CN108129487B

CN108129487B - Coumarin thiophenol fluorescent probe and preparation method thereof

Info

Publication number: CN108129487B
Application number: CN201611089946.5A
Authority: CN
Inventors: 张大同; 潘韵霖; 郝雪
Original assignee: Qilu University of Technology
Current assignee: DOUHUANGJIN FOOD Co.,Ltd.
Priority date: 2016-12-01
Filing date: 2016-12-01
Publication date: 2020-07-03
Anticipated expiration: 2036-12-01
Also published as: CN108129487A

Abstract

The invention discloses a fluorescent probe for specifically identifying thiophenol and a preparation method thereof. The probe is obtained by catalyzing 2,3,6, 7-tetrahydro-10-hydroxy-1H, 5H-quinolizino (9,1-GH) coumarin and 2, 4-dinitrofluorobenzene in an organic solvent by alkali. In an acetonitrile/PBS buffer system, the probe selectively reacts with thiophenol to release a strong fluorescent substance 2,3,6, 7-tetrahydro-10-hydroxy-1H, 5H-quinolizino- (9,1-GH) coumarin, so that the reaction system has a strong fluorescent signal at the wavelength of 519 nm. Experimental results show that the probe can specifically identify thiophenol, has the characteristics of low detection limit (only 36nmol/L), large Stokes displacement (128 nm) and the like, and can realize qualitative and quantitative detection of thiophenol by applying the fluorescence enhancement response of the probe to thiophenol. The probe can be obtained by chemical synthesis, the synthesis process is simple and easy to implement, and the detection method is simple to operate, high in sensitivity and good in selectivity, so that the probe has practical application value in the biological field.

Description

Coumarin thiophenol fluorescent probe and preparation method thereof

Technical Field

The invention relates to preparation of a coumarin structure-based thiophenol fluorescent probe and selective fluorescence detection of thiophenol by using the coumarin structure-based thiophenol fluorescent probe, and belongs to the field of organic small-molecule fluorescent probes.

Background

Thiophenol belongs to thiol compounds, is colorless liquid, has special odor, and is an important chemical raw material. However, thiophenol has great toxicity and half lethal dose of 0.01 mmol/L-0.4 mmol/L to fish. Exposure to thiophenol liquids or gases in humans can cause serious central nervous system damage and other related system injuries, including shortness of breath, muscle weakness, nausea, vomiting, coma and even death. Thiophenols are exposed to open fire, high heat or contact with oxidizing agents, which can cause the risk of combustion and explosion. Considering the harm of the thiophenol to the environment and the human health, the development of a simple and effective method for detecting the thiophenol has important significance.

Disclosure of Invention

A thiophenol fluorescent probe based on a 2,3,6, 7-tetrahydro-10-hydroxy-1H, 5H-quinolizino (9,1-GH) coumarin structure is as follows:

。

the preparation method of the probe comprises the following steps:

the probe is synthesized by taking 2,3,6, 7-tetrahydro-10-hydroxy-1H, 5H-quinolizino (9,1-GH) coumarin and 2, 4-dinitrofluorobenzene as raw materials through base catalysis in an organic solvent. Wherein 2,3,6, 7-tetrahydro-10-hydroxy-1H, 5H-quinolizino (9,1-GH) coumarin: 2, 4-dinitrofluorobenzene: reacting at a base molar ratio of 1: 1-1.5: 2-3, wherein the base is potassium carbonate or triethylamine; the organic solvent is dichloromethane, acetonitrile or tetrahydrofuran; the reaction temperature is room temperature; the reaction time is 6-12 hours. The reaction equation is as follows:

。

the thiophenol fluorescent probe of the invention has the following remarkable advantages: (1) has a novel structure; (2) thiophenols can be specifically detected in a buffer system; (3) the detection limit is low (only 36nmol/L), and the Stokes displacement is large (128 nm); (4) the reaction with thiophenol shows fluorescence enhancement, and qualitative and quantitative detection can be carried out on the thiophenol; (5) the preparation method has easily obtained raw materials, and the product is easy to separate and purify.

