CN107337654A - A kind of fluorescence probe for analyzing mercury ion, preparation method and application - Google Patents

Abstract

The invention relates to a fast and highly selective mercury ion fluorescent probe. It is a thiocarbonate compound, and its structural formula is . This type of probe can realize at least one of the following technical effects: highly selective recognition of mercury ions; rapid response to mercury ions; large Stokes shift; stable properties and long-term storage and use; Strong anti-interference ability.

Description

Fluorescent probe for analyzing mercury ions, preparation method and application

technical field

The invention relates to a thiocarbonate compound used as a fluorescent probe for analyzing mercury ions and a preparation method, which can rapidly and sensitively identify mercury ions with high selectivity, or can measure the concentration of mercury ions in a sample.

Background technique

Mercury is a silvery white shiny heavy liquid, chemically stable, insoluble in acid and alkali. Mercury can evaporate at room temperature, and mercury vapor and mercury compounds are highly toxic (chronic). Mercury is widely used in the manufacture of scientific measuring instruments (such as thermometers, etc.), medicines, electrodes, catalysts, etc. Mercury is ubiquitous in nature. Generally, animals and plants contain trace amounts of mercury. Therefore, there are trace amounts of mercury in our food, which can be metabolized through excretion and hair without affecting health.

Mercury is a heavy metal pollutant with strong biological toxicity in the environment. It is difficult to be excreted after entering the organism, which seriously threatens human health. The harm of mercury to the human body mainly involves the central nervous system, digestive system and kidneys, and also has certain effects on the respiratory system, skin, blood and eyes. In the past ten years, the concentration of mercury in the environment has continued to rise around the world, which has attracted great attention from governments and environmental protection organizations, and has become another global environmental problem after climate change.

In view of this, it is extremely important and meaningful to develop analytical methods that can effectively detect mercury ions. The analytical methods reported to detect mercury ions include volumetric analysis, optical analysis, ion chromatography (IC), mercury ion selective electrode method, on-line analysis and other methods. Among these numerous detection methods, fluorescent probes have become the focus of researchers because of their unique advantages. However, the currently reported fluorescent probes still have some problems, including insufficient selectivity, insufficient response speed, and complex synthesis. Due to other ions in the environment such as cadmium ions, lead ions, magnesium ions, aluminum ions, ferrous ions, ferric ions, chromium ions, calcium ions, zinc ions, sodium ions, potassium ions, cobalt ions and nickel ions, etc. Other metal ions will potentially interfere with the detection of mercury ions. Therefore, rapid development, high selectivity, high sensitivity, and simple synthesis of fluorescent probes for mercury ions are urgent problems for those skilled in the art.

Contents of the invention

There is an urgent need in the field for a simple, rapid and highly selective fluorescent probe for mercury ions, which can effectively detect mercury ions. And provide a preparation method with simple synthesis, high selectivity, high sensitivity, and rapid identification of mercury ions.

Specifically, the present invention provides a fluorescent probe for mercury ion evolution, which is a thiocarbonate compound, and its structure is shown in Formula 1:

Formula 1

Among them: R1, R2, R3, R4, R5, R6, R7, R8, R9, R10, R11, R12, R13, R14, R15 are hydrogen atoms, straight chain or branched chain alkyl groups with 1-3 carbon atoms, Straight chain or branched alkoxy, sulfonic acid, ester, carboxyl with 1-3 carbon atoms; R1, R2, R3, R4, R5, R6, R7, R8, R9, R10, R11, R12, R13 , R14 and R15 are the same or different.

Preferably, the fluorescent probe of the present invention has a structural formula as shown in formula 2:

Formula 2.

The present invention also provides the preparation method of mercury ion fluorescent probe, it is characterized in that adopting following steps:

Dissolve the hexamethylenetetramine shown in Formula 4 in 20 mL of trifluoroacetic acid, then add p-cresol shown in Formula 3, the molar ratio of the two is 4:1, and heat at reflux at 100°C for 6 hours, And shading treatment, after the reaction, the product was added to ice water, and after the solid was precipitated, suction filtration was carried out to obtain the pure product formula 5

Formula 3 Formula 4 Formula 5

Dissolve the compound shown in Formula 5 in absolute ethanol, then add aminobenzenethiol shown in Formula 6, the molar ratio of the two is 1:1, stir at room temperature, add 37% hydrochloric acid and 30% hydrogen peroxide, reacted for 30 min, then carried out suction filtration to obtain pure product formula 8

Formula 6 Formula 5 Formula 8

Dissolve the compound shown in formula 8 in methylene chloride, add N,N-diisopropylethylamine (DIPEA) for catalysis, then add phenyl thiochloroformate shown in formula 9, the molar ratio of the two is 1:3, heated to reflux at 50°C for 10h, and then filtered with suction to obtain pure product Formula 2

Formula 8 Formula 9 Formula 2.

