Multifunctional fluorescent probe responding to sulfur dioxide and polarity

Abstract

The invention discloses a multifunctional fluorescent probe based on a coumarin derivative and a preparation method of the multifunctional fluorescent probe. The probe can efficiently detect sulfur dioxide (SO2) and respond to the change of environmental polarity. A double bond and benzo [B] thiophene-3 (2H)-ketone 1, 1-dioxide are introduced into a coumarin molecule to serve as SO2 and a polarity sensitive group respectively, so that the coumarin shows excellent fluorescence performance in different polarity environments. According to the probe, rapid and real-time detection of SO2 and polarity is realized under the wavelength of 480 nm and 580 nm respectively. The method has the remarkable advantages of short response time, high sensitivity, good selectivity, strong specificity and the like. Through the probe, the concentration of SO2 in the environment and the change of polarity of the SO2 can be conveniently monitored, and the probe has important application value. In conclusion, the multifunctional fluorescent probe disclosed by the invention opens up a new direction for research and application in related fields while providing real-time monitoring, and has wide market potential and application prospect.

Description

Technical Field

[0001] The present invention belongs to the technical field of organic small molecule fluorescent probes. Specifically, the present invention relates to a coumarin-based multifunctional fluorescent probe for simultaneously detecting sulfur dioxide (SO 2 ), and polarity, and a preparation method thereof. Background Art

[0002] Sulfur dioxide (SO 2 ), is an important environmental pollutant, mainly derived from activities such as burning fossil fuels, industrial emissions, and transportation. The increase in its concentration in the air not only has a serious impact on the ecological environment but also poses a threat to human health, potentially leading to respiratory diseases and cardiovascular diseases. Therefore, developing sensitive, rapid, and selective SO 2 detection methods is particularly important. Most traditional SO 2 detection methods rely on physical and chemical analysis techniques such as gas chromatography and mass spectrometry. Although they have high sensitivity, they usually require complex equipment and cumbersome sample pretreatment, and are not suitable for on-site rapid detection. As an emerging detection technology, fluorescent probes have gradually become a research hotspot in the field of environmental monitoring due to their high sensitivity, good selectivity, and simple operation.

[0003] In a physiological environment, the polarity of a solution can significantly affect the behavior and reactions of molecules. Polarity refers to the phenomenon of separation of positive and negative charges caused by uneven charge distribution in a molecule, and this property plays an important role in the interactions between biomolecules. Many physiological processes, such as enzymatic reactions, signal transduction, and material transport, are affected by the environmental polarity. Understanding and monitoring the polarity changes in the physiological environment are crucial for studying biochemical reactions and their roles in health and disease.

[0004] Developing a fluorescent probe that can simultaneously respond to SO 2 and the polarity of the physiological environment is of great significance. With the intensification of environmental pollution, the monitoring of SO 2 becomes increasingly important, and the polarity changes in the physiological environment may also affect the biological effects of SO 2 . Therefore, a probe that can monitor both in real time will help to more comprehensively evaluate the impact of the environment on organisms. In recent years, the research on coumarin-based fluorescent probes for detecting SO 2 has gradually increased. These probes usually rely on chemical reactions or physical interactions, resulting in significant changes in fluorescence intensity. However, existing probes often have problems such as slow reaction rate, poor selectivity, and background signal interference when responding to SO 2 , which limits their effectiveness and reliability in practical applications. Therefore, there is an urgent need to develop new multifunctional fluorescent probes to improve the detection of SO 2The sensitivity and selectivity, and simultaneously respond to the polarity changes in the physiological environment. SUMMARY OF THE INVENTION

[0005] The present invention aims to design a multifunctional fluorescent probe based on coumarin, which can not only efficiently detect SO 2 while simultaneously responding to changes in polarity. By introducing specific functional groups, the fluorescence properties of coumarin can be optimized to exhibit excellent fluorescence properties in a polar environment. Specifically, the present invention enhances its selective reaction to SO 2 by introducing a double bond and benzo[B]thiophen-3(2H)-one 1,1-dioxide into the coumarin molecule. At the same time, the introduction of this group improves the sensitivity of the probe to changes in the polarity of the solution, enabling effective response to environmental polarity. This research direction will show broad application prospects in the fields of environmental monitoring, public health, and biomedicine.

[0006] In a first aspect, the present invention provides a multifunctional fluorescent probe COU based on a coumarin derivative, and its structural formula is as follows: .

[0007] The synthesis route of the above-mentioned multifunctional fluorescent probe COU based on a coumarin derivative is: In a second aspect, the present invention provides a preparation method of the multifunctional fluorescent probe COU based on a coumarin derivative, including the following steps: Stir and reflux 7-(diethylamino)coumarin-3-carbaldehyde (Compound 1), benzo[B]thiophen-3(2H)-one 1,1-dioxide (Compound 2), and piperidine in an anhydrous ethanol solution at 80 °C. After cooling, remove ethanol and piperidine, and purify the crude product by column chromatography to obtain a blue-black solid, which is the fluorescent probe.

