CN113651828B

CN113651828B - Near-infrared fluorescent probe for detecting chromium ions and aluminum ions as well as preparation method and application of near-infrared fluorescent probe

Info

Publication number: CN113651828B
Application number: CN202110975505.XA
Authority: CN
Inventors: 许志红; 周起航; 杨莉; 刘高兵; 杨涵熙; 许泗林; 许锐杰; 高志颖; 段颖颖; 潘明霞
Original assignee: Xuchang University
Current assignee: Xuchang University
Priority date: 2021-08-24
Filing date: 2021-08-24
Publication date: 2022-05-17
Anticipated expiration: 2041-08-24
Also published as: CN113651828A

Abstract

The invention belongs to the technical field of organic synthesis, and particularly relates to a near-infrared fluorescent probe for detecting chromium ions and aluminum ions, a preparation method and application thereof, wherein the fluorescent probe takes rhodamine hemicyanine as a fluorophore and 8-hydroxyquinoline-2-formaldehyde as a recognition group, so that the specific recognition response to the chromium ions and the aluminum ions is realized, and the probe has the following structure:

Description

Near-infrared fluorescent probe for detecting chromium ions and aluminum ions as well as preparation method and application of near-infrared fluorescent probe

Technical Field

The invention belongs to the technical field of organic synthesis, and particularly relates to a near-infrared fluorescent probe for detecting chromium ions and aluminum ions, and a preparation method and application thereof.

Background

The application of the metal ions in our daily life is very wide, such as in our heavy industry, light industry, textile industry, food processing and drug production processes, and great convenience is brought to our life and production. We live for a long time in an environment associated with metal ions. Different metal ions are accumulated in a human body through a food chain, and a series of diseases caused by the metal ions, such as growth retardation, dysmnesia, cancer and the like, can be caused after the accumulation. Therefore, health problems caused by metal ions are also drawing increasing attention.

Al³⁺The food packaging box is particularly common in the living environment of people, such as kitchen ware of our home, food additives, food packaging, medicines and the like. There are many times when Al is intentionally and unintentionally taken in³⁺，Al³⁺Can stay in cells and tissues for a longer time and then be excreted by the body. However, too much intake, Al³⁺Easily gather in the body, thereby possibly causing various diseases including osteomalacia, breast cancer and the like, Al³⁺(also known as neurotoxic agents) also affect the central nervous system, Parkinson's disease, Alzheimer's disease, and the like. World Health Organization (WHO) published weekly human pairs of Al³⁺The intake was made at 7 mg/kg. Thus, development of detection of Al³⁺There is a great need for a fluorescent sensor that is highly sensitive and applicable to biological organisms.

Cr³⁺Plays an important role in the agricultural and industrial production (battery production, electroplating, alloy manufacturing, leather tanning and the like). At the same time Cr³⁺And also become one of the major environmental pollutants. Cr (chromium) component³⁺Is one of the trace elements necessary for the organism to maintain normal life activities, and participates in various physiological processes of the organism, such as protein synthesis, nucleic acid metabolism and the like. Cr (chromium) component³⁺The increase or decrease of the content can affect the normal physiological activity and increase the risk of diseases. Due to Cr³⁺The paramagnetic property of the material is easy to quench fluorescence, and the fluorescence quenching is not easy to identify the output of a fluorescence signal, so that the fluorescence imaging of cells is interfered, and the detection of Cr is limited³⁺Development of the probe of (1). In particular, the development of Cr which can be used at the subcellular level in organisms³⁺It is difficult and challenging for a near infrared fluorescence sensor to be fluorescence-enhanced.

At present, Al is aimed at³⁺Or Cr³⁺The fluorescence sensor of (1), the fluorescence sensor that excites and emits in the near infrared region is rare. Thus developingDesigned for detecting Al³⁺Or Cr³⁺The near infrared fluorescence sensor utilizes the characteristics of near infrared, and the research on applying the near infrared fluorescence sensor to the fluorescence imaging of cells or organisms is necessary, so that the near infrared fluorescence sensor can be better applied to medicine and biology.

