Novel rhodamine Al < 3 + > fluorescent probe as well as preparation method and application thereof

Abstract

The invention belongs to the field of fluorescent probes, and discloses a novel rhodamine Al < 3 + > fluorescent probe as well as a preparation method and application thereof. The method comprises the following steps: synthesizing an intermediate R from cyanuric chloride serving as a framework and p-hydroxybenzaldehyde, synthesizing rhodamine B hydrazide from rhodamine B and hydrazine hydrate under the protection of nitrogen, and finally synthesizing the final product probe R1 from the intermediate R and the rhodamine B hydrazide. After 10 times of KSCN is added into the probe R1 as a color developing agent, the interference of Fe < 3 + >, Cu < 2 + > and Zn < 2 + > ions is masked, and the probe R1 shows high sensitivity and selectivity to Al < 3 + >. Furthermore, the lowest detection limit of Al < 3 + > is calculated to be 0.1 mu mol / L through fluorescence titration test. Therefore, the probe R1 prepared by the invention can realize high-selectivity fluorescent recognition on Al < 3 + > ions, and can be used as a fluorescent probe for detecting metal ions to be applied to the related fields of environment detection, biomedical treatment, agriculture and the like.

Description

Technical Field

[0001] The present invention belongs to the field of fluorescent probes, and more specifically relates to a novel rhodamine-based Al 3+ Fluorescent probe and its preparation method and application. Background Art

[0002] Aluminum is a non-essential element for the human body and is widely present in nature. A large number of studies have shown that excessive aluminum entering human cells through the blood will interfere with their normal metabolism, causing symptoms such as anemia and reduced immunity. In severe cases, it may also lead to diseases such as Alzheimer's disease and Parkinson's disease.

[0003] The detection methods of aluminum mainly include titration, ultraviolet spectrophotometry, graphite furnace atomic absorption spectrometry, inductively coupled plasma atomic emission spectrometry, inductively coupled plasma mass spectrometry, etc. At present, due to the high cost of inductively coupled plasma emission instruments and graphite furnace atomic absorption instruments, it is difficult to popularize them at the grassroots level. Spectrophotometry is more suitable for the monitoring needs of aluminum content by grassroots testing institutions and processing and production enterprises in my country at this stage, but the operation steps are relatively cumbersome and require the addition of many types of reagents. The development of a highly sensitive and highly selective aluminum ion fluorescent probe can provide effective technical support for aluminum ion-related detection.

[0004] Therefore, how to provide a new type of rhodamine Al 3+ Fluorescent probes and their preparation methods and applications are issues that need to be urgently addressed by those skilled in the art. Summary of the invention

[0005] In order to overcome the shortcomings and deficiencies in the prior art, the present invention provides a novel rhodamine-based Al 3+ Fluorescent probe and its preparation method and application.

[0006] In order to achieve the above object, the present invention adopts the following technical solution:

[0007] A new type of rhodamine Al 3+ Fluorescent probe, the fluorescent probe structure is as follows:

[0008]

[0009] The present invention also discloses a novel rhodamine Al 3+ The method for preparing a fluorescent probe comprises the following steps:

[0010] (1) Synthesis of intermediate R

[0011] Add p-hydroxybenzaldehyde and THF to a reaction container. After the p-hydroxybenzaldehyde is completely dissolved, add an excess of triethylamine dropwise at a uniform speed. After the dropwise addition is completed, place the mixture in an ultrasonic oscillation reactor and mix evenly. Then weigh cyanuric chloride in a beaker, add THF to dissolve the mixture to obtain a cyanuric chloride solution. Transfer the cyanuric chloride solution to a constant pressure funnel, and dropwise add the mixture to the reaction container at a speed of 2 to 3 drops / s. Perform ultrasonic oscillation reaction while dropping the mixture. After the dropwise addition is completed, heat the mixture to 80° C. and continue ultrasonic oscillation for 40 minutes. After the reaction is completed, cool the mixture to room temperature, add 100 mL of a 10% sodium carbonate solution by mass, stir evenly, filter the mixture, wash with distilled water, dry the mixture, and recrystallize the mixture to obtain a white powder, namely, intermediate R.