[0008] In some embodiments, the molar ratio of 7-(diethylamino)coumarin-3-carbaldehyde to benzo[B]thiophen-3(2H)-one 1,1-dioxide is 1:(1.2 - 2).

[0009] In some embodiments, the feeding amount of piperidine relative to the mass of 7-(diethylamino)coumarin-3-carbaldehyde is 2% - 5%.

[0010] In some embodiments, the reactants are stirred and refluxed for 6 - 24 hours, preferably 12 hours.

[0011] In some embodiments, the eluent used for column chromatography purification is dichloromethane and methanol, and the volume ratio is (100 - 20):1, preferably 50:1.

[0012] In some embodiments, the yield of the fluorescent probe COU is 50% - 70%.

[0013] In a third aspect, the present invention provides the use of the multifunctional fluorescent probe COU based on coumarin derivatives in detecting sulfur dioxide (SO 2 ) and the polarity of the physiological environment.

[0014] In some embodiments, the multifunctional fluorescent probe COU based on coumarin derivatives has fluorescence emission wavelengths of 480 nm and 580 nm for SO 2 and the physiological environment polarity respectively, and there is no phenomenon of mutual crosstalk.

[0015] In some embodiments, the lowest detection limit of the multifunctional fluorescent probe COU based on coumarin derivatives for SO 2 is 40.2 nM.

[0016] The present invention has the following beneficial effects: (1) The multifunctional fluorescent probe of the present invention has a unique design, combines the dual response capabilities to SO 2 and the polarity of the physiological environment, has a simple synthesis method, and is easy to separate and purify; the probe can achieve high - selective recognition of SO 2 , effectively reducing the influence of common interfering substances, thus ensuring the accuracy of the measurement results; the probe has extremely high sensitivity to SO 2 , with a lowest detection limit of up to 40.2 nM, a fast response time, and can reach the response in only 1 minute, which can meet the monitoring requirements in practical applications; at the same time, the probe can monitor the change of the polarity of the solution, providing more comprehensive information for evaluating the impact of the environment on organisms.

[0017] (2) The mechanism of the multifunctional fluorescent probe COU of the present invention for recognizing SO 2 and polarity is as Figure 1 shown.

[0018] The coumarin - based fluorophores selected in the present invention have the advantages of convenient synthesis, easy structural adjustment, high fluorescence quantum yield, large Stokes shift, etc. Therefore, they are widely used as fluorescent staining agents and small - molecule fluorescent probes. According to the literature report, SO 2 is prone to Michael addition reaction with unsaturated double bonds, destroying the conjugate mechanism of the probe. Therefore, the present invention selects coumarin as the fluorophore, and through the aldol condensation reaction, a group with a strong electron - withdrawing effect is introduced into coumarin to enhance the sensitivity of the double bond to SO 2 . The probe COU itself does not have fluorescence emission. When it reacts with SO 2After the response occurs, its fluorescence at 480 nm increases rapidly. In addition, when the probe is in a highly polar environment, its fluorescence at 580 nm is very weak, while when in a low-polarity solvent, the fluorescence at 580 nm is significantly enhanced. Therefore, as a multifunctional fluorescent probe, probe COU can achieve the differential detection of SO 2 and polarity simultaneously.

[0019] (3)The multifunctional fluorescent probe of the present invention has significant advantages in responding to SO 2 and the polarity change of the physiological environment. The fluorescence emission wavelengths of this probe are located at 480 nm and 580 nm, without mutual crosstalk. It has a large Stokes shift, a fast response time, strong selectivity and high specificity, making it have fast and efficient performance in the detection of SO 2 and polarity. Therefore, the present invention is a non-invasive probe that can achieve in-situ and real-time monitoring of SO 2 in vivo, and at the same time evaluate the polarity change of the physiological environment, with broad application potential. In the fields of environmental monitoring and biomedicine, such multifunctional probes show good application prospects and provide a powerful detection tool for research and practice. BRIEF DESCRIPTION OF THE DRAWINGS

[0020] In order to more clearly illustrate the technical solutions in the embodiments of the present invention or the prior art, the following will briefly introduce the drawings required for the description of the embodiments or the prior art. Obviously, the following drawings are only some embodiments of the present invention. For those of ordinary skill in the art, without creative efforts, other drawings can also be obtained based on these drawings.

