A method for detecting Cu in aqueous solution 2+ Fluorescent probe and preparation method and application thereof

Abstract

The present invention relates to a method for detecting Cu in aqueous solution. 2+ The fluorescent probe and its preparation method and application are as follows, using methyl hydrazinecarboxylate and 7-diethylaminocoumarin-3-formaldehyde as raw materials, and the primary amine on methyl hydrazinecarboxylate and the aldehyde group on 7-diethylaminocoumarin-3-formaldehyde undergo nucleophilic addition reaction to generate a C=N double bond, and then 1 H NMR, 13 C NMR and ESI-MS were used to characterize its structure. The probe showed bright green fluorescence in aqueous solution, and the fluorescence was quenched after adding copper ions. After mixing the copper ion solution with a concentration gradient change with the probe SJL solution, the fluorescence intensity was measured, and then a graph was drawn with the copper ion concentration as the abscissa and the fluorescence intensity of the mixed system as the ordinate to establish a standard line for the relationship between the concentration gradient change of the copper ion solution and the fluorescence intensity change of the fluorescent probe solution. This compound can be used as a highly sensitive and selective method for detecting trace Cu in aqueous solution. 2+ The complex constant of the fluorescent probe is 1.98×10 5 M ‑1 , with a detection limit as low as 0.6 μM.

Description

Technical Field

[0001] The invention discloses a fluorescent probe for detecting Cu2+ in an aqueous solution and a preparation method and application thereof, belonging to the technical field of fluorescent probes for heavy metal ion detection. Background Art

[0002] Fluorescent probes are based on the organic combination of molecular recognition and fluorescence technology. They selectively bind to specific objects through receptors, express molecular binding information with fluorescent signals, and achieve in-situ real-time detection at the molecular level. Fluorescent probes for metal ion detection provide a detection method with wide application value. The photophysical and photochemical issues involved in their design, synthesis, identification and application have attracted extensive attention from researchers in different fields.

[0003] Fluorescent probes have unparalleled advantages over other analytical methods, such as atomic absorption spectroscopy, inductively coupled plasma atomic emission spectroscopy, solid phase microextraction combined with high performance liquid chromatography and electrochemical analysis methods. That is, the presence and concentration range of ions can be directly reflected through changes in fluorescence intensity and color, and they have high sensitivity and selectivity. The recognition mechanisms of common ion recognition fluorescent probes include photoinduced electron transfer (PET), intramolecular charge transfer (ICT), and fluorescence resonance energy transfer (FRET).

[0004] Coumarin derivatives are an important class of natural heterocyclic compounds widely found in nature. In addition to their unique biological activity, these compounds also have excellent optical properties. Their basic structure, benzopyran, is a colorless substance without fluorescence. However, if an electron-donating group is introduced at the 7th position and an electron-withdrawing group is introduced at the 3rd or 4th position, a "push-pull" electron system is formed to obtain a strong fluorescent substance. Coumarin compounds have a large molar absorption coefficient, a high fluorescence quantum yield, a large Stokes shift, and good photostability. They can effectively reduce the background fluorescence of cell components and physiological fluids, and are highly favored in the design of fluorescent probes. The design and synthesis of new coumarin fluorescent compounds are of great significance.

[0005] Copper is a common metal widely present in the environment. 2+ It is beneficial to maintain the normal operation of the body. 2+ Metabolic imbalance can cause diseases. Due to the widespread use of copper in the industrial field, it has caused serious environmental pollution through the discharge of factory wastewater, threatening human life and health. 2+There are endless fluorescent probes for Cu in aqueous solution, but most of them are based on organic solvents, which may cause harm to organisms or environmental pollution during the detection process, limiting their practical application. Therefore, it is necessary to establish a fluorescent probe that has low toxicity, good biocompatibility, high selectivity, high sensitivity and is easy to synthesize. 2+ The fluorescent probe is of great significance. Summary of the invention

[0006] The present invention overcomes the shortcomings of the prior art and provides a fluorescent probe for detecting Cu2+ in aqueous solution and a preparation method and application thereof. The designed fluorescent probe molecule uses methyl hydrazinecarboxylate as an ion receptor and coumarin as a fluorescent response group to synthesize a fluorescent probe that has low toxicity, good biocompatibility, high selectivity, high sensitivity and is easy to synthesize and can detect trace amounts of Cu2+ in pure water. 2+ Fluorescent probe reagents, synthetic methods and applications in trace Cu 2+ Determination of.

