolve 232 mg of methacrylic acid, 30 mg of methyl methacrylate, 4.6 mg of fluorescent monomer CAMA, and 12 mg of crosslinking agent polyethylene glycol diacrylate PEGDA in 1 ml of dimethyl sulfoxide, then add 3 mg of AIBN and mix evenly to obtain a precursor solution. Place it at 65 °C for polymerization for 6 h to obtain an organic gel; wash the unreacted monomers in the gel with DMSO and then soak the organic gel in a large amount of pure water for 48 h, changing the water every 6 h. A fluorescent hydrogel P(MAA-MMA-CAMA) based on coumarin acylhydrazone derivatives is obtained through solvent exchange; the structures and all synthetic routes of compound CA, fluorescent monomer CAMA, and fluorescent hydrogel P(MAA-MMA-CAMA) are shown in the following formula:

[0030]

[0031] Based on the coumarin acylhydrazone derivative CA 1 H NMR spectra and 13 C NMR spectra are shown in the attached specification Figure 1 , 2; for the fluorescent monomer CAMA 1 H NMR spectra and 13 C NMR spectra are shown in the attached specification Figure 3 , 4.

[0032] The chemical compositions and structures of the fluorescent monomer CAMA, P(MAA-MMA) hydrogel, and P(MAA-MMA-CAMA) fluorescent hydrogel were characterized by Fourier transform infrared spectroscopy (FT-IR). Strong absorption peaks (shoulder peaks) in the range of 1600 - 1700 cm -1 are attributed to the stretching vibration of C=O in MAA and MMA, and the broad peak at 3350 cm -1 is due to the O-H stretching vibration. Compared with the P(MAA-MMA) hydrogel sample, two new characteristic peaks appeared in the P(MAA-MMA-CAMA) fluorescent hydrogel. The absorption peak at 1510 cm -1 is the bending vibration peak of the N-H bond in the acylhydrazone structure of the fluorescent probe, and the absorption peak at 1346 cm -1 is the stretching vibration peak of the C-N bond in the acylhydrazone structure. The appearance of these peaks can prove that the fluorescent monomer CAMA was successfully polymerized into the hydrogel system. The relevant infrared spectra are shown in the attached specification Figure 5 .

[0033] Example 2

[0034] Detection ability of the fluorescent hydrogel P(MAA-MMA-CAMA) for Cu 2+ To verify that the fluorescent hydrogel P(MAA-MMA-CAMA) has excellent selectivity and anti-interference for the recognition of Cu

[0035] 2+ , the hydrogel sensor was immersed in solutions containing different metal ions (Na + , K + , Li + , Ca 2+ , Zn 2+ , Mg 2+- , Al 3+ , Fe 2+ , Fe 3+ , Cu 2 + , Mn 2+ , Cr 3+ , Cd 2+ , Co 2+ , Hg​2+ , Ag + , Pb 2+ ) aqueous solution, after standing for a period of time, the hydrogel sample was tested by fluorescence spectroscopy. As shown in the appendix Figure 6 , it can be seen that only the fluorescence of the hydrogel sample soaked in Cu 2+ solution was significantly quenched, and other samples showed strong green fluorescence. And after being soaked in Cu 2+ solution under natural light, the hydrogel sample showed a visible color change from yellow to brown. Subsequently, Cu 2+ was added to the solutions containing different metal ions, and the hydrogel sample was soaked again. After a period of time, the fluorescence intensity of the hydrogel at 425 nm was recorded and plotted as a bar chart. It can be seen that the presence of other metal interfering ions does not affect the specific recognition of Cu 2+ by the hydrogel sensor.