The present invention also provides the use of the probe of the present invention in preparing a preparation for detecting mercury ion concentration in a sample.

For the above-mentioned application, first prepare the probe stock solution: weigh 5 mg of the compound shown in formula 2 and add it to the colorimetric tube, add 1 mL of dichloromethane, shake well to dissolve the probe, and then use anhydrous Dilute ethanol to 10 mL and prepare 1 mM probe stock solution;

Then, add 5 mL of absolute ethanol to the colorimetric tube, pipette 50 µL of the probe stock solution (1 mM) into the colorimetric tube, add 2-3 mL of distilled water, and then add 0.5 mL of 4-hydroxyethyl Piperazineethanesulfonic acid (HEPES), then distilled water to 10mL; pipette 200 µL mercury ion solution (1 mM) into the colorimetric tube, shake well, after 20min, use a fluorescence spectrophotometer (Horiba FluoroMax-4) Measure the fluorescence spectrum.

The mercury ion fluorescent probe of the invention can interact with mercury ions to produce changes in fluorescence spectrum, thereby realizing quantitative detection of mercury ions.

Specifically, the mercury ion fluorescent probe of the present invention is combined with cadmium ions, lead ions, magnesium ions, aluminum ions, ferrous ions, ferric ions, chromium ions, calcium ions, zinc ions, sodium ions, potassium ions, respectively. The interaction of other ions such as ions, cobalt ions, and nickel ions cannot cause a significant change in the fluorescence spectrum, thereby achieving selective identification of mercury ions, which can optionally be used to exclude these cadmium ions, lead ions, magnesium ions, and aluminum ions. , Ferric ions, ferric ions, chromium ions, calcium ions, zinc ions, sodium ions, potassium ions, cobalt ions and nickel ions interfere with the quantitative determination of mercury ions.

The Stokes shift of the mercury ion fluorescent probe of the present invention is very large, so that it is beneficial to reduce the influence of the self-absorption spectrum or the self-fluorescence spectrum on the fluorescence intensity.

Optionally, the mercury ion fluorescent probe of the present invention has good stability, and thus can be stored and used for a long time.

Furthermore, the mercury ion fluorescent probe of the present invention is a fast and highly selective mercury ion fluorescent probe, and the synthesis is simple, which is favorable for commercial promotion and application.

The beneficial effect of the invention is that the invention relates to a fast and highly selective mercury ion fluorescent probe. Specifically, the probe of the present invention is a class of thiocarbonate compounds, which can be used as mercury ion fluorescent probes for the detection of mercury ions. This type of probe can realize at least one of the following technical effects: highly selective recognition of mercury ions; rapid response to mercury ions; large Stokes shift; stable properties and long-term storage and use; Strong anti-interference ability.

Description of drawings

Figure 1 is the fluorescence spectrum of the probe (5µM) before and after adding Hg2+ (20µM).

Fig. 2 Effect of different concentrations of Hg2+ (0-40 µM) on the fluorescence spectrum of the probe (5 µM).

Figure 3 Effect of different ion analytes (20 µM) on the fluorescence intensity of the probe (5 µM).

detailed description:

The invention will be illustrated in more detail below by means of the following examples. The following examples are illustrative only, and it should be understood that the present invention is not limited by the following examples.

Embodiment 1: preparation formula 5 and formula 8 compounds

Dissolve 11.2 g of hexamethylenetetramine shown in formula 4 in 20 mL of trifluoroacetic acid, then add 2.16 g of p-cresol shown in formula 3, the molar ratio of the two is 4:1, 100 °C Heating to reflux for 6 hours, and shading treatment, after the reaction, the product was added to ice water, and after the solid was precipitated, suction filtration was performed to obtain the pure product Formula 5.