Drawings

FIG. 1 shows acetonitrile/PBS buffer solution (10 mmol/L, pH = 7.4) of the fluorescent probe prepared in example 1 (the volume ratio of acetonitrile to PBS buffer solution is 3:7, the probe concentration is 10 μmol/L) and fluorescence emission spectra (p-methoxythiophenol, thiophenol, p-aminophenylthiophenol concentration is 200 μmol/L; other analytes concentration is 500 μmol/L, including potassium fluoride, sodium chloride, potassium bromide, potassium iodide, sodium carbonate, sodium acetate, sodium nitrate, sodium sulfide, sodium sulfite, sodium sulfate, potassium thiocyanate, sodium thiosulfate, sodium dithionite, sodium nitrite, hydrogen peroxide, thioglycolic acid, ethanethiol, alanine, glycine, cysteine, glutathione, homocysteine), the ordinate represents the fluorescence intensity and the abscissa represents the wavelength. The excitation wavelength was 395 nm.

FIG. 2 is a graph showing acetonitrile/PBS buffer (10 mmol/L, pH = 7.4) solution (acetonitrile/PBS buffer volume ratio of 3:7, probe concentration of 10. mu. mol/L) of the fluorescent probe prepared in example 1 for different analytes (1 probe; 2 potassium fluoride; 3 sodium chloride; 4 potassium bromide; 5 potassium iodide; 6 sodium carbonate; 7 sodium acetate; 8 sodium nitrate; 9 sodium sulfide; 10 sodium sulfite; 11 sodium sulfate; 12 potassium thiocyanate; 13 sodium thiosulfate; 14 sodium dithionite; 15 sodium nitrite; 16 hydrogen peroxide; 17 thioglycolic acid; 18 ethanethiol; 19 alanine; 20 glycine; 21 cysteine; 22 glutathione; 23 homocysteine; 24-p-methoxyphenylphenol; 25 phenylphenol; 26-p-aminophenol, wherein the concentration of 24-p-methoxyphenylphenol, 25 phenylphenol, 26-p-aminophenol is 200. mu. mol/L, concentration of other analytes is 500 [ mu ] mol/L) in 519nm, excitation wavelength is 395 nm. In fig. 2, the ordinate represents fluorescence intensity and the abscissa represents different analytes; the first column 1 from the left indicates no addition of analyte; the left side of each subsequent group of columns shows the fluorescence intensity of the solution only in the presence of the analyte, and the right side shows the fluorescence intensity of the solution in the presence of both 500 mu mol/L of the analyte and 200 mu mol/L of thiophenol; the last three columns represent the fluorescence intensity of the solution in the presence of 200 μmol/L methoxythiophenol (24), thiophenol (25), p-aminophenol (26). The excitation wavelength was 395 nm.

FIG. 3 is a graph showing the change of fluorescence intensity at 519nm with time of a solution of different concentrations of thiophenol added to a solution of the fluorescent probe in acetonitrile/PBS buffer (10 mmol/L, pH = 7.4) (volume ratio of acetonitrile to PBS buffer 3:7, probe concentration 10. mu. mol/L), with fluorescence intensity on the ordinate and time on the abscissa. The excitation wavelength was 395 nm.

FIG. 4 is a fluorescence response spectrogram of acetonitrile/PBS buffer solution (10 mmol/L, pH = 7.4) (the volume ratio of acetonitrile to PBS buffer solution is 3:7, the probe concentration is 10 mu mol/L) of the fluorescent probe at 50 minutes after adding thiophenol with different concentrations, wherein the thiophenol concentration is 0-250 mu mol/L, the ordinate represents the fluorescence intensity, and the abscissa represents the wavelength. The excitation wavelength was 395 nm.

FIG. 5 is a scatter diagram showing the change of the solution fluorescence intensity of the acetonitrile/PBS buffer solution (10 mmol/L, pH = 7.4) (the volume ratio of acetonitrile to PBS buffer solution is 3:7, the probe concentration is 10. mu. mol/L) solution of the fluorescent probe at 519nm along with the thiophenol concentration (0-250. mu. mol/L), the ordinate represents the fluorescence intensity, and the abscissa represents the thiophenol concentration. The excitation wavelength was 395 nm.

FIG. 6 is a graph showing the change of fluorescence intensity at 519nm after the solution of acetonitrile/PBS buffer (10 mmol/L) (volume ratio of acetonitrile to PBS buffer 3:7, probe concentration 10 μmol/L) of the fluorescent probe is acted on 200 μmol/L thiophenol for 50 minutes under different pH conditions, the ordinate represents the fluorescence intensity, and the abscissa represents the pH. The excitation wavelength was 395 nm.