[0021] Figure 1 Shows the mechanism of the multifunctional fluorescent probe COU of the present invention for recognizing SO 2 and polarity; Figure 2 Is the proton nuclear magnetic resonance spectrum of the fluorescent probe COU of the present invention (the solvent is CDCl 3 ); Figure 3 Is the carbon nuclear magnetic resonance spectrum of the fluorescent probe COU of the present invention (the solvent is CDCl 3 ); Figure 4 Is the high-resolution mass spectrum of the fluorescent probe COU of the present invention; Figure 5 Is the fluorescence titration diagram of the fluorescent probe COU of the present invention for recognizing SO 2 , with an excitation wavelength of 420 nm; Figure 6 Is the fluorescence titration diagram of the fluorescent probe COU of the present invention for recognizing SO 2The lowest detection limit graph, excitation wavelength 420 nm, emission wavelength 480 nm; Figure 7 The fluorescence probe COU of the present invention recognizes SO 2 The fluorescence selectivity graph of, excitation wavelength 420 nm; Figure 8 The fluorescence probe COU of the present invention recognizes SO 2 The kinetic curve graph, excitation wavelength 420 nm, emission wavelength 480 nm; Figure 9 The fluorescence emission curve graph of the fluorescence probe COU of the present invention in different polar solvents, excitation wavelength 530 nm; Figure 10 The fluorescence emission curve graph of the fluorescence probe COU of the present invention in a dioxane / water mixed solution with different ratios, excitation wavelength 530 nm. Specific embodiments

[0022] In order to make the objectives, technical solutions and advantages of the present invention clearer and more understandable, the present invention will be further described in detail below in conjunction with specific embodiments.

[0023] The technical solutions of the present invention will be clearly and completely described below in conjunction with the embodiments of the present invention. Obviously, the described embodiments are only a part of the embodiments of the present invention, rather than all of the embodiments. All other embodiments obtained by those of ordinary skill in the art based on the embodiments of the present invention without creative efforts shall fall within the scope of protection of the present invention.

[0024] Example 1 The multifunctional fluorescence probe molecule for SO 2 and the preparation method for physiological environment polarity detection are as follows: Dissolve 7-(diethylamino)coumarin-3-carbaldehyde (245.3 mg, 1 mmol), benzo[b]thiophen-3(2H)-one 1,1-dioxide (273.3 mg, 1.5 mmol) and 1 drop of piperidine in anhydrous ethanol (10 ml). The mixed solution is stirred and refluxed at 80 °C for 12 h. After cooling, ethanol and piperidine are removed, and the crude product is purified by column chromatography (dichloromethane / methanol = 50 / 1) to obtain a blue-black solid, which is the fluorescence probe COU (280.0 mg, 68%).

[0025] Nuclear magnetic resonance hydrogen spectrum determination: 1 H NMR (400 MHz, DMSO-d 6 ) δ 8.93 (s, 1H), 8.47 (s,1H), 8.11 (d, J= 7.4 Hz, 1H), 8.04 (d, J = 7.6 Hz, 1H), 7.92 (td, J = 7.6, 1.2 Hz,1H), 7.84 (td, J = 7.5, 1.1 Hz, 1H), 7.49 (d, J = 9.1 Hz, 1H), 6.67 (dd, J = 9.1,2.5 Hz, 1H), 6.48 (d, J = 2.3 Hz, 1H), 3.51 (q, J = 7.2 Hz, 4H), 1.28 (t, J = 7.1Hz, 6H). 1H NMR measurement: 13 C NMR (101 MHz, DMSO-d 6 ) δ 177.9, 161.0, 158.5,153.9, 149.0, 143.6, 138.9, 135.9, 134.1, 132.9, 127.7, 124.7, 121.1, 110.6,109.8, 97.3, 45.6, 12.5。

[0026] The 1H NMR spectrum is as shown in Figure 2 Figure [X], and the 13C NMR spectrum is as shown in Figure 3 Figure [X].

[0027] High-resolution mass spectrometry measurement: HR-MS (m / z) calcd for chemical formula C 22 H 20 NO 5 + [M] + : 410.1057, Found: 410.1055. The high-resolution mass spectrum is as shown in Figure 4 Figure [X].

[0028] Example 2 Accurately weigh the probe COU prepared in Example 1 and dissolve it in dimethyl sulfoxide (DMSO) solution to prepare a 1 mM solution for standby.

[0029] SO 2 (10 mM) solution preparation: Weigh NaHSO 3 and dissolve it in 10 ml of deionized water, transfer it to a 10 mL volumetric flask and make up to the mark, and store it at -20 °C (prepare and use immediately).

[0030] The fluorescence in a PBS solution containing the probe COU (10 μM) was measured using a fluorescence spectrometer.

[0031] As Figure 5 shown, when 420 nm was used as the excitation wavelength, the individual probe showed almost no fluorescence emission at 480 nm; subsequently, after adding solutions of different concentrations of NaHSO 3 (0 - 20 μM), the fluorescence at 480 nm increased significantly.