Formula 3 Formula 4 Formula 5

Take 656 mg shown in Formula 5 and dissolve in 20 mL of absolute ethanol, then add 500 mg of aminobenzenethiol shown in Formula 6, the molar ratio of the two is 1:1, stir at room temperature, and add dropwise during the stirring process 1184 mg of 37% hydrochloric acid and 2720 mg of 30% hydrogen peroxide were reacted for 30 min, and then filtered by suction to obtain the pure product Formula 8.

Formula 6 Formula 5 Formula 8

Example 2

Formula 8 Formula 9 Formula 2

(Scheme 1) Dissolve 374 mg (1 mmol) of the compound of formula 8 prepared in Example 1 in 10 mL of dichloromethane, add 129 mg (1 mmol) of DIPEA, and then add 173 mg (1 mmol) of benzene thiochloroformate The ester was refluxed at 50° C. for 10 h, and then subjected to suspension evaporation using a rotary evaporator to obtain a solid, which was the crude product of the compound represented by Formula 2. If you want to obtain a purer product, you can use a mixed system of dichloromethane and petroleum ether (for example, v/v, 1:1) to obtain 423.3 mg of an orange pure product by silica gel column chromatography. The rate is 83%.

(Scheme 2) Dissolve 374 mg (1 mmol) of the compound of formula 8 prepared in Example 1 in 10 mL of dichloromethane, add 387 mg (3 mmol) of DIPEA, and then add 519 mg (3 mmol) of benzene thiochloroformate The ester was refluxed at 50° C. for 10 h, and then subjected to suspension evaporation using a rotary evaporator to obtain a solid, which was the crude product of the compound represented by Formula 2. If you want to get a purer product, you can use a mixed system of dichloromethane and petroleum ether (for example, v/v, 1:1) to obtain 464.1 mg of an orange pure product by silica gel column chromatography. The rate is 91%.

(Scheme 3) Dissolve 374 mg (1 mmol) of the compound represented by formula 8 in 10 mL of dichloromethane, add 645 mg (5 mmol) of DIPEA, then add 865 mg (5 mmol) of phenyl thiochloroformate and reflux at 50 °C 10h, and then use a rotary evaporator to carry out suspension evaporation to obtain a solid, which is the crude product of the compound shown in formula 2. If you want to get a purer product, you can use a mixed system of dichloromethane and petroleum ether (for example, v/v, 1:1) to obtain 443.7 mg of an orange pure product by silica gel column chromatography. The rate is 87%.

(Scheme 4) Dissolve 374 mg (1 mmol) of the compound of formula 8 prepared in Example 1 in 15 mL of dichloromethane, add 258 mg (2 mmol) of DIPEA, and then add 346 mg (2 mmol) of benzene thiochloroformate The ester was refluxed at 50° C. for 10 h, and then subjected to suspension evaporation using a rotary evaporator to obtain a solid, which was the crude product of the compound represented by Formula 2. If you want to obtain a purer product, you can use a mixed system of dichloromethane and petroleum ether (for example, v/v, 1:1) to obtain 433.5 mg of an orange pure product by silica gel column chromatography. The rate is 85%.

(Scheme 5) Dissolve 374 mg (1 mmol) of the compound of formula 8 prepared in Example 1 in 10 mL of dichloromethane, add 387 mg (3 mmol) of DIPEA, and then add 519 mg (3 mmol) of phenyl thiochloroformate Reflux at 50° C. for 15 h, and then perform suspension evaporation using a rotary evaporator to obtain a solid, which is the crude product of the compound represented by Formula 2. If you want to get a purer product, you can use a mixed system of dichloromethane and petroleum ether (for example, v/v, 1:1) to obtain 479.4 mg of an orange pure product by silica gel column chromatography. The rate is 94%.

The NMR characterization (i.e. hydrogen spectrum, carbon spectrum) data of the mercury ion fluorescent probe of the present invention is as follows:

1H-NMR (400 MHz, DMSO-d6) δ (*10-6): 2.59(s, 3H), 7.21(d, J = 8.0 Hz, 2H), 7.35(t, J = 8.0 Hz, 1H), 7.49(t, J = 8.0 Hz, 2H), 7.57(t, J = 8.0 Hz, 2H), 7.65(t, J = 8.0 Hz, 2H), 8.19(d, J = 8.0 Hz, 2H), 8.33( d, J = 8.0 Hz,2H), 8.34 (s, 2H). 13C-NMR (100 MHz, DMSO-d6) δ (*10-6): 20.90, 115.69, 121.94, 122.97, 123.63, 126.55, 127.48, 127.73, 129.82, 130.55, 133.46, 135.56, 138.70, 145.61, 152.86, 153.62, 161.72, 193.28. ESI-MS calcd for C28H19N2O2S3 [M + H]+ 511.1, found 51

Example 3

Using the compounds of Scheme 5, probes were prepared. Weigh 5 mg of the compound represented by formula 2 into a colorimetric tube, add 1 mL of dichloromethane, shake well to dissolve the probe, and then dilute to 10 mL with absolute ethanol to prepare a 1 mM probe. Needle stock solution.

