Octapolar conjugated cyanine dye fluorescent probe and its application

Abstract

The present invention relates to an octapole conjugated cyanine dye fluorescent probe and its application in detecting hypochlorite and bisulfite ions. The octapole conjugated cyanine fluorescent probe utilizes a hexyl-substituted iodinated alkane and 1,1,2-trimethyl-1H-benz[e]indole to react to form a corresponding quaternary ammonium salt, which is then condensed with 2,4,6-trihydroxy-1,3,5-benzenetricarboxaldehyde. The probe is a dual-function fluorescent probe capable of detecting hypochlorite and bisulfite ions, exhibits good sensitivity to a variety of interfering ions, and can achieve both qualitative and quantitative detection.

Description

Technical Field

[0001] The invention relates to an octapole conjugated cyanine dye fluorescent probe and application thereof in detecting hypochlorite and bisulfite ions. Background Art

[0002] Hypochlorous acid (HClO) / hypochlorite (ClO - ) ions are important signal molecules in reactive oxygen species and play an important role in biomedicine and environmental safety. As a disinfectant in our daily lives, it can kill bacteria in water, so HClO is often used to sterilize domestic water. However, excessive HClO will produce trihalomethanes (THMs), which cause serious harm to humans, animals and plants. In biological systems, hypochlorous acid is produced by hydrogen peroxide and chloride ions under the catalysis of myeloperoxidase (MPO). Bisulfite ion (HSO3 - ) is widely present in the human living environment. Due to rapid industrial development, SO2 in the air has gradually increased, which not only easily causes hazards such as acid rain, but can also enter the human body through respiration, easily generating sulfites or bisulfites in the body. Excessive sulfites and bisulfites can easily cause asthma, allergies, gastrointestinal discomfort and other diseases. In addition, bisulfite is an important antioxidant and preservative, widely used in food, medicine, alcohol, and cosmetics. Excessive bisulfite can cause great harm to the human body and is classified as a Group III carcinogen by the World Health Organization and the International Agency for Research on Cancer. The Food and Agriculture Organization of the United Nations and the World Health Organization recommend that the bisulfite level in the human body should be below 0.7mg / kg.

[0003] So far, some cyanine fluorescent probes have been developed to detect ClO - and HSO3 - For example, Sun et al. reported a near-infrared fluorescent probe based on oxidative cleavage for the detection of ClO -(M. Sun, H. Yu, H. Zhu, et al. Anal. Chem. 2014, 86, 671-677.). Zhang et al. developed a probe targeting mouse lungs by linking a cyanine derivative with an aliphatic chain to an amino group, which can effectively detect the content of HClO in the body (X. Zhang, W.Zhao, B. Li, et al. Chem. Sci., 2018,). Xiong et al. used the strong oxidizing properties of HOCl to develop a fluorescent probe for detecting endogenous hypochlorous acid and successfully applied it to the detection of HOCl in 84 disinfectants (K. Xiong, F. Huo, Y.Zhang, et al. Anal. Methods 2019, 11, 1751–1756.). Yin et al. reported the detection of HSO3 - (G. Yin, Y. Gan, T. Yu, et al. Talanta 2019, 191, 428–434.). However, the fluorescent probes for hypochlorite and bisulfite ions reported so far still have many shortcomings, such as only fluorescence color change, low sensitivity, detection affected by other ions, and long reaction time. In addition, it is not possible to simultaneously detect ClO - and HSO3 - There has been no report on the octapole conjugated cyanine fluorescent probe for ions so far, so this invention has important pioneering significance in this field. Summary of the Invention

[0004] The present invention provides a method for selectively detecting ClO - and HSO3 - The dual-function fluorescent probe has not only fluorescence ratio changes but also color ratio changes. Its fluorescence ratio and color ratio changes are similar to those of ClO - and HSO3 - The concentration of ClO is linearly correlated, and ClO can be detected qualitatively and quantitatively. - and HSO3 - concentration.

[0005] The advantages of the present invention are as follows:

[0006] 1. The raw materials for the synthesis of this compound are readily available, the synthetic operation is simple, and the preparation is convenient;

[0007] 2. Under the conditions described, the compound reacts with ClO - and HSO3 - The reaction is sensitive and highly selective, and can be used for qualitative and quantitative detection of ClO - and HSO3 - .