Disclosure of Invention

In order to overcome the problems in the prior art, the invention provides a near-infrared fluorescent probe for detecting chromium ions and aluminum ions, which realizes specific recognition response to the chromium ions and the aluminum ions, is not influenced by interfering ions, and can judge whether the chromium ions and the aluminum ions exist or not by direct visual observation.

The invention also provides a preparation method and application of the near-infrared fluorescent probe.

In order to achieve the purpose, the technical scheme of the invention is as follows:

a near-infrared fluorescent probe for detecting chromium ions and aluminum ions, the molecular formula of the probe is C₄₈H₄₇O₅N₄The structural formula is as follows:

。

the synthesis route for detecting the chromium ions and the aluminum ions is as follows:

the preparation method specifically comprises the following steps:

(1) dissolving cyclohexanone in concentrated sulfuric acid, adding LF-1, stirring at 92-98 ℃ for reaction till the reaction is complete, cooling to room temperature, adding perchloric acid, performing suction filtration, washing with water, and drying to obtain a compound LF-2;

(2) adding acetic anhydride into LF-2 and Fischer aldehyde under the protection of nitrogen, stirring at 50 +/-5 ℃ until the mixture is complete, then adding ice water into the mixed solution, extracting with dichloromethane and water, extracting an organic phase with a saturated sodium bicarbonate solution, finally washing the organic phase with deionized water, drying to obtain a crude product, and separating the crude product by using column chromatography to obtain a pure compound LF-3;

(3) under the protection of nitrogen, dichloromethane is added into a compound LF-3, a Kate condensing agent (BOP) is dissolved in dichloromethane and added into methane solution of LF-3, then hydrazine hydrate is added into reaction liquid, the reaction liquid is stirred at room temperature until the reaction is completed, dichloromethane and water are used for extraction, drying is carried out, a crude product LF-4 is obtained, and the obtained crude product is separated by column chromatography to obtain a pure compound LF-4;

(4) and adding a compound LF-4 and 8-hydroxyquinoline-2-formaldehyde into ethanol, refluxing at 100 +/-5 ℃ until the reaction is complete, separating out a solid after the reaction is finished, washing with absolute ethyl alcohol, and drying to obtain the fluorescent probe.

Preferably, the molar ratio of LF-1, cyclohexanone and perchloric acid in the step (1) is 1 (2-3) to (10-16).

Preferably, the molar ratio of the compound LF-2 to the Fischer aldehyde in the step (2) is 1 (1.1-1.2).

Preferably, in the step (3), the molar ratio of the compound LF-3 to BOP to hydrazine hydrate is 1 (1.1-1.3) to (5-12).

Preferably, the molar ratio of the compound LF-4 to the 8-hydroxyquinoline-2-carbaldehyde in the step (4) is 1 (1.1-1.3).

The near-infrared fluorescent probe is applied to the fluorescent detection of chromium ions and aluminum ions, and is particularly used for the fluorescent detection, visual qualitative detection and cell imaging detection of the contents of the chromium ions and the aluminum ions.

Compared with the prior art, the invention has the beneficial effects that:

1. after the probe is added with chromium ions and aluminum ions, the fluorescence intensity is obviously enhanced and is accompanied with obvious eye color change, the selected interference ions and the like almost have no influence on the detection effect, and the specific identification response to the chromium ions and the aluminum ions is realized.

2. The detection limit of the near-infrared fluorescent probe of the invention is Al³⁺Has a detection limit of 0.9023 μ M, Cr of³⁺The detection limit of (2) is 0.594. mu.M.