[0007] In order to solve the above technical problems, the technical solution adopted by the present invention is: a fluorescent probe for detecting Cu2+ in aqueous solution, using 7-diethylaminocoumarin-3-carboxaldehyde as a carbon source and methyl hydrazinecarboxylate as a nitrogen source to form a Schiff base bond with strong coordination ability. The compound name is 7-diethylamino-2-oxo-2H-chromene-3-methylenehydrazine-1-carboxylic acid methyl ester, and the chemical structural formula of the compound is:

[0008]

[0009] Molecular formula: C 16 H 19 N3O4

[0010] Molecular weight: 317.35

[0011] Melting point: 212.3-217.2℃

[0012] Solubility: Soluble in water, easily soluble in methanol, ethanol, dichloromethane and other organic solvents.

[0013] Spectral properties: The maximum ultraviolet absorption wavelength in aqueous solution is 444nm, and the fluorescence emission wavelength is 515nm.

[0014] The structural characterization is as follows: 1H NMR (DMSO-d6, 400 MHz, TMS): δ (ppm) 11.17 (s, 1H), 8.23 ​​(s, 1H), 8.06 (s, 1H), 7.62 (d, J = 9.2 Hz, 1H), 6.73 (dd, J1 = 2.4 Hz, J2 = 2.4 Hz, 1H), 6.55 (d, J = 2.0 Hz, 1H), 3.69 (s, 3H), 3.45 (q, J = 6.8 Hz, 4H), 1.13 (t, J = 7.2 Hz, 6H). 13 C NMR (DMSO-d6, 600MHz, TMS): δ (ppm): 161.22, 156.78, 154.29, 154.27, 151.58,138.40,131.12,113.24,110.10,108.48,96.86,52.36,44.67,12.82.HRMS(ESI):m / z calcd for C 16 H 20 O4N3[(M+H) + ]:318.1454,found 318.1448;C 16 H 19 O4N3Na[(M+Na) + ]:340.1274,found 340.1268.

[0015] The technical route is as follows:

[0016]

[0017] The preparation method is as follows:

[0018] Synthesis of probe SJL

[0019] The molar ratio of methyl hydrazinecarboxylate and compound 2 was controlled to be 1.0-1.2:1, and anhydrous ethanol was used as the solvent. Refluxed at 85°C for 5h, after complete reaction, cooled to room temperature, solid precipitated, filtered, and the filter cake was washed with ice ethanol to obtain a yellow solid. The yellow solid was separated and purified by silica gel column chromatography using dichloromethane and methanol as eluents to obtain an orange solid SJL.

[0020] Compared with the prior art, the present invention has the following beneficial effects: the present invention establishes a method for identifying Cu in aqueous solution with low toxicity, good biocompatibility, high selectivity, high sensitivity and easy synthesis. 2+ The designed fluorescent probe uses methyl hydrazinecarboxylate as an ion receptor and coumarin as a fluorescent response group to synthesize a fluorescent probe that has low toxicity, good biocompatibility, high selectivity, high sensitivity and is easy to synthesize and can detect trace amounts of Cu in pure water. 2+Fluorescent probe reagents, synthetic methods and applications in trace Cu 2+ Determination of. BRIEF DESCRIPTION OF THE DRAWINGS

[0021] The present invention will be further described below in conjunction with the accompanying drawings.

[0022] Figure 1 is the chemical structure of the fluorescent probe SJL of the present invention;

[0023] Figure 2 is the H NMR spectrum of the fluorescent probe SJL of the present invention;

[0024] Figure 3 is the NMR carbon spectrum of the fluorescent probe SJL of the present invention;

[0025] Figure 4 This is the high-resolution mass spectrum of the fluorescent probe SJL of the present invention;

[0026] Figure 5 The ultraviolet spectrum curves of the fluorescent probe SJL of the present invention in response to different metal cations;

[0027] Figure 6 The fluorescent probe SJL of the present invention is sensitive to different Cu 2+ UV spectroscopy for concentration titration;

[0028] Figure 7 The fluorescence spectra of the fluorescent probe SJL of the present invention in response to different metal cations;

[0029] Figure 8 This is an ion competition experiment of the fluorescent probe SJL of the present invention;

[0030] Fig. 9 The fluorescent probe SJL of the present invention is sensitive to different Cu 2+ Fluorescence spectrum curve of concentration titration;

[0031] Fig.10 The fluorescent probe SJL of the present invention is Cu 2+ The detection limit straight line;