[0008] The present invention adopts the following technical scheme: using hexyl-substituted iodinated alkane and 1,1,2-trimethyl-1H-benz[e]indole to react to generate the corresponding quaternary ammonium salt, which is then condensed with 2,4,6-trihydroxy-1,3,5-benzenetricarboxaldehyde to form an octapolar conjugated cyanine fluorescent probe. (1) The double bond in its structure can react with ClO - Nucleophilic addition reaction can occur to generate ethylene oxide and ethylene glycol, causing fluorescence and color changes, indicating ClO - The presence or quantitative detection of ClO - (2) By adding dilute HCl, the ketone group in the structure undergoes proton transfer under the catalysis of acid to form enol, which can react with HSO3 - An addition reaction occurs, and under the induction of visible light, the fluorescence and color change, indicating HSO3 - The presence or quantitative detection of HSO3 - concentration.

[0009] The structural formula of the fluorescent probe is as follows:

[0010]

[0011] The detection of ClO - The method is as follows: the probe is prepared into a methanol solution of a certain concentration, and then added to a solution containing ClO with a gradient concentration. - After the reaction is balanced, the fluorescence intensity of each sample is measured, and then the ClO - The concentration of ClO in the test solution is plotted as the horizontal axis and the fluorescence intensity of the system after the reaction is plotted as the vertical axis. The ClO in the test solution can be read from the graph based on the fluorescence intensity. - content.

[0012] The detection of HSO3 - The method is as follows: the probe is prepared into a methanol solution of a certain concentration, 1M dilute hydrochloric acid is added, and then the solution is added into a solution containing HSO3 with a gradient concentration. - After irradiation with visible light for 2 to 3 minutes, the UV intensity of each sample was measured after the reaction was balanced, and then the UV intensity of each sample was measured with HSO3 - The concentration of HSO3 in the solution to be tested is plotted as the horizontal axis and the UV intensity (275 nm) of the system after the reaction is plotted as the vertical axis. The HSO3 in the solution to be tested can be read from the graph based on the UV intensity. - content. BRIEF DESCRIPTION OF THE DRAWINGS

[0013] Figure 1 . Probe for ClO - The results of the selective experiment.

[0014] Figure 2 . Probe for ClO - Quantitative detection experimental results.

[0015] Figure 3 . Probe for HSO3 - The results of the selective experiment.

[0016] Figure 4 . Probe for HSO3 - Quantitative detection experimental results. DETAILED DESCRIPTION

[0017] The examples are provided to further illustrate the present invention, but the present invention is not limited to the examples.

[0018] Example 1 (Synthesis of Probe): This was carried out in two steps. In the first step, a mixture of 1.23 g (6.00 mmol) of 1,1,2-trimethyl-1H-benzo[e]indole and 3.81 g (18.00 mmol) of 1-iodohexane was refluxed in 80 mL of acetonitrile for 5 hours. After cooling to room temperature, a crude solid precipitated. This solid was filtered and washed with diethyl ether to yield 2.04 g of pink crystals of 3-propyl-1,1,2-trimethyl-1H-benzo[e]indole-3-iodide (yield: 81%). In the second step, 3-hexyl-1,1,2-trimethyl-1H-benzo[e]indole-3-iodide (0.632 g, 1.5 mmol) and 2,4,6-trihydroxy-1,3,5-benzenetricarboxaldehyde (0.105 g, 0.5 mmol) were heated to reflux in 5 mL of pyridine for 10 minutes. After cooling, the oily substance was washed several times with ether. The crude product was eluted through silica gel (CH2Cl2:CH3OH, 50:1, v / v) to obtain 0.21 g (41%) of the probe product.1H NMR (600 MHz, CDCl3): δ9.03 (dd, J = 18.1, 14.1 Hz, 2H), 8.94 (dd, J = 19.3, 13.9 Hz, 1H), 8.31 –8.05 (m, 6H), 7.91 (dd, J = 8.6, 3.1 Hz, 3H),7.88 (dt, J = 8.0, 3.8 Hz, 3H),7.57 (dt, J = 14.7, 7.5 Hz, 3H), 7.40 (dt, J = 9.3, 6.4 Hz, 3H), 7.30–7.25(m, 3H), 4.11 (dq, J = 24.5, 8.5, 7.9 Hz,6H), 2.15 (s, 18H), 1.96 (d, J = 7.0Hz, 6H), 1.63–1.36 (m, 18H), 0.96 (dt, J = 31.5, 7.2 Hz, 9H). 13C NMR (150MHz, CDCl3): δ 188.93, 186.48, 184.96, 175.19, 174.47, 173.87, 147.36,146.66, 146.53, 145.80, 140.64,140.55, 140.49, 140.38, 132.85, 132.64,132.57, 132.37, 130.91, .74, 118.78, 110.09, 110.06, 109.97,109.95, 99.20, 98.70, 98.60, 50.26, 50.10, 49.92, 43.74, 43.66, 43.59, 31.59,31.50, 28.84, 28.74, 27.05, 27.02, 26.91, 26.76, 22.67, 22.65,14.14, 14.07.

