Fluorescent probe based on 2-hydrazinopyridine modified aminobenzophenoxazinone derivative and application of fluorescent probe

Abstract

The invention discloses a fluorescent probe based on a 2-hydrazinopyridine modified amino benzophenoxazinone derivative and application of the fluorescent probe. The invention relates to a 2-hydrazinopyridine modified aminobenzophenoxazinone derivative, which is simply recorded as CdP, and the structure of the 2-hydrazinopyridine modified aminobenzophenoxazinone derivative is as follows: imgabs0 #. According to the invention, an amino benzophenoxazinone derivative is modified by 2-hydrazinopyridine, so that an off-on type fluorescent probe is obtained. The probe can identify Cd < 2 + > in an aqueous solution and an electronic cigarette compound, has obviously enhanced fluorescence intensity, and provides a powerful tool for rapid detection of the content of Cd < 2 + > in the electronic cigarette compound.

Description

Technical Field

[0001] The present invention belongs to the field of fluorescent materials, and particularly relates to a fluorescent probe based on 2-hydrazinylpyridine modified aminobenzophenoxazinone derivative and its application. Background Art

[0002] Cadmium (Cd) is a heavy metal that has a serious toxic effect on human health, which can lead to renal insufficiency, calcium metabolism disorders, and increase the incidence of various cancers. Due to the extensive application of cadmium in industry and agriculture, a large amount of cadmium has accumulated in soil and water areas, and then enters the human body through the food chain. Therefore, rapid and efficient detection of Cd 2+ is of great significance for environmental monitoring and disease prevention.

[0003] A fluorescent probe is a chemical reagent that can detect specific molecules or ions through changes in fluorescence signals. When excited by light of a specific wavelength, the fluorescent probe emits fluorescence of a specific wavelength, and the change in its intensity or wavelength can reflect the presence and concentration of the target ion.

[0004] In recent years, many cadmium ion fluorescent probes have been reported and applied in many aspects such as environmental monitoring and in vivo biological research. However, these probes may encounter some problems, such as low quantum yield and small Stokes shift. Aminobenzophenoxazinone derivative dyes are selected to improve the optical performance of the probe, solve the problems of low quantum yield, small Stokes shift, and low fluorescence intensity, because of their good photostability, emission wavelength in the near-infrared region, large Stokes shift, high quantum yield, and easy to observe with the naked eye. At the same time, due to the structural rigidity of benzophenoxazinone, it can specifically bind to cadmium ions, making its selectivity much higher than other existing probes. In summary, the Cd 2+ fluorescent probe of the present invention has broad application prospects in environmental monitoring and biomedical research due to its high selectivity, sensitivity, and rapid response characteristics. Summary of the Invention

[0005] The present invention provides a fluorescent probe based on 2-hydrazinylpyridine modified aminobenzophenoxazinone derivative and its application. The present invention modifies aminobenzophenoxazinone derivative with 2-hydrazinylpyridine to obtain a "turn-off-turn-on" type fluorescent probe. This probe can identify Cd 2+ in aqueous solution and electronic cigarette aerosol, and has significantly enhanced fluorescence intensity, providing a powerful tool for the rapid detection of Cd 2+ content in electronic cigarette aerosol.

[0006] The fluorescent probe of the present invention based on 2-hydrazinylpyridine modified aminobenzophenoxazinone derivative has a molecular formula of C 22 H 15 N 5 O2 , abbreviated as CdP, and its structure is as follows:

[0007] .

[0008] The preparation method of the fluorescent probe CdP of the present invention includes the following steps:

[0009] Step 1: Dissolve 5.2 g (30 mmol) of 2-hydroxy-1,4-naphthoquinone and 4.6 g (30 mmol) of 2-hydroxy-4-nitroaniline in an 80% aqueous acetic acid solution, stir and react at 100 °C for 12 h, rotary evaporate to remove most of the acetic acid, pour the remaining liquid into water, adjust the pH to neutral with sodium bicarbonate, filter to collect the precipitate, extract with ethyl acetate and water, collect the organic phase, and separate with a silica gel column to obtain 6.54 g of compound 1 (yellow solid powder, yield 74.6%).

[0010] Step 2: Dissolve 2.92 g (10 mmol) of compound 1 in absolute ethanol, add 5.23 g (80 mmol) of zinc powder and 2 mL of dilute hydrochloric acid, reflux and react at 80 °C for 4 h, filter by suction, collect the filtrate, and perform column chromatography with petroleum ether and ethyl acetate (1:1) as the mobile phase to obtain 2.28 g of compound 2 (black solid powder, yield 87.7%).

[0011] Step 3: Dissolve 1.31 g (5 mmol) of compound 2 and 1.28 g (10 mmol) of dichloromethyl methyl ether in dichloromethane, stir at room temperature for 5 min, then add 3.8 g (20 mmol) of titanium tetrachloride and continue to stir for 30 min, add ice water to quench the reaction, extract with ethyl acetate and collect the organic phase, and perform column chromatography with petroleum ether and ethyl acetate (2:1) as the mobile phase to obtain 1.15 g of compound 3 (dark red solid powder, yield 79.3%).