Detailed Description

Synthesis of thiophenol fluorescent probe of 2,3,6, 7-tetrahydro-10-hydroxy-1H, 5H-quinolizino (9,1-GH) coumarin structure:

example 1: and (3) synthesizing a probe. 50.0mg (0.20mmol) 2,3,6, 7-tetrahydro-10-hydroxy-1H, 5H-quinolizino (9,1-GH) coumarin are dissolved in 3ml dichloromethane and 58.0mg (0.40 mmol) K are added₂CO₃37.2mg (0.20mmol) of 2, 4-dinitrofluorobenzene, stirring at room temperature for reaction, tracking the reaction by TLC, completely reacting for 12 hours, extracting by ethyl acetate, drying an organic phase by anhydrous sodium sulfate, filtering, evaporating the solvent under reduced pressure, and separating and purifying by silica gel column chromatography to obtain 62.3 deep red solid with the yield of 73.6 percent.¹H NMR (400 MHz, CDCl₃): δ 8.86(s, 1 H), 8.32 (d,J= 6.8 Hz, 1 H),7.53 (s, 1 H), 7.10 (d,J= 9.2 Hz, 1 H), 6.88 (s, 1 H), 3.29-3.33 (m, 4 H),2.77-2.90 (m, 4 H), 1.99-2.09 (m, 4 H).¹³C NMR (100 MHz, CDCl₃): δ 157.13,155.44, 150.25, 146.30, 141.65, 138.64, 133.16, 131.39, 128.69, 125.10,122.25, 119.76, 117.64, 106.90, 50.05, 49.65, 27.53, 21.25, 20.36. HRMS (ESI)(C₁₃H₁₅NO₃) m/z: calculated for [M+H]⁺: 424.1145. Found [M+H]⁺: 424.1139.

Example 2: and (3) synthesizing a probe. 50.0mg (0.20mmol) of 2,3,6, 7-tetrahydro-10-hydroxy-1H, 5H-quinolizino (9,1-GH) coumarin was dissolved in 3mL of dichloromethane and 0.06mL (0.40 mmol) of Et was added₃N and 37.2mg (0.20mmol) of 2, 4-dinitrofluorobenzene are stirred for reaction at room temperature, TLC is used for tracking reaction, the reaction is completed within 10 hours, ethyl acetate is used for extraction, an organic phase is dried by anhydrous sodium sulfate, the solvent is removed by evaporation under reduced pressure, and the deep red solid of 73.9mg is obtained by silica gel column chromatography separation and purification, and the yield is 87.4%.

Example 3: and (3) synthesizing a probe. 50.0mg (0.20mmol) of 2,3,6, 7-tetrahydro-10-hydroxy-1H, 5H-quinolizino (9,1-GH) coumarin are dissolved in 3mL of acetonitrile and 0.09mL (0.60 mmol) of Et is added₃N and 37.2mg (0.20mmol) of 2, 4-dinitrofluorobenzene are stirred for reaction at room temperature, TLC tracking reaction is carried out, the reaction is completed within 8 hours, ethyl acetate is used for extraction, an organic phase is dried by anhydrous sodium sulfate, the solvent is removed by evaporation under reduced pressure, and the reddish brown solid with the yield of 82.2 percent is obtained by silica gel column chromatography separation and purification.

Example 4: and (3) synthesizing a probe. 50.0mg (0.20mmol) of 2,3,6, 7-tetrahydro-10-hydroxy-1H, 5H-quinolizino (9,1-GH) coumarin are dissolved in 3mL of acetonitrile and 0.09mL (0.60 mmol) of Et is added₃N and 55.8mg (0.30mmol) of 2, 4-dinitrofluorobenzene are stirred at room temperature for reaction, TLC tracking reaction is carried out, the reaction is completed within 6 hours, ethyl acetate is used for extraction, an organic phase is dried by anhydrous sodium sulfate, the solvent is removed by evaporation under reduced pressure, and the red brown solid of 74.5mg is obtained by silica gel column chromatography separation and purification, and the yield is 88.0%.

Example 5: and (3) synthesizing a probe. 50.0mg (0.20mmol) of 2,3,6, 7-tetrahydro-10-hydroxy-1H, 5H-quinolizino (9,1-GH) coumarin are dissolved in 3mL of tetrahydrofuran and 0.09mL (0.60 mmol) of Et are added₃N and 55.8mg (0.30mmol) of 2, 4-dinitrofluorobenzene are stirred at room temperature for reaction, TLC is used for tracking reaction, the reaction is completed within 8 hours, ethyl acetate is used for extraction, an organic phase is dried by anhydrous sodium sulfate, the solvent is removed by evaporation under reduced pressure, and the reddish brown solid with the yield of 82.0 percent is obtained by silica gel column chromatography separation and purification.