[0032] Fig.11 The fluorescent probe SJL of the present invention is Cu 2+ The complex constant straight line;

[0033] Fig.12 This is the Job's Plot curve of the fluorescent probe SJL of the present invention. DETAILED DESCRIPTION

[0034] like Figures 1 to 12As shown, the present invention is a fluorescent probe for detecting Cu2+ in aqueous solution, the name of the compound is 7-diethylamino-2-oxo-2H-chromene-3-methylenehydrazine-1-carboxylic acid methyl ester, and the chemical structural formula of the compound is:

[0035]

[0036] Molecular formula: C 16 H 19 N3O4

[0037] Molecular weight: 317.35

[0038] Melting point: 212.3-217.2℃

[0039] Solubility: Soluble in water, easily soluble in methanol, ethanol, dichloromethane and other organic solvents

[0040] Spectral properties: The maximum ultraviolet absorption wavelength in aqueous solution is 444nm, and the fluorescence emission wavelength is 515.5nm.

[0041] The fluorescent probe is of coumarin-based Schiff base type.

[0042] A method for preparing a fluorescent probe for detecting Cu2+ in an aqueous solution is implemented according to the following steps:

[0043] Step 1: Synthesize compound 7-diethylaminocoumarin-3-carboxaldehyde (hereinafter referred to as compound 2);

[0044] Step 2: Control the molar ratio of methyl hydrazinecarboxylate and compound II to 1.0-1.2:1, use anhydrous ethanol as solvent; reflux at 85°C for 5h, cool to room temperature, filter out solid precipitation, wash the filter cake with ice ethanol to obtain a yellow solid.

[0045] Step 3: The yellow solid is separated and purified by silica gel column chromatography using dichloromethane and methanol as eluents to obtain an orange solid SJL, which is the fluorescent probe.

[0046] An application of a fluorescent probe for detecting Cu2+ in an aqueous solution, characterized in that the Schiff base compound can be used as a fluorescent probe for detecting Cu2+ in an aqueous solution. 2+ The fluorescent probe is Cu 2+ The concentration of Cu was plotted as the horizontal axis and the fluorescence intensity of the mixed system was plotted as the vertical axis to establish the Cu 2+ The standard line of the change of solution concentration gradient and the change of fluorescence intensity of the probe SJL solution is used to read the Cu in the solution to be tested from the graph according to the fluorescence intensity. 2+ The detection limit was as low as 0.6 μM.

[0047] In order to further illustrate the present invention, the present invention is described in further detail in conjunction with the accompanying drawings and embodiments, so that those skilled in the art can implement the present invention by referring to the description.

[0048] Figure 5 In the figure, the horizontal axis is wavelength and the vertical axis is absorbance. In aqueous solution, the absorbance of the ultraviolet spectrum is 2+ The change of the solution color from yellow-green to brown-yellow indicates that the probe SJL is sensitive to Cu 2+ Have a certain naked eye recognition ability.

[0049] Figure 6 In the pure water system, Cu 2+ The concentrations ranged from 0 to 13.9 times the concentration of the probe SJL. Figure 5 It shows that the absorbance of probe SJL increases with the increase of Cu 2+ The concentration of SJL increased significantly, and the maximum absorption wavelength red-shifted from 444nm to 473nm. Isoconcentration points appeared at 363nm and 443nm, respectively, indicating that the probe SJL and the complex SJL-Cu 2+ In dynamic balance.

[0050] Figure 7 In the pure water system, the horizontal axis is the wavelength and the vertical axis is the fluorescence intensity. Compared with other ions, the probe SJL 2+ Afterwards, the fluorescence intensity decreased significantly, and the quenching rate was as high as 82.3%.

[0051] Figure 8 By comparing the fluorescence intensity of each solution at the same emission wavelength (444 nm), it can be seen that the probe SJL has a strong affinity for Cu 2+ The selective recognition is not affected by other ions.

[0052] Fig. 9 In the 2+ As the concentration of Cu increases, the fluorescence intensity of the probe SJL gradually decreases. 2+ When the concentration increased to 25.9 times, the fluorescence intensity hardly changed. This also shows that the probe SJL has a strong affinity for Cu 2+ Has higher sensitivity.