Drawings

FIG. 1 is a nuclear magnetic hydrogen spectrum of the fluorescent probe prepared in example 1;

FIG. 2 is a nuclear magnetic carbon spectrum of the fluorescent probe prepared in example 1;

FIG. 3 is a fluorescence spectrum of the fluorescent probe prepared in example 1 after reacting with various ions;

FIG. 4 is a graph showing fluorescence intensities of the fluorescent probes prepared in example 1 after reacting with different ions;

FIG. 5 shows the fluorescent probe prepared in example 1 and Al³⁺、Cr³⁺Fluorescence spectra of the reaction over time; wherein (a) is a fluorescent probe pair Al³⁺The fluorescence spectrum of the reaction changes with time, (b) is the fluorescent probe to Cr³⁺Fluorescence spectra of the reaction over time;

FIG. 6 shows fluorescent probes prepared in example 1 with different Al concentrations³⁺、Cr³⁺Fluorescence spectra after ion response; wherein (a) is fluorescent probe for different concentrations of Al³⁺Fluorescence spectrum after ion response, and (b) fluorescence probe for different concentrations of Cr³⁺Fluorescence spectra after ion response;

FIG. 7 is the fluorescence intensity of the solution with Al³⁺、Cr³⁺A linear relationship between ion concentrations; wherein (a) is a fluorescent probe pair Al³⁺Is a linear relationship diagram of (b) is a fluorescent probe pair Cr³⁺A linear relationship graph;

FIG. 8 shows the reaction of the fluorescent probe prepared in example 1 with Al in different pH system solutions³⁺、Cr³⁺Fluorescent light after ion response

Spectra. Wherein (a) is a fluorescent probe pair to Al at different pH³⁺A spectrogram of fluorescence intensity change after ion response, and (b) a pair of fluorescence probes for Cr under different pH values³⁺And (3) a fluorescence intensity change spectrogram after ion response.

Detailed Description

The invention is further illustrated, but not limited, by the following examples and the accompanying drawings.

Example 1

The preparation method of the fluorescent probe comprises the following steps:

(1) preparation of Compound LF-2

8 mL of concentrated sulfuric acid (commercially available 98% concentrated sulfuric acid) was placed in a 50 mL pear-shaped flask and stirred at 0 ℃ for 20 min. Slowly injecting cyclohexanone (0.2941 g, 3.0 mmol) into a bottle, stirring for 15-20 min, adding LF-1 (0.4691 g, 1.5 mmol) into the reaction solution in small amount for multiple times, controlling the addition within 30 min, gradually changing the color of the solution from colorless to red, heating at 92 ℃ for 4 h after the solid is completely dissolved, stopping heating after the reaction is completely finished, cooling to room temperature, slowly pouring the reaction solution into a beaker filled with 480mL of ice water, stirring, and dropwise adding 1.5 mL of HClO₄Accompanied by precipitation of a red solid, the precipitate was filtered with suction, washed with cold water and dried. 0.5462 g of a red solid was finally obtained (yield: 95.6%), which was directly subjected to the next step without purification;

(2) preparation of Compound LF-3

LF-2 (0.3262 g, 0.8 mmol), Fisher's aldehyde (0.1812 g, 0.9 mmol) were placed in a 50 mL two-necked pear-shaped flask, nitrogen was added, acetic anhydride (5 mL) was added, stirring was carried out at 50 ℃ for 2 hours, ice water (typically 2-3 times the volume of acetic anhydride) was added to the mixture, extraction was carried out with dichloromethane and water, the organic phase was extracted three times with saturated sodium bicarbonate solution, washed three times with deionized water, and Na was added₂SO₄Drying to obtain a crude product, and separating and purifying by using a chromatographic column, wherein an eluent is dichloromethane: methanol = 20:1, solvent was spin dried, dried to yield 0.2564 g of green solid (55.3% yield).¹H NMR (400 MHz, CDCl₃) δ 8.24- 8.15 (m, 1H), 8.10 (d, J = 13.2 Hz, 1H), 7.55-7.44 (m, 2H), 7.28 (s, 1H), 7.03 (q, J = 6.3, 4.4 Hz, 2H), 6.86 (d, J = 8.4 Hz, 2H), 6.55 (d, J = 9.2 Hz, 1H), 6.45 (s, 1H), 5.63 (d, J = 13.2 Hz, 1H), 5.29 (s, 1H), 3.45 (t, J = 7.3 Hz, 4H), 3.36 (s, 3H), 2.58 (tq, J = 16.3, 9.7, 7.5 Hz, 2H), 2.42 (dt, J = 14.1, 5.4 Hz, 1H), 2.10 (dd, J = 16.3, 6.3 Hz, 1H), 1.98 (s, 1H), 1.84 (dd, J = 13.4, 6.4 Hz, 1H), 1.23 (t, J = 7.1 Hz, 6H).