[0012] Step 4: Dissolve 1.45 g (5 mmol) of compound 3 and 0.55 g (5 mmol) of 2-hydrazinopyridine in ethanol, add 0.5 mL of acetic acid, reflux and stir for 8 h, filter to obtain 1.65 g of the target product 4 (CdP, dark red solid powder, yield 86.6%).

[0013] The synthetic route is as follows:

[0014]

[0015] The application of the fluorescent probe CdP of the present invention in the preparation of Cd 2+ detection reagent.

[0016] Furthermore, Cd 2+In the equivalent range of 0.1 - 0.6 eq in concentration, the fluorescence intensity of the fluorescent probe at 610 nm has a linear relationship with the concentration of Cd 2+ and the detection limit is 1.54 nM.

[0017] Recognition mechanism of the fluorescent probe (CdP):

[0018]

[0019] Compared with the prior art, the probe of the present invention has better selectivity. Due to the structural rigidity of benzophenoxazine ketone, it can specifically bind to cadmium ions. After binding, the solution changes from yellow to dark red and emits red fluorescence with a longer wavelength. The color change is more obvious under natural light and can be observed more clearly with the naked eye, and it can be judged without the aid of instruments.

[0020] In addition, due to the limited detection of low-concentration cadmium ions by existing Cd 2+ fluorescent probes, and the low concentration of Cd 2+ in e-cigarette aerosols, it is difficult to displace the zinc ions in the existing Cd 2+ fluorescent probes, and the fluorescence change is not obvious. Therefore, the existing Cd 2+ fluorescent probes cannot be applied to the detection of Cd 2+ in e-cigarette aerosols.

[0021] Compared with the prior art, the beneficial effects of the present invention are reflected in:

[0022] Due to the PET effect of 2-hydrazinopyridine in the probe CdP of the present invention, the fluorescence of the aminobenzophenoxazine ketone skeleton is in the off state. After the reaction, due to the binding of Cd 2+ , the PET effect and the high quantum yield of the aminobenzophenoxazine ketone skeleton itself are blocked, resulting in a bright color change visible to the naked eye and red fluorescence.

[0023] Before the reaction, the solution of the probe CdP of the present invention is yellow, and after the reaction, it turns to dark red, and the phenomenon is obvious and can be observed with the naked eye. Its fluorescence is significantly enhanced at 610 nm, and it is a "Turn-on" type probe.

[0024] The probe CdP of the present invention can quickly and specifically recognize Cd 2+ in aqueous solutions and e-cigarette aerosols. Brief Description of the Drawings

[0025] Figure 1 are photos of the fluorescent probe CdP before and after the reaction with Cd 2+ under 365 nm ultraviolet light. As can be seen from Figure 1 , after the reaction of CdP with Cd 2+ , the fluorescence changes from darker yellow fluorescence to bright red.

[0026] Figure 2 is the UV spectrum after the reaction of the fluorescent probe CdP (20 μM) with Cd 2+ (20 μM). It can be seen from Figure 2 that there is a red shift at the maximum absorption peak after the reaction.

[0027] Figure 3 is the fluorescence intensity change curve graph of the fluorescent probe CdP after reacting with Cd 2+ (0 - 18 μM) in the range of 475 nm to 725 nm. It can be seen from Figure 3 that the fluorescence emission gradient of CdP enhances after reacting with Cd 2+ (0 - 18 μM).

[0028] Figure 4 is the linear fitting graph of the relative fluorescence intensity of the fluorescent probe CdP at 610 nm after reacting with Cd 2+ (0 - 20 μM). It can be seen from Figure 4 that the relative fluorescence intensity of CdP after reacting with Cd 2+ (0 - 20 μM) has a good linear correlation with C Cd 2+ / C CdP (the equivalent ratio of the two).

[0029] Figure 5 is the selectivity test graph of the fluorescent probe CdP. It can be seen from Figure 5 that CdP has good selectivity for Cd 2+ .

[0030] Figure 6 is the interference test graph of the fluorescent probe CdP. It can be seen from Figure 6 that when Cd 2+ coexists with potential interfering substances, the reaction of CdP with Cd 2+ is not affected. Detailed implementation manners

[0031] The technical solution of the present invention will be further analyzed and explained through specific embodiments below.

[0032] Example 1: Preparation of CdP

[0033]

[0034] Dissolve 1.45 g (5 mmol) of Compound 3 and 0.55 g (5 mmol) of 2-hydrazinopyridine in ethanol, add 0.5 mL of acetic acid, reflux and stir for 8 h. After stopping the reaction, filter to obtain 1.65 g of Compound 4 (CdP, dark red solid powder, yield 86.6%). 1 H NMR (600 MHz, Chloroform-d) δ 9.07 (s, 2H), 8.66 (d, J = 0.6 Hz,2H), 8.39 (dd, J = 4.3, 1.7 Hz, 2H), 8.14 (dd, J = 7.5, 1.3 Hz, 2H), 7.99(dd, J = 7.6, 1.3 Hz, 2H), 7.74 (td, J = 7.0, 1.5 Hz, 2H), 7.70 (s, 2H), 7.68(td, J = 7.6, 1.3 Hz, 2H), 7.53 (td, J = 7.8, 1.3 Hz, 2H), 7.37 (dd, J = 7.0,1.4 Hz, 2H), 7.14 – 7.09 (m, 2H), 6.95 (s, 2H), 6.49 (s, 2H), 6.08 (s, 4H).