[0012] (2) Preparation of Rhodamine B Hydrazide

[0013] Rhodamine B and methanol are added to a reactor to obtain a Rhodamine B solution, wherein the molar concentration of the Rhodamine B solution is 0.1 mol / L. Under nitrogen protection, 80% hydrazine hydrate solution is slowly added dropwise in multiple times. After the addition is completed, the solution is refluxed and stirred, and the color of the solution changes from pink to brown. After the reaction is completed, the reaction solution is transferred to a large amount of deionized water to precipitate a pink precipitate. The precipitate is filtered, washed with distilled water, and dried to obtain Rhodamine B hydrazide;

[0014] (3) Synthesis of fluorescent probe R1

[0015] Rhodamine B hydrazide and intermediate R are added to a reaction vessel, anhydrous ethanol is used as the solvent, a condensation reflux device is installed, and reflux and boil the reaction under magnetic stirring. The solution generates a light red precipitate, which is filtered, recrystallized, and vacuum dried to obtain a pink-white solid, namely the fluorescent probe R1.

[0016] Preferably, in step (1), the temperature is raised to 80° C. and the ultrasonic reaction time is 40 min.

[0017] Preferably, in step (1), the molar ratio of triethylamine to p-hydroxybenzaldehyde is 5:4, and the molar ratio of cyanuric chloride to p-hydroxybenzaldehyde is 1:4.

[0018] Preferably, the solvent for recrystallization in step (1) is tetrahydrofuran and ethyl acetate in a volume ratio of 1:2.

[0019] Preferably, in step (2), the temperature of the oil bath is 75° C., and the stirring reflux time is 12 h.

[0020] Preferably, in step (2), the molar ratio of hydrazine hydrate to rhodamine B is 16:1.

[0021] Preferably, in step (3), the molar ratio of rhodamine B hydrazide to intermediate R is 3.1:1.

[0022] Preferably, the molar concentration of the intermediate R in anhydrous ethanol in step (3) is 0.02 mol / L.

[0023] Preferably, the reflux boiling reaction time in step (3) is 14 to 16 hours.

[0024] Preferably, the solvent for recrystallization in step (3) is methanol and ethanol in a volume ratio of 1:2.

[0025] The present invention also discloses the novel rhodamine Al 3+ Application of fluorescent probes in environmental monitoring, biomedicine, and agriculture.

[0026] The synthetic route of the fluorescent probe of the present invention is as follows:

[0027] (1) Synthesis of intermediate R

[0028]

[0029] (2) Preparation of Rhodamine B Hydrazide

[0030]

[0031] (3) Synthesis of fluorescent probe R1

[0032]

[0033] It can be seen from the above technical solution that, compared with the prior art, the present invention provides a new type of rhodamine Al 3+ Fluorescent probes and preparation methods and applications thereof have the following beneficial effects:

[0034] 1. The present invention first uses cyanuric chloride as a skeleton and p-hydroxybenzaldehyde to synthesize an intermediate R, then synthesizes rhodamine B hydrazide with hydrazine hydrate under nitrogen protection, and finally synthesizes the final product probe R1 with the intermediate R and rhodamine B hydrazide. The structure of R1 is verified by nuclear magnetic resonance hydrogen spectroscopy and mass spectrometry, which proves that R1 is successfully synthesized. The fluorescence response of R1 to 17 metal ions is tested by fluorescence spectroscopy, and the results show that R1 has a strong affinity to Al 3+ , Fe 3+ , Cu 2+ and Zn 2+ ions have obvious fluorescence response, among which Al 3+ , Fe 3+ The ion fluorescence red-shifted by 20nm, and the fluorescence was significantly enhanced, making it an enhanced (TURN-ON) fluorescence probe.