[0032] As Figure 6 shown, the fluorescence of the probe COU at 480 nm was linearly fitted with the concentration of the added NaHSO 3 , obtaining the linear equation Y = 340.4X + 620.4, and having a high linear relationship. The detection limit was calculated to be 40.2 nM according to the lowest detection line equation LOD = 3σ / k. This indicates that the probe COU has high sensitivity for the detection of SO 2 .

[0033] Example 3 The concentration of the fluorescence probe COU in Example 1 was maintained at 10 μM, and the selectivity of this probe for SO 2 was investigated.

[0034] As Figure 7 shown, in the PBS solution, the individual probe showed almost no fluorescence emission. After adding other analytes, almost no fluorescence enhancement of the probe was caused. Only after adding NaHSO 3 (20 μM), a significant fluorescence enhancement (30 - fold) of the probe at 480 nm was induced. Although the addition of TBHP and Na 2 S 2 O 4 caused partial fluorescence enhancement, the fluorescence enhancement amplitude of the probe for SO 2 was more. The above experimental results indicate that the probe has high selectivity for the detection of SO 2 .

[0035] Example 4 The concentration of the probe COU in Example 1 was maintained at 10 μM, and the response time of this probe for SO 2 was investigated. In the PBS solution of the probe COU (10 μM), NaHSO 3 (0, 5 μM, 10 μM, 20 μM) was added, and the fluorescence change of the probe at 480 nm was recorded.

[0036] As Figure 8 shown, the probe COU showed no fluorescence emission without adding NaHSO 3At this time, its fluorescence at 480 nm shows almost no change within 0 - 600 s. When adding NaHSO 3 (5 μM, 10 μM, 20 μM), its fluorescence signal increases significantly within 1 minute. The above experiments indicate that the probe has a fast response time for the detection of SO 2 .

[0037] Example 5 Keep the concentration of the probe COU in Example 1 at 10 μM, and investigate the fluorescence emission of this probe in different polar solvents. As Figure 9 shown, when the probe COU is in low-polarity solvents such as n-hexane, toluene, and 1,4-dioxane, it has strong fluorescence emission (around 580 nm).

[0038] However, in DMSO, water, acetonitrile, and DMF, the fluorescence emission is relatively weak. This shows that the probe has different fluorescence emissions for solutions with different polarities and can have a good response to polarity.

[0039] Example 6 Keep the concentration of the probe COU in Example 1 at 10 μM, and investigate the fluorescence emission of this probe in a mixed solvent of 1,4-dioxane / water with different ratios.

[0040] As Figure 10 shown, the probe COU has obvious fluorescence emission at 580 nm in 1,4-dioxane (100%). When the proportion of water increases, that is, the polarity of the solution increases, the fluorescence emission decreases significantly. This shows that the probe is highly sensitive to polarity.

[0041] The above are only the preferred embodiments of the present invention and are not used to limit the present invention. Any modifications, equivalent replacements, improvements, etc. made within the spirit and principle of the present invention shall be included in the protection scope of the present invention.

Claims

1. A multifunctional fluorescent probe based on coumarin derivatives, the structural formula of which is as follows: 。 2. The method for preparing the multifunctional fluorescent probe according to claim 1, comprising the following steps: 7-(Diethylamino)coumarin-3-carboxaldehyde, benzo[B]thiophene-3(2H)-one 1,1-dioxide and piperidine are stirred and refluxed at 80° C. in an anhydrous ethanol solution. After cooling, ethanol and piperidine are removed, and the crude product is purified by column chromatography to obtain a blue-black solid, which is the fluorescent probe.

3. The preparation method according to claim 2, wherein The molar ratio of 7-(diethylamino)coumarin-3-carboxaldehyde to benzo[B]thiophene-3(2H)-one 1,1-dioxide is 1:(1.2-2).

4. The preparation method according to claim 3, wherein The molar ratio of 7-(diethylamino)coumarin-3-carboxaldehyde to benzo[B]thiophene-3(2H)-one 1,1-dioxide is 1:1.

5.

5. The preparation method according to claim 2, wherein: The reaction was stirred at reflux for 6-24 hours.

6. The preparation method according to claim 5, wherein: The reaction was stirred at reflux for 12 hours.

7. The preparation method according to claim 2, wherein: The eluent used for column chromatography purification was dichloromethane and methanol in a volume ratio of (100-20):

1.

8. The preparation method according to claim 7, wherein: The eluent used for column chromatography purification was dichloromethane and methanol in a volume ratio of 50:

1.

9. Use of the multifunctional fluorescent probe based on coumarin derivatives according to claim 1 in detecting sulfur dioxide and polarity of physiological environment.

10. The use according to claim 9, wherein The minimum detection limit of the multifunctional fluorescent probe based on coumarin derivatives for SO2 is 40.2 nM.

Images