[0036] The detection limit and binding constant are important indicators for evaluating the performance of fluorescence sensors. Therefore, the fluorescent hydrogel P(MAA-MMA-CAMA) sample was soaked in Cu 2+ solutions with different concentrations (0 - 200 μM) for 2 h, and then its fluorescence intensity was recorded. As the concentration of Cu 2+ increased, the fluorescence intensity of the hydrogel gradually decreased, and the color gradually deepened under visible light. As Figure 7 shows, after the hydrogel sample was treated with Cu 2+ solutions with concentrations of 0 - 100 μM, the fluorescence intensity at a wavelength of 525 nm showed a significant linear relationship with the ion concentration, and the linear equation was y = -1095.2191x + 194788.8864 (R 2 = 0.9973). From formula 1 and formula 2, the detection limit of the probe can be obtained as where S is the slope of the linear fitting straight line equation, and SD is the standard deviation of the fluorescence intensity ratio at 550 nm and 650 nm in the fluorescence spectra of the DQC solution measured ten times. It can be calculated that the detection limit of the fluorescent hydrogel P(MAA-MMA-CAMA) for Cu 2+ is 2.53 μM. The images under ultraviolet light and natural light show that the green fluorescence of the fluorescent hydrogel gradually weakened and the color gradually deepened after being treated with different concentrations of Cu 2+ . This visual detection ability makes it more intuitive and convenient in practical applications, especially having significant advantages in on-site rapid detection and qualitative analysis.

[0037] Example 3

[0038] Mechanical properties and anti-swelling properties of the fluorescent hydrogel P(MAA-MMA-CAMA)

[0039] As the most basic mechanical property test, the tensile stress-strain experiment can explore the tensile strength and elongation at break of the hydrogel. In an environment with constant temperature and humidity, a CMT4204 electronic universal testing machine (SANS Co., Ltd., China) was used to conduct a uniaxial tensile test on the hydrogel sample. The rectangular hydrogel sample had a thickness of 1 mm, a width of 10 mm, and a length of 30 mm. The gauge length was set to 18 mm, and the tensile speed was 20 mm / min. The stress-strain curve was recorded, and each sample was tested 3 times, and the average value was taken. Attached Figure 8 (B) The solid line in shows the tensile stress-strain curve of the hydrogel sample before swelling. It can be seen that the mechanical properties of the fluorescent hydrogel with the introduction of the fluorescent monomer CAMA are significantly improved. The tensile strength increases from 0.32 MPa to 0.68 MPa, and the elongation at break increases from 175% to 275%. This is because the introduction of hydrophobic groups in the fluorescent probe forms hydrophobic microdomains, and the aggregation of hydrophobic groups can form physical cross-linking points, which act together with the chemical cross-linking network to enhance the overall structural stability of the hydrogel while improving the energy dissipation mechanism, thereby improving its mechanical properties. After soaking in Cu 2+ aqueous solution, the mechanical properties of the fluorescent hydrogel are further improved to reach 1.05 MPa, and the elongation at break reaches 285%. Due to the coordination effect of metal ions, the cross-linking point density of the hydrogel can be increased, forming a more compact network structure, thereby improving its rigidity and strength.

[0040] The swelling performance of the hydrogel was measured by the mass analysis method. In an environment with constant temperature and humidity, the hydrogel samples swollen for different times were taken out, and the excess water on the surface of the samples was removed and then weighed. W x represents the mass of the hydrogel after different swelling times, and W0 represents the mass of the hydrogel after 48 h of solvent exchange. The obtained results are the average values of three measurements for each sample. The swelling ratio calculation formula:

[0041] Most hydrogels will inevitably expand in volume in an aqueous environment, the internal structure is damaged, accompanied by mechanical weakening, and the service life is shortened. During the solvent exchange process, water continuously diffuses into the solution, promoting gelation. The hydrogel can effectively capture water through interfacial hydrogen bonds. After 48 hours of solvent exchange, although DMSO has been completely removed, the absolute swelling equilibrium of the hydrogel in water has not been reached.

[0042] The swelling behavior of the hydrogel material was studied by the mass analysis method, and the mechanical properties of three hydrogel samples after swelling for 7 days were tested by the tensile test. By Figure 8(A) As can be seen from the blue dotted line graph, the swelling ratio of the hydrogel sample without the addition of fluorescent monomers during polymerization continuously increases with the increase of swelling time, and tends to be stable after swelling for 48 h. The final swelling ratio is about 32%, and the mechanical properties decrease sharply. The green dotted line graph represents the change of the swelling ratio of the fluorescent hydrogel in water. It increases slowly with the increase of time, and the final swelling ratio reaches 7.5%, which is significantly lower than that of the blank hydrogel sample, and the mechanical properties remain good. This shows that after introducing fluorescent monomers into the hydrogel system, the anti-swelling performance is significantly enhanced. The green dotted line graph represents the fluorescent hydrogel soaked in Cu 2+ aqueous solution. After soaking in water for 7 days, the swelling ratio is less than 3% and the tensile properties remain almost unchanged. Therefore, the fluorescent hydrogel exhibits excellent anti-swelling performance, can maintain structural stability and functional persistence in complex environments, and improve the use efficiency and safety of materials.