[0035] 2. After adding 10 times the amount of KSCN as a color developer to R1, Fe 3+ , Cu 2+ and Zn2+ Ion interference, probe R1 to Al 3+ The results showed high sensitivity and selectivity. The Al 3+ The minimum detection limit is 0.1 μmol / L. Therefore, the probe R1 prepared by the present invention can detect Al 3+ The highly selective fluorescence recognition of ions can be used as a fluorescent probe for detecting metal ions in environmental testing, biomedicine, agriculture and other related fields. BRIEF DESCRIPTION OF THE DRAWINGS

[0036] In order to more clearly illustrate the embodiments of the present invention or the technical solutions in the prior art, the drawings required for use in the embodiments or the description of the prior art will be briefly introduced below. Obviously, the drawings described below are only embodiments of the present invention. For ordinary technicians in this field, other drawings can be obtained based on the provided drawings without paying creative work.

[0037] Figure 1 This is the hydrogen nuclear magnetic resonance spectrum of the probe R1 obtained in Example 1.

[0038] Figure 2 This is the mass spectrometry characterization diagram of the probe R1 obtained in Example 1.

[0039] Figure 3 This is the fluorescence emission spectrum of the probe R1 obtained in Example 1.

[0040] Figure 4 This is the precision diagram of probe R1 obtained in Example 1.

[0041] Figure 5 This is a comparison chart of the fluorescence responses of the probe R1 obtained in Example 1 to various metal ions.

[0042] Figure 6 10 times KSCN with Al 3+ , Fe 3+ 、Zn 2+ , Cu 2+ Fluorescence response diagram.

[0043] Figure 7 The probe R1 obtained in Example 1 and Al 3+ Fluorescence titration curve.

[0044] Figure 8 This is the Hill-Plot titration linear graph at the emission wavelength of 581 nm. DETAILED DESCRIPTION

[0045] The technical solutions in the embodiments of the present invention are described clearly and completely below. Obviously, the described embodiments are only part of the embodiments of the present invention, not all of them. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without creative work are within the scope of protection of the present invention.

[0046] Example 1

[0047] The present invention also discloses a novel rhodamine Al 3+ The method for preparing a fluorescent probe comprises the following steps:

[0048] (1) Synthesis of intermediate R

[0049] Add p-hydroxybenzaldehyde (4.8846 g, 0.04 mol) and 50 mL THF into a reaction vessel. After the p-hydroxybenzaldehyde is completely dissolved, add triethylamine (7 mL, 0.05 mol) dropwise at a uniform speed. After the addition is complete, place the mixture in an ultrasonic oscillation reactor and mix well. Add cyanuric chloride (1.8440 g, 0.01 mol) into 30 mL THF and mix thoroughly to obtain a cyanuric chloride solution. Transfer the cyanuric chloride solution into a constant pressure funnel, and then dropwise add the mixture into the reaction vessel at a speed of 2 to 3 drops / s. Ultrasonic oscillation reaction is performed while the addition is completed. After the addition is completed, the mixture is heated to 80° C., ultrasonically reacted for 40 min, cooled to room temperature, poured into 100 mL of a sodium carbonate solution having a mass fraction of 10%, stirred and mixed at a uniform speed, and filtered, washed with distilled water, dried, and recrystallized to obtain a white powder, namely, intermediate R, with a mass of 2.9179 g and a yield of 66.17%.

[0050] (2) Preparation of Rhodamine B Hydrazide

[0051] Rhodamine B (9.58 g, 0.02 mol) was dissolved in 150 mL of methanol solution. Under nitrogen protection, 20 mL of 80% hydrazine hydrate solution was slowly added dropwise several times. The molar ratio of hydrazine hydrate to rhodamine B was 16:1. The mixture was refluxed and stirred in a 75°C oil bath for 12 h. The color of the solution changed from pink to brown. After the reaction, the reaction solution was poured into a large amount of deionized water to precipitate a pink precipitate. After suction filtration, the precipitate was washed with distilled water and dried to obtain rhodamine B hydrazide with a mass of 3.92 g and a yield of 54.14%.