(3) Preparation of Compound LF-4

LF-3 (0.2018 g, 0.36 mmol) was placed in a two-necked flask, nitrogen blanketed, 4 mL of methylene chloride was injected into the flask, the Kate condensing agent (BOP, 0.43 mmol) was dissolved in methylene chloride, slowly injected into the flask, and stirred for 20-30 minutes, then 3mL of hydrazine hydrate was injected into the flask via syringe, and stirred vigorously (1800 + 2500 rpm) at room temperature. The solution changes from green to yellow or red with the addition of hydrazine hydrate, and after stirring for 3 hours at room temperature, the solution is extracted three times with dichloromethane and water, and then Na is added₂SO₄Drying to obtain crude product, and separating and purifying with chromatographic column to obtain eluent only using dichloromethane. The product was collected and dried to yield 0.1624 g of a yellow solid (76.18% yield).¹H NMR (400 MHz, CDCl₃) δ 7.93-7.85 (m, 1H), 7.56-7.38 (m, 3H), 7.23-7.11 (m, 3H), 6.65-6.58 (m, 1H), 6.40-6.23 (m, 3H), 5.37 (d, J = 12.6 Hz, 1H), 3.65 (s, 2H), 3.35 (q, J = 7.1 Hz, 4H), 3.15 (s, 3H), 2.65-2.41 (m, 2H), 2.04 (s, 1H), 1.71 (d, J = 6.4 Hz, 6H), 1.37 (d, J = 1.1 Hz, 1H), 1.26 (t, J = 7.1 Hz, 2H), 1.18 (t, J = 7.0 Hz, 6H)。

(4) Preparation of Probe LFK

Compound LF-4 (0.1211 g, 0.21 mmol) was weighed into a 50 mL pear-shaped flask, then 8-hydroxyquinoline-2-carbaldehyde (0.0401 g, 0.23 mmol) was added to the flask, and 8.5 mL absolute ethanol was added, the temperature was set to reflux at 105 ℃, after the reaction was over 6 hours, cooled to room temperature, a tan solid precipitated, which was filtered with suction and washed several times with absolute ethanol. The solid was collected and placed in a drying oven for drying. The product was 120.5 mg of a tan solid (yield 78.8%), and its hydrogen and carbon spectra are shown in FIGS. 1 and 2.¹H NMR (400 MHz，CDCl3) δ 13.76 (s, 1H), 8.85 (s, 1H), 8.74-8.67 (m, 1H), 8.58 (dd, J = 7.3, 1.2 Hz, 1H), 8.34 (s, 1H), 7.99 (d, J = 7.5 Hz, 1H), 7.70-7.64 (m, 1H), 7.62 (s, 1H), 7.61-7.55 (m, 2H), 7.51 (td, J = 7.5, 1.1 Hz, 1H), 7.24-7.18 (m, 2H), 6.88 (t, J = 7.4 Hz, 1H), 6.64 (d, J = 7.8 Hz, 1H), 6.50 (s, 1H), 6.47 (s, 1H), 6.45 (d, J = 2.6 Hz, 1H), 6.28 (dd, J = 8.9, 2.6 Hz, 1H), 5.39 (d, J = 12.7 Hz, 1H), 3.36 (qd, J = 7.2, 3.9 Hz, 4H), 3.16 (s, 3H), 2.49 (s, 2H), 1.70-1.64 (m, 2H), 1.60 (s, 6H), 1.44-1.38 (m, 2H), 1.19 (t, J = 7.0 Hz, 6H),¹³C NMR (101 MHz，CDCl3 ) δ 164.36，163.92，161.89，158.23，152.30，150.31，149.19，148.44，147.69，145.32，138.98，134.16，133.82，132.54，130.03，128.73，128.70，127.71，127.47，125.90，123.59，123.41，122.32，121.59，120.57，119.61，119.40，113.42，108.61，105.77，103.99，102.57，98.14，92.05，77.22，67.93，45.60，44.31，40.07，30.25，29.15，28.70，28.41，25.22，22.97，22.13，20.36，13.84，12.64 .