The prepared probe was used for detection, and the fluorescence detection method was as follows: add 5 mL of absolute ethanol to the colorimetric tube, pipette 50 µL of the probe stock solution (1 mM) into the colorimetric tube, add 2- Add 3mL of distilled water, then add 0.5mL of 4-hydroxyethylpiperazineethanesulfonic acid (HEPES), then dilute to 10mL with distilled water, pipette 200 µL of mercury ion solution (1 mM) into the colorimetric tube. Shake well, and after 20 min, measure the fluorescence spectrum with a fluorescence spectrophotometer (Horiba FluoroMax-4).

The test results are shown in Figure 1.

Figure 1 is the fluorescence spectrum of the probe (5 µM) before and after adding Hg2+ (20 µM). We can see that the fluorescence changes are very obvious through the illustration.

Example 4

Probes prepared using compounds of Scheme 2. Weigh 5 mg of the compound represented by formula 2 into a colorimetric tube, add 1 mL of dichloromethane, shake well to dissolve the probe, and then dilute to 10 mL with absolute ethanol to prepare a 1 mM probe. Needle stock solution.

To detect the fluorescence spectrum, the specific method is: add 5mL of absolute ethanol to the colorimetric tube, pipette 50 µL of the probe stock solution (1 mM) into the colorimetric tube, add 2-3mL of distilled water, and then add 0.5 mL of 4-hydroxyethylpiperazineethanesulfonic acid (HEPES), then distilled water to 10 mL, pipette 50-400 µL of mercury ion solution (1 mM) into the colorimetric tube. Shake well, and after 20 min, measure the fluorescence spectrum with a fluorescence spectrophotometer (Horiba FluoroMax-4).

The result is shown in Figure 2.

Figure 2 Effects of different concentrations of Hg2+ (0-40 µM) on the fluorescence spectrum of the probe (5 µM). It can be seen from the figure that the response of the probe (5 µM) to different concentrations of Hg2+ (0-16 µM) satisfies a good linear relationship .

It can be seen that with the increase of Hg2+ concentration in the probe solution, the fluorescence intensity gradually increases, and within the range of (0-16 μM) Hg2+ concentration, the response of the probe to the concentration of Hg2+ is linear. Therefore, the probe of the present invention can more accurately determine the content of mercury ions in the sample to be tested.

Example 5

Using the compounds of Scheme 1, probes were prepared. Weigh 5 mg of the compound represented by formula 2 into a colorimetric tube, add 1 mL of dichloromethane, shake well to dissolve the probe, and then dilute to 10 mL with absolute ethanol to prepare a 1 mM probe. Needle stock solution.

The prepared probe was used for detection, and the fluorescence intensity detection method was as follows: add 5 mL of absolute ethanol to the colorimetric tube, pipette 50 μL of the probe stock solution (1 mM) into the colorimetric tube, add 2 -3mL of distilled water, then add 0.5 mL of 4-hydroxyethylpiperazineethanesulfonic acid (HEPES), then dilute to 10mL with distilled water, pipette 200 µL of different ion analytes (1 mM) into the colorimetric tube. Shake well, and after 20 min, measure the fluorescence spectrum with a fluorescence spectrophotometer (Horiba FluoroMax-4).

The result is shown in Figure 3.

Figure 3 Fluorescence intensity of different ion analytes (20 µM) versus probe (5 µM). All measurements are made after 20 minutes, and the response time is relatively fast.