[0052] (3) Synthesis of fluorescent probe R1

[0053] Rhodamine B hydrazide and intermediate R in a molar ratio of 3.1:1 were added to the reactor, with anhydrous ethanol as the solvent, the molar concentration of intermediate R in anhydrous ethanol was 0.02 mol / L, and a condensation reflux device was installed. The solution was refluxed and boiled under magnetic stirring for 16 hours, and a light red precipitate was generated in the solution. The solution was filtered, recrystallized, and vacuum dried to obtain a powdery white solid, namely the fluorescent probe R1, with a mass of 0.8785 g and a yield of 52.14%.

[0054] The solvent for recrystallization in step (1) is tetrahydrofuran and ethyl acetate in a volume ratio of 1:2.

[0055] The solvent for recrystallization in step (3) is methanol and ethanol in a volume ratio of 1:2.

[0056] The fluorescent probe R1 obtained in Example 1 was subjected to the following tests:

[0057] 1. Nuclear Magnetic Resonance Proton Spectrum Characterization

[0058] Nuclear magnetic resonance hydrogen spectrum test, TMS as internal standard, 1 The H-NMR operating frequency is 400 MHz. The structure of probe R1 is as follows:

[0059]

[0060] The H NMR spectrum of probe R1 is shown in Figure 1 The H NMR spectrum analysis is shown in Table 1.

[0061] Table 1 H NMR spectrum analysis of probe R1

[0062]

[0063]

[0064] 2. Mass spectrometry characterization

[0065] The mass spectrum of probe R1 is as follows Figure 2 As shown, m / z = 1757.63 is the quasi-molecular ion peak of R1 [M+H + ], the test results show that the ligand (probe R1) obtained by the reaction of the intermediate (R) with rhodamine B hydrazide was successfully synthesized. H NMR spectrum and mass spectrometry analysis showed that probe R1 was the target product and was pure.

[0066] 3. Fluorescence recognition study of probe R1 and common metal ions

[0067] R1 was prepared to a concentration of 1×10 -5 mol / L ethanol (trace DMF solvent) solution; Cr 3+ 、Ag + 、Cd 2+, Ca 2+ , Mn 2+ , Ba 2+ 、Zn 2+ 、Ni 2+ , Pb 2+ 、Co 2+ Cr 3+ 、Na + , Hg 2+ Mg 2+ 17 common metal ions were prepared into aqueous solutions with a concentration of 0.1 mol / L; Al 3+ , Cu 2+ , Fe 3+ The three metal ions were prepared into aqueous solutions with concentrations of 0.01 mol / L, 0.1 mol / L and 0.1 mg / L respectively; the color developer KSCN was 0.1 mol / L; and the concentration of HEPES buffer solution (hydroxyethylpiperazine ethanesulfonic acid) was 0.01 mol / L.

[0068] (1) Fluorescence spectrum and precision test of probe R1

[0069] Take 3mL of R1 solution and add it to a 1cm wide quartz cuvette. Scan the R1 fluorescence emission spectrum at a speed of 1000nm / min with an excitation wavelength of 480nm, an emission slit of 10nm, and a gain intensity of medium. Figure 3 As shown in the figure, it can be seen that R1 has a maximum fluorescence emission peak at a wavelength of 564nm.

[0070] Precision can well reflect the stability of R1 performance. By repeating the test of R1 ten times, the fluorescence spectrum is obtained, such as Figure 4 As shown. Then through The calculated precision is 7.38%, indicating that the measurement has high precision and provides more accurate data support for subsequent experimental data.

[0071] (2) Fluorescence response of probe R1 to 17 metal ions

[0072] Take 3mL of R1 (10μM, anhydrous ethanol) solution in a quartz cuvette and add 3μL of metal ion solution (0.1mM, 10 times the concentration of R1). 3+ 、Ag + 、Cd 2+ , Ca 2+ , Mn 2+ , Ba 2+ 、Zn 2+ 、Ni 2+ , Pb 2+ 、Co 2+ Cr 3+ 、Na+ , Hg 2 + Mg 2+ 17 kinds of metals were simultaneously tested for fluorescence detection and identification. The test results are as follows Figure 5 As shown, it can be seen that R1 has a great influence on Al 3+ , Cu 2+ 、Zn 2+ , Fe 3+ Metal ions have a fluorescence response phenomenon, in which the fluorescence emission of R1 is red-shifted. 3+ , Fe 3+ The ion emission wavelength red-shifted from 564nm to 584nm, and the fluorescence response was strong and the fluorescence was significantly enhanced; 2+ 、Zn 2+ The ion emission wavelength also showed a red shift, from 564nm to 576nm, and the fluorescence was significantly enhanced; but the fluorescence response to the other 13 metal ions was not obvious or even no response.