[0035] Example 2: Spectral properties of CdP

[0036] First, prepare a 1 mM mother liquor of CdP with dimethyl sulfoxide (DMSO), and then dilute it to 20 μM with ethanol; the cadmium ion standard solution is obtained by dissolving cadmium chloride in deionized water. The absorption spectrum of CdP in an ethanol solution (20 μM) is approximately at 463 nm. When we add Cd 2+ (20 μM) to the probe solution, we find that immediately after the addition of Cd 2+ a reaction occurs between the probe and Cd 2+ accompanied by a change in the color of the solution, and the absorption peak at 463 nm blue-shifts to 520 nm. ( Figure 1 、 Figure 2 )

[0037]

[0038] For quantitative analysis, subsequently, we record the fluorescence intensity (Ex: 543 nm, Em = 610 nm) after the reaction of the probe (20 μM) with 0 - 20 μM Cd 2+ ( Figure 3 ), and find that when the probe CdP is added to Cd 2+The fluorescence intensity at 610 nm is linearly related to the Cd concentration in the range of 0.1 - 0.6 eq, and a standard curve is plotted ( 2+ ). From this, the detection limit of the probe is calculated to be as low as 1.54 nM, and through multiple calibrations and validations, the high precision and high reliability of the measurement results are ensured. Figure 4 ).

[0039]

[0040] Then, we used a variety of common interfering ions to test the probe and determined that the probe CdP has significant high selectivity and anti-interference ability for cadmium ions ( Figure 5 , Figure 6 ).

[0041] Subsequently, the present invention used the standard addition method. By adding Cd 2+ standards to the e-cigarette oil, the detection concentration was expressed as the ratio of its concentration to that of the detection probe CdP (equivalent ratio), and its gradient was set to 0.15, 0.25, 0.35, 0.45, 0.55. The fluorescence intensity after the reaction with the fluorescence probe CdP was detected using a fluorescence spectrophotometer in the optimized reaction system, and according to the obtained linear fitting equation, the corresponding Cd 2+ concentration was calculated. The results are shown in Table 3, and the recovery rate ranges from 97.8% - 103.6%, indicating that CdP can accurately detect Cd in the e-cigarette oil in the concentration range of 0 - 0.6 equivalent. 2+ .

[0042]

Claims

1. A 2-hydrazinopyridine-modified aminobenzophenoxazinone derivative, referred to as CdP, characterized in that Its structure is as follows: 。 2. The method for preparing the 2-hydrazinopyridine-modified aminobenzophenoxazinone derivative according to claim 1, characterized in that The steps include: Step 1: dissolving 2-hydroxy-1,4-naphthoquinone and 2-hydroxy-4-nitroaniline in an aqueous acetic acid solution, stirring and reacting at 100° C., removing most of the acetic acid by rotary evaporation after the reaction, pouring the remaining liquid into water, adjusting the pH to neutral with sodium bicarbonate, collecting the precipitate by filtration, extracting with ethyl acetate and water, collecting the organic phase, and separating with a silica gel column to obtain compound 1; Step 2: Compound 1 was dissolved in anhydrous ethanol, zinc powder and dilute hydrochloric acid were added, and the mixture was reacted under reflux. After the reaction was completed, the mixture was filtered, and the filtrate was collected and separated by column chromatography to obtain compound 2; Step 3: Compound 2 and dichloromethyl methyl ether were dissolved in dichloromethane, stirred at room temperature, and then titanium tetrachloride was added and the reaction was continued with stirring for 30 min. Ice water was added to quench the reaction, and the organic phase was extracted with ethyl acetate and collected. Compound 3 was obtained by column chromatography. Step 4: dissolve compound 3 and 2-hydrazinopyridine in ethanol, add acetic acid, reflux and stir to react, and filter after the reaction to obtain the target product 4; The synthetic route is as follows: 。 3. The method of claim 1 for preparing Cd based on 2-hydrazinopyridine modified aminobenzophenoxazinone derivative 2+ Application in detection reagents.

4. The use according to claim 3, characterized in that: The detection reagent can specifically identify Cd in aqueous solution and / or electronic cigarette aerosol 2+ .

5. The use according to claim 4, characterized in that: In the detection system, Cd 2+ When the concentration is in the equivalent range of 0.1-0.6 eq, the fluorescence intensity of the detection reagent at 610 nm is similar to that of Cd 2+ The concentration is linear.

6. The use according to claim 5, characterized in that: The detection reagent is effective for Cd 2+ The detection limit was 1.54 nM.

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