Example 6: the application study of the photophysical detection of the thiophenol by the fluorescent probe based on the 2,3,6, 7-tetrahydro-10-hydroxy-1H, 5H-quinolizino (9,1-GH) coumarin structure.

The fluorescent probe based on the 2,3,6, 7-tetrahydro-10-hydroxy-1H, 5H-quinolizino (9,1-GH) coumarin structure prepared in example 1 was dissolved in 3mL of acetonitrile to prepare a stock solution having a concentration of 1mmol/L, and then a probe test solution having a concentration of 10. mu. mol/L was prepared using acetonitrile/PBS buffer (10 mmol/L, pH = 7.4) (the volume ratio of acetonitrile to PBS buffer is 3: 7). The probe solution has weaker fluorescence emission at 519nm under the excitation of 395nm, when thiophenol, p-aminophenol or p-methoxythiophenol is added into the probe solution, the fluorescence intensity is obviously enhanced, and the Stokes displacement reaches 128 nm; the adding amount of thiophenol is within 0-50 [ mu ] mol/L, the fluorescence intensity of the probe solution at 519nm and the concentration of thiophenol form good linearity, and the detection limit is only 36 nmol/L; no significant change in fluorescence intensity of the probe solution was caused after addition of other common analytes.

As can be seen from fig. 1, after adding 500 μmol/L of other analytes to the solution of the probe, including potassium fluoride, sodium chloride, potassium bromide, potassium iodide, sodium carbonate, sodium acetate, sodium nitrate, sodium sulfide, sodium sulfite, sodium sulfate, potassium thiocyanate, sodium thiosulfate, sodium dithionite, sodium nitrite, hydrogen peroxide, thioglycolic acid, ethanethiol, alanine, glycine, cysteine, glutathione, homocysteine, fluorescence is not significantly enhanced; after adding thiophenol or p-aminophenol or p-methoxy thiophenol, the fluorescence emission intensity of the probe solution is obviously enhanced.

As can be seen from fig. 2, when 500 μmol/L of other analytes are present in a 10 μmol/L acetonitrile/PBS buffer solution (10 mmol/L, pH = 7.4) (the volume ratio of acetonitrile to PBS buffer solution is 3:7) of the probe, including potassium fluoride, sodium chloride, potassium bromide, potassium iodide, sodium carbonate, sodium acetate, sodium nitrate, sodium sulfide, sodium sulfite, sodium sulfate, potassium thiocyanate, sodium thiosulfate, sodium dithionite, sodium nitrite, hydrogen peroxide, thioglycolic acid, ethanethiol, alanine, glycine, cysteine, glutathione, homocysteine, 200 μmol/L of thiophenol is added to the solution, and the column height indicates that the response of the probe to thiophenol cannot be interfered under the presence of other analytes, and the probe exhibits good anti-interference capability.

As can be seen from FIG. 3, when different concentrations of thiophenol were added to the probe solution, the fluorescence intensity of the solution increased with time, with the higher concentration increasing more rapidly and the lower concentration increasing more slowly.

As can be seen from FIGS. 4 and 5, when the probe concentration is 10 μmol/L, the fluorescence intensity gradually increases with the increase of the thiophenol concentration; the concentration of thiophenol is within 0-50 mu mol/L, and the fluorescence intensity of the probe solution at 519nm and the concentration of thiophenol form good linearity.

As can be seen from FIG. 6, the probe has a significant pH dependence on the detection of thiophenol.

It is concluded that the fluorescent probe based on the 2,3,6, 7-tetrahydro-10-hydroxy-1H, 5H-quinolizino (9,1-GH) coumarin structure prepared by the invention has high selectivity and sensitivity on thiophenol, and can be used for qualitative and quantitative detection of thiophenol.

Claims

1. The application of a 2,3,6, 7-tetrahydro-10-hydroxy-1H, 5H-quinolizino (9,1-GH) coumarin structural fluorescent probe is characterized in that: the method can realize the selective qualitative and quantitative detection of the thiophenol in a water-containing system;

the aqueous system is acetonitrile/PBS buffer solution, the concentration of the PBS buffer solution is 10mmol/L, the pH is 7.4, and the volume ratio of acetonitrile to the PBS buffer solution is 3: 7;

the structural formula of the 2,3,6, 7-tetrahydro-10-hydroxy-1H, 5H-quinolizino (9,1-GH) coumarin fluorescent probe is as follows:

。