[0053] Fig.10 In Cu 2+ The fluorescence intensity of the probe SJL was 0.1 to 0.7 times that of Cu 2+ The concentration showed a good linear relationship, and the linear equation was y = -978.57x + 2175.64, R 2 =0.99038. This indicates that within the above concentration range, the probe can detect Cu2+ The concentration can be quantitatively detected. According to the equation, the slope m = -978.57 × 10 5 At the same time, we used the linear relationship and the detection limit calculation formula LOD = 3σ / m to calculate the minimum detection limit of probe SJL to be 0.6μM, indicating that the sensitivity of probe SJL is relatively high.

[0054] Fig.11 The fitted linear equation y = 4.07 × 10 -9 x+8.05×10 -4 , R 2 =0.99241, according to the complex constant calculation formula, the complex constant is Ka = 1.98 × 10 5 M -1 This indicates that the probe SJL and Cu 2+ A complex with good stability was formed.

[0055] Fig.12 In the experiment, the Job curve was determined. When the total concentration remained unchanged, the Cu 2+ With the increase of concentration ratio, the fluorescence intensity of probe SJL at 515nm gradually decreases. 2+ When the concentration of Cu is 0.47 times, the curve has an inflection point, indicating that the probe and Cu 2+ The coordination ratio of the ions is 1:1.

[0056] Synthesis of probe SJL

[0057]

[0058] Weigh 90.08 mg (1 mmol) of methyl hydrazinecarboxylate and add it to a 50 mL round-bottom flask, add 10 mL of anhydrous ethanol, stir at room temperature to completely dissolve it, add 245 mg (1 mmol) of 7-diethylaminocoumarin-3-carboxaldehyde to the reaction bottle, stir at 60 ° C for 30 min, and drop a drop of glacial acetic acid into the reaction bottle. The mixture is heated to 85 ° C and refluxed for 5 h. TLC monitoring, after the raw materials are completely reacted, the mixture is cooled to room temperature, solids are precipitated, filtered, and dried to obtain 240 mg of yellow solids. Use silica gel column chromatography to separate and purify it, use CH2Cl2 / CH3OH (300:1→200:1→100:1, v / v) as eluent, and finally obtain the target product SJL 120 mg, with a yield of 63%.

[0059] The embodiments of the present invention are described in detail above in conjunction with the accompanying drawings, but the present invention is not limited to the above embodiments, and various changes can be made within the knowledge scope of ordinary technicians in this field without departing from the purpose of the present invention.

Claims

1. A fluorescent probe for detecting Cu2+ in aqueous solution, characterized in that: The name of the compound is 7-diethylamino-2-oxo-2H-chromene-3-methylenehydrazine-1-carboxylic acid methyl ester, and the chemical structural formula of the compound is: ， It is denoted as probe SJL; Molecular formula: C 16 H 19 N3O4 Molecular weight: 317.35 Melting point: 212.3-217.2℃ Solubility: Soluble in water, easily soluble in organic solvents including methanol, ethanol, and dichloromethane Spectral properties: The maximum ultraviolet absorption wavelength in aqueous solution is 444 nm, and the fluorescence emission wavelength is 515.5 nm.

2. A fluorescent probe for detecting Cu2+ in aqueous solution according to claim 1, characterized in that: The fluorescent probe is a coumarin-based Schiff base compound.

3. A method for preparing a fluorescent probe for detecting Cu2+ in aqueous solution according to claim 1, characterized in that: Follow the steps below to implement: Step 1: Synthesize compound 7-diethylaminocoumarin-3-carboxaldehyde, hereinafter referred to as compound 2; Step 2: Control the molar ratio of methyl hydrazinecarboxylate and compound II to 1.0-1.2:1, use anhydrous ethanol as solvent; reflux at 85°C for 5h, cool to room temperature, filter solid precipitation, wash the filter cake with ice ethanol to obtain a yellow solid; Step 3: The yellow solid is separated and purified by silica gel column chromatography using dichloromethane and methanol as eluents to obtain an orange solid SJL, which is the fluorescent probe.

4. A use of a fluorescent probe for detecting Cu2+ in aqueous solution according to claim 2, characterized in that: The coumarin-based Schiff base compound can be used as a detector for Cu in aqueous solution. 2+ The fluorescent probe is Cu 2+ The concentration of Cu was plotted as the horizontal axis and the fluorescence intensity of the mixed system was plotted as the vertical axis to establish the Cu 2+ The standard line of the change of solution concentration gradient and the change of fluorescence intensity of the probe SJL solution is used to read the Cu in the solution to be tested from the graph according to the fluorescence intensity. 2+ The detection limit was as low as 0.6 μM.

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