The application test of the near-infrared fluorescent probe prepared by the experiment is as follows:

(1) preparation of stock solution for detection

a. The concentration is 1.00X 10^-3Preparing mol/L probe LFK stock solution: the probe LFK is weighed 7.27 mg by ten-thousandth of an electronic balance, and then the weighed probe LFK is dissolved in 10 mL of acetonitrile, and the solution is heated and shaken well and sonicated at 500W for 8 minutes.

b. Various cations (zinc chloride, copper chloride dihydrate, magnesium chloride, cadmium chloride, ferric chloride hexahydrate, aluminum trichloride, cobalt chloride hexahydrate, chromium chloride hexahydrate, calcium chloride, nickel chloride hexahydrate, silver nitrate, lithium chloride, barium chloride, manganese chloride, potassium chloride, mercury chloride) were formulated to 1.00X 10 with deionized water^-2mol/L solution, and diluting to 1.00X 10^-3 mol/L。

The buffer solution used in the following detection is a mixed solution of deionized water and absolute ethyl alcohol in a volume ratio of 4: 6.

(2) Detection assay

3mL of the buffer was added to a solution containing 15uL of 1.0X 10^-3Adding 10 equivalents of the above various ion stock solutions into mol/L probe stock solution,the fluorescence spectrum was detected, and the results are shown in fig. 3, wherein the fluorescence parameters were set to 5nm for excitation slit width, 2.5nm for emission slit width, and Ex = 660nm for excitation wavelength. As can be seen from FIG. 3, Al is added separately³⁺、Cr³⁺The fluorescence spectrum shows that the fluorescence intensity of the probe is obviously enhanced, and the fluorescence intensity is not obviously changed after other ions are added, so that the probe is used for Al³⁺、Cr³ ⁺There is good selectivity for ions, but no specific selectivity for other ions. The fluorescence intensity of the fluorescent probe after reacting with different ions is shown in FIG. 4. As can be seen from FIG. 4, the fluorescent probe reacts with Al³⁺、Cr³⁺The fluorescence intensity after reaction is obviously higher than that of a blank sample and other interfering ions, so that the probe can be used for detecting Al with high selectivity³⁺、Cr³⁺The fluorescence-enhanced near-infrared probe of (1).

3mL of the buffer was added to a solution containing 15uL of 1.0X 10^-3The mol/L stock solution of the probe was added with 15uL of 1.0X 10^-2mol/L of Al³⁺、Cr³⁺Solution, the time change of the fluorescence spectrum was detected as shown in FIG. 5. The excitation wavelength Ex = 660nm, the excitation slit width is 5nm, the emission slit width is 2.5nm, and the scanning range is 680-900 nm. FIGS. 5 (a) and (b) show the probes LFK and Al, respectively³⁺、Cr³⁺In response to the change pattern, both of which changed from colorless to green, the fluorescence intensity reached the maximum almost instantaneously, and the reaction was substantially completed in 1 minute. The fluorescence intensity does not change with time basically, and the probes LFK and Al are proved³⁺、Cr³⁺The reaction is rapid and stable.