[0043] Example 4

[0044] Application of Fluorescent Hydrogel P(MAA-MMA-CAMA) in Actual Samples

[0045] To evaluate the practicality of the designed fluorescent probe in actual samples, it was applied to the detection of Cu 2+ in drinking water and vegetables. The drinking water was taken from purified drinking water, filtered through a 0.22 μm microporous membrane, and used after adjusting the pH. Lettuce was purchased from the local market, washed, crushed, ground into a pulp, added with deionized water, ultrasonically extracted for 30 min, centrifuged, and the supernatant was taken, filtered through a microporous membrane, and used after adjusting the pH.

[0046] The main sources of copper ion intake in the human body are drinking water and food intake. The residues of copper pipes and copper-based pesticides may lead to the over-standard of Cu 2+ in drinking water and vegetables. Developing efficient probes for the detection of copper ions in drinking water and food has important value. To evaluate the practicality of the P(MAA-MMA-CAMA) hydrogel material, the Cu 2+ concentrations in the drinking water samples of Nanjing Forestry University and common vegetable samples (lettuce and leeks) were detected by the standard addition recovery test.

[0047] 20 μM, 40 μM, and 60 μM Cu 2+ solutions were added to the drinking water and vegetable samples respectively. The fluorescent hydrogel samples were soaked in them, and the fluorescence intensity of each group of hydrogels was measured. Each group of samples was measured three times (n = 3), and the average value and standard deviation were obtained. Based on the regression equation y = -1095.2191x + 194788.8864, the samples were analyzed, and the results are as shown in the appendix Figure 9 as shown, Cu 2+The spiked recovery rate is in the range of 95.61% to 103.11%, and the relative standard deviation is in the range of 1.24% to 4.16%. This indicates that the fluorescent hydrogel P(MAA-MMA-CAMA) is expected to become an efficient and reliable fluorescent detection tool for detecting trace amounts of Cu in water and vegetables 2+ .

Claims

1. Preparation and application of a fluorescent hydrogel P(MAA-MMA-CAMA) based on coumarin acylhydrazone derivatives, the structure of the material is as follows:

2. A preparation method of a fluorescent hydrogel P(MAA-MMA-CAMA) based on coumarin acylhydrazone derivatives as described in claim 1, comprising the following steps: 0.76 g of coumarin acylhydrazone derivative CA and 0.4 ml of triethylamine were fully dissolved in 10 ml of N-methylpyrrolidone, heated and stirred at 50 °C for 9 hours under a N2 atmosphere. After the reaction, the mixture was poured into pure water to obtain a precipitate, which was filtered, washed, and dried to obtain an orange fluorescent monomer CAMA; 232 mg of methacrylic acid, 30 mg of methyl methacrylate, 4.6 mg of fluorescent monomer CAMA, and 12 mg of cross-linking agent polyethylene glycol diacrylate PEGDA were dissolved in 1 ml of dimethyl sulfoxide, and then 3 mg of AIBN was added and mixed evenly to obtain a precursor solution. After polymerization at 65 °C for 6 h, an organogel was obtained; the unreacted monomers in the gel were washed away with DMSO, and then the organogel was soaked in a large amount of pure water for 48 h, and the water was changed every 6 h to obtain a fluorescent hydrogel P(MAA-MMA-CAMA) based on coumarin acylhydrazone derivatives as described in claim 1; Among them, the structures and all synthetic routes of compound CA, fluorescent monomer CAMA, and fluorescent hydrogel P(MAA-MMA-CAMA) are shown in the following formula:

3. Use of the fluorescent hydrogel P(MAA-MMA-CAMA) based on coumarin acylhydrazone derivatives as described in claim 1 for non-diagnostic purposes, characterized in that: A fluorescent hydrogel P(MAA-MMA-CAMA) based on coumarin acylhydrazone derivatives as described in claim 1 is used to detect Cu2+ in the aqueous phase.

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