Analytes include: cadmium ions (Cd2+), lead ions (Pb2+), magnesium ions (Mg2+), aluminum ions (Al3+), ferric ions (Fe2+), ferric ions (Fe3+), chromium ions (Cr3+), Calcium ion (Ca2+), zinc ion (Zn2+), sodium ion (Na+), potassium ion (K+), cobalt ion (Co2+) and nickel ion (Ni2+). Their concentrations were all 20 µM. All test conditions were done in distilled water, absolute ethanol (v/v, 5:5) and 4-hydroxyethylpiperazineethanesulfonic acid (HEPES), and the probes used were those prepared in Scheme 1, And all spectra were measured at 25°C 20 min after the addition of the analytes. Specifically, add 5 mL of absolute ethanol to the colorimetric tube, pipette 50 µL of the probe stock solution (1 mM) into the colorimetric tube, add 2-3 mL of distilled water, and then add 0.5 mL of 4-hydroxyethyl piperazineethanesulfonic acid (HEPES), then distilled water to 10mL, pipette 200µL of different ion analytes (1 mM) into the colorimetric tube. Shake well and measure after 20 min. The result is shown in Figure 3.

It can be seen from Figure 3 that the probe only responds to mercury ions, and the common ions in the environment will not significantly interfere with the fluorescence intensity of the probe to mercury ions, so the probe has good selectivity and anti-interference.

As shown in Figure 3, the left side is the fluorescence intensity of the analytical stock solution containing only the following ions, and the right side is the fluorescence intensity after adding Hg2+. (1) Hg2+; (2) Cd2+; (3) Pb2+; (4) Mg2+; (5) Al3+; (11) Na+; (12) K+; (13) Co2+; (14) Ni2+

Although the present invention has been described with the above embodiments, it should be understood that, without departing from the spirit of the present invention, the present invention can be further modified and changed, and these modifications and changes are within the protection scope of the present invention .

Claims (5)

1. a kind of fluorescence probe for analyzing mercury ion, it is thiocarbonic acid esters compound, and its structure is as shown in Equation 1：

Formula 1

Wherein：R1, R2, R3, R4, R5, R6, R7, R8, R9, R10, R11, R12, R13, R14, R15 are hydrogen atom, and carbon atom is 1-3 straight or branched alkyl, carbon atom be 1-3 straight or branched alkoxyl, sulfonic group, ester group, carboxyl；R1, R2, R3, R4, R5, R6, R7, R8, R9, R10, R11, R12, R13, R14 and R15 are identical or different.

2. fluorescence probe according to claim 1, it is characterized in that, structural formula is as shown in Equation 2：

Formula 2.

3. the preparation method of the fluorescence probe described in a kind of claim 2, it is characterised in that using following steps:

Hexamethylenetetramine shown in formula 4 is dissolved in 20 mL trifluoroacetic acid, adds the p-methyl phenol shown in formula 3, two Person's mol ratio is 4:1,100 DEG C is heated to reflux 6 hours, and shading treatment, adds product in frozen water after having reacted, and separates out solid Filtered after body, obtain sterling formula 5

The formula 5 of 3 formula of formula 4

Compound shown in formula 5 is dissolved in absolute ethyl alcohol, adds the amino benzenethiol shown in formula 6, the two mol ratio is 1: 1, stirring at normal temperature, 37% hydrochloric acid and 30% hydrogen peroxide are added dropwise in whipping process, react 30 min, then filtered Obtain sterling formula 8

The formula 8 of 6 formula of formula 5

Compound shown in formula 8 is dissolved in dichloromethane, DIPEA (DIPEA) catalysis is added, adds formula 9 Shown thio phenyl chloroformate, the mol ratio of the two are 1:3,50 DEG C are heated to reflux 10h, are then filtered, obtain sterling Formula 2

The formula 2 of 8 formula of formula 9.

4. a kind of fluorescence probe of claim 1 or 2 is being prepared for detecting the use in sample in the preparation of ion concentration of mercury On the way.

5. application according to claim 4, it is characterised in that：

Match somebody with somebody manufacturing probe storing solution first：Weigh the compound shown in 5 mg formula 2 to be added in colorimetric cylinder, add 1mL dichloro Methane, shake up, dissolve probe, then with absolute ethyl alcohol constant volume to 10 mL, be configured to 1 mM probe storing solution；

Then, 5mL absolute ethyl alcohol is added in colorimetric cylinder, the probe storing solution (1 mM) for pipetting 50 μ L is put into colorimetric cylinder, 2-3mL distilled water is added, then adds 0.5 mL 4- hydroxyethyl piperazineethanesulfonic acids（HEPES）, then arrived with distilled water constant volume 10mL；Pipette 200 μ L mercury ions solution (1 mM) to add in colorimetric cylinder, shake up, after 20min, use sepectrophotofluorometer （Horiba FluoroMax-4）Determine fluorescence spectrum.