3mL of the buffer was added to a solution containing 15uL of 1.0X 10^-3Adding Al with different equivalent weight into mol/L probe stock solution³⁺、Cr³⁺The solution was shaken well (0.1 eq, 0.2eq, 0.3 eq, 0.4 eq, 0.5 eq, 0.6 eq, … … 3.5.5 eq, 3.6 eq, 3.7 eq, 3.8 eq, 3.9 eq, 4.0 eq, respectively, based on the molar amount of the probe) to detect the change in fluorescence intensity. As shown in FIG. 6, with Al³⁺、Cr³⁺The fluorescence intensity gradually increased with increasing concentration, and FIG. 6 (a) and (b) are the probe and Al with different concentrations³⁺、Cr³⁺The fluorescence spectrum of (2). When Al is present³⁺、Cr³⁺The concentration is 0-4 × 10^-2mol/L, fluorescence intensity of solution and Al³⁺、Cr³⁺There was a good linear relationship between the ion concentrations, and the results are shown in fig. 7. By linear fitting, a corresponding one-dimensional linear equation can be obtained, wherein FIG. 7 (a) probes for Al³⁺For y =196.82161+157.67917x, the correlation coefficient squared (R) is obtained by calculation²) 0.99176, indicating a good fit. The LOD detection limit is finally calculated from the formula LOD =3 σ/k (σ is the standard deviation of 18 pure probes LFK tested consecutively), and its Al³⁺The detection limit was 0.9023. mu.M. In which FIG. 7 (b) probe is directed to Cr³⁺For y =176.26332+174.18236x, the correlation coefficient squared (R) is obtained by calculation²) 0.99786, indicating a good fit. The LOD detection limit is finally calculated from the formula LOD =3 σ/k (σ is the standard deviation of 19 pure probes LFK tested consecutively), and its Al³⁺The detection limit was 0.594. mu.M.

Adjusting the pH value of water to 3-13 by using 0.01mol/L NaOH solution or 0.01mol/L HCl solution, mixing the water solution with the pH value of 3-13 and absolute ethyl alcohol according to the volume ratio of 4:6 to prepare buffer solution, taking 3mL of the buffer solution with different pH values, adding 15uL of the buffer solution with the concentration of 1.0 multiplied by 10^-3The mol/L stock solution of the probe was added with 15uL of 1.0X 10^-2mol/L of Al³⁺、Cr³⁺And (4) dissolving, and detecting the change of the fluorescence spectrum. The results are shown in FIG. 8. As can be seen from fig. 8 (a), the fluorescence spectrum probes for Al between pH =5-8.5³⁺The response of the ions has good stability, so that the probe can detect Al in a physiological environment³⁺Ions. As can be seen from fig. 8 (b), the fluorescence spectrum probes for Al between pH =5-8.5³⁺The response of the ions has good stability, so that the probe can detect Cr in a physiological environment³⁺Ions.

Claims

1. A near-infrared fluorescent probe for detecting chromium ions and aluminum ions is characterized in that the structural formula of the probe is as follows:

。

2. the method for preparing the near-infrared fluorescent probe for detecting chromium ions and aluminum ions as claimed in claim 1, wherein the synthetic route is as follows:

the method specifically comprises the following steps:

3. The method for preparing a near-infrared fluorescent probe for detecting chromium ions and aluminum ions as claimed in claim 2, wherein the molar ratio of LF-1, cyclohexanone and perchloric acid in step (1) is 1 (2-3) to (10-16).

4. The method for preparing the near-infrared fluorescent probe for detecting chromium ions and aluminum ions according to claim 2, wherein the molar ratio of the compound LF-2 to the Fischer aldehyde in the step (2) is 1 (1.1-1.2).

5. The method for preparing a near-infrared fluorescent probe for detecting chromium ions and aluminum ions as claimed in claim 2, wherein the molar ratio of the compounds LF-3, BOP and hydrazine hydrate in step (3) is 1 (1.1-1.3) to (5-12).

6. The method for preparing the near-infrared fluorescent probe for detecting chromium ions and aluminum ions according to claim 2, wherein the molar ratio of the compound LF-4 to the 8-hydroxyquinoline-2-formaldehyde in the step (4) is 1 (1.1-1.3).

7. The use of the near-infrared fluorescent probe according to claim 1 in the fluorescent detection of chromium and aluminum ions, characterized in that it is used for the fluorescent detection, visual qualitative detection and cell imaging detection of the contents of chromium and aluminum ions.