[0019] Example 2 (Probe to ClO -Ion selectivity): Take 9 μL of 1 mM probe methanol solution, 45 μL of 1 mM NaClO solution, 45 μL of 10 mM various interfering ions (AcO - 、CN - 、SCN - 、S2O3 2- , CO3 2- , H2PO4 - 、HSO3 - 、NO3 - , F - , Cl - Br - , I - ), dilute to 3 mL with methanol, mix well and add 2 mL of the solution into a cuvette, and measure the fluorescence emission spectrum of the working solution (λex = 460 nm, grating width 5 nm, 5 nm). - The selective experimental results are as follows Figure 1 As shown in the figure, when ClO is added - After addition of ClO, the fluorescence intensity of the probe changed significantly and the fluorescence was quenched, while other interfering ions in the sample hardly caused any change in the spectrum. - The color of the sample with α-aminobutyric acid changed to pink, while the color of other analytes did not change.

[0020] Example 3 (Probe to ClO - Quantitative detection of ions): 9 μL of 1 mM probe methanol solution and 9 μL of sodium hypochlorite solution with different concentrations were added to 3 mL of methanol, mixed and 2 mL of the solution was added to the cuvette, and the fluorescence emission spectrum of the working solution was measured (λex = 460 nm, grating width 5 nm, 5 nm). - The experimental results of Figure 2 As shown in the results, it can be seen that the absorption intensity is linearly related to the concentration, which can be used to quantitatively detect ClO - concentration.

[0021] Example 4 (Probe to HSO3 - Ion selectivity): 9 μL of 1 mM probe methanol solution, 12 μL of 1 M dilute hydrochloric acid, 72 μL of 1 mM sodium bisulfite solution, 36 μL of 10 mM various interfering ions (AcO - , CN - , SCN - , CO3 2- ,H2PO4 - , S2O3 2- , NO3 - , F- ,Cl - , Br - , I - , ClO - ) Dilute to 3 mL with methanol, mix well and irradiate with visible light for 2 to 3 minutes, take 2 mL of solution and add it to the cuvette, and measure the UV-visible absorption spectrum (200-900 nm) of the working solution. - The selective experimental results are as follows Figure 3 As shown in the figure, when HSO3 is added - After addition of HSO3, the probe's UV spectrum shifted blue and the absorption intensity changed significantly, while other interfering ions in the sample hardly caused any spectral changes. - The color of the sample changed to pink, while the color of other analytes did not change. From the results, it can be seen that the absorption intensity is linearly related to the concentration, which can be used to quantitatively detect HSO3 - concentration.

[0022] Example 5 (Probe to HSO3 - Quantitative detection of ions): 9 μL of 1mM probe methanol solution, 12 μL of 1M dilute hydrochloric acid and 9 μL of sodium bisulfite solution with different concentrations were added to 3 mL with methanol in turn. After mixing, 2 mL of the solution was added to a cuvette and the UV-visible absorption spectrum of the working solution was measured (200-900 nm). The probe reacted with different concentrations of HSO3 - The experimental results of Figure 4 As shown. From the results, it can be seen that the absorption intensity at 275 nm is linearly related to the concentration, which can be used to quantitatively detect HSO3 - concentration.

Claims

1. A fluorescent probe, characterized in that The structure of the fluorescent probe is as follows:

2. Use of the fluorescent probe according to claim 1 in the detection of hypochlorite ions for non-disease diagnosis purposes, characterized in that: With an excitation wavelength of 460 nm, the fluorescence intensity of the probe is linearly correlated with the concentration of hypochlorite ions, thereby achieving qualitative or quantitative detection of hypochlorite ions.

3. Use of the fluorescent probe according to claim 1 in the detection of bisulfite ions, characterized in that: The method comprises the following steps: adding 1M dilute hydrochloric acid to the probe solution, irradiating the solution with visible light for 2 to 3 minutes, and then measuring the ultraviolet absorption intensity at 275 nm. The intensity is linearly correlated with the concentration of bisulfite ions, thereby achieving qualitative or quantitative detection of bisulfite ions.

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