[0073] (3) R1 to Al 3+ Selective research

[0074] 1) Fluorescence recognition study after adding KSCN color developer

[0075] R1 was dissolved in 10 mL DMF to prepare a 0.01 mol / L solution, which was then dissolved in anhydrous ethanol. After adding 10 times the amount of KSCN color developer, 10 eq Zn 2+ 、10eqCu 2+ 、10eqFe 3+ and 10eqAl 3+ After solution, its fluorescence response is as Figure 6 As shown in the experimental results, it can be seen that at a wavelength of 584nm, R1 has a significant influence on Al 3+ The fluorescence response of ions after adding KSCN is still very strong, and compared with Al 3+ For Zn 2+ , Cu 2+ and Fe 3+ The fluorescence response of R1 decreased. 3+ High selectivity.

[0076] 2) Probe R1 and Al 3+ Fluorescence detection limit test

[0077] Add probe R1 and 10 times the amount of KSCN color developer to the (ethanol: buffer solution = 9:1) solution, and gradually add 10 to 400 μmol / L Al 3+ Metal ions, such as Figure 7 and Figure 8As shown in the figure, it is linearly correlated in the concentration range of 0 to 100 μmol / L, and the linear equation is F = 82.25 + 1.12 × 107χ (correlation coefficient: 0.99607). According to the detection limit equation D = 3S0 / B, the correlation coefficient of R1 to Al 3+ The detection limit is 0.1μmol / L, with high sensitivity.

[0078] The above description of the disclosed embodiments enables one skilled in the art to implement or use the present invention. Various modifications to these embodiments will be apparent to one skilled in the art, and the general principles defined herein may be implemented in other embodiments without departing from the spirit or scope of the present invention. Therefore, the present invention will not be limited to the embodiments shown herein, but rather to the widest scope consistent with the principles and novel features disclosed herein.

Claims

1. A new type of rhodamine Al 3+ A fluorescent probe, characterized in that The structural formula of the fluorescent probe R1 is shown below: 。 2. A novel rhodamine Al according to claim 1 3+ A method for preparing a fluorescent probe, characterized in that: The following steps are involved: Rhodamine B hydrazide and intermediate R are added to a reaction vessel, anhydrous ethanol is used as the solvent, a condensation reflux device is installed, and reflux and boil the reaction under magnetic stirring. The solution generates a light red precipitate, which is filtered, recrystallized, and vacuum dried to obtain a pink-white solid, namely the fluorescent probe R1.

3. A novel rhodamine Al according to claim 2 3+ A method for preparing a fluorescent probe, characterized in that: The structural formula of the intermediate R is shown below:

4. A novel rhodamine Al according to claim 2 3+ A method for preparing a fluorescent probe, characterized in that: The structural formula of the rhodamine B hydrazide is as follows:

5. A novel rhodamine Al according to claim 2 3+ A method for preparing a fluorescent probe, characterized in that: The molar ratio of the rhodamine B hydrazide to the intermediate R is 3.1:

1.

6. A novel rhodamine Al according to claim 2 3+ A method for preparing a fluorescent probe, characterized in that: The molar concentration of the intermediate R in anhydrous ethanol is 0.02 mol / L.

7. A novel rhodamine Al according to claim 2 3+ A method for preparing a fluorescent probe, characterized in that: The reflux boiling reaction time is 14 to 16 hours.

8. A novel rhodamine Al according to claim 2 3+ A method for preparing a fluorescent probe, characterized in that: The solvent for the recrystallization is methanol and ethanol in a volume ratio of 1:

2.

9. The novel rhodamine Al according to claim 1 3+ Application of fluorescent probes in environmental monitoring, biomedicine, and agriculture.

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