CN108997249B

CN108997249B - Application of probe

Info

Publication number: CN108997249B
Application number: CN201810948737.4A
Authority: CN
Inventors: 王小锋
Original assignee: Nanjing Xiaozhuang University
Current assignee: Nanjing Xiaozhuang University
Priority date: 2018-01-31
Filing date: 2018-01-31
Publication date: 2020-07-17
Anticipated expiration: 2038-01-31
Also published as: CN108863974B; CN108863975B; CN108863974A; CN108148013A; CN108997249A; CN108863975A; CN108148013B

Abstract

The invention discloses an application of a probe, and belongs to the field of analysis and detection. The method takes a compound I as a raw material, and reacts with chloroacetyl chloride under the action of a weakly alkaline catalyst to obtain a compound II; and reacting the compound II with salicylaldehyde hydrazone under the condition of weak alkaline salt to obtain a compound III. The synthetic method is simple, is easy for industrial production, and has low detection limit and high selectivity when being used as a probe molecule for detecting zinc ions.

Description

Application of probe

The application is as follows: 2018-01-31, with the application number: 2018100937972, the name is: a zinc ion probe, a preparation method and application thereof.

Technical Field

The invention belongs to the field of analysis and detection, and particularly relates to application of a probe.

Background

Zinc ions are an essential trace element in organisms, widely exist in cells and body fluid, and play an important role in physiological processes such as growth and development, reproduction, nerve signal transmission, gene entry, nucleic acid identification, cell growth regulation, apoptosis and the like. Excessive or insufficient intake of zinc ions can cause growth and development disorder of organisms, and various diseases such as appetite reduction, low immunity, diabetes, prostate cancer and the like are related to zinc ion imbalance. Therefore, the development and research of novel zinc ion detection methods are of great significance to both biological science and environmental science.

Disclosure of Invention

The invention provides a zinc ion probe, a synthetic method and application aiming at the defects in the prior art.

The purpose of the invention can be realized by the following technical scheme:

a zinc ion probe, the structural formula of the probe is as follows:

the preparation method of the zinc ion probe comprises the following reaction route:

the method specifically comprises the following steps:

the first step is as follows: the method takes a compound I as a raw material, and reacts with chloroacetyl chloride under the action of a weakly alkaline catalyst to obtain a compound II;

in some embodiment aspects: the solvent used in the first step is dichloromethane, tetrahydrofuran and toluene. The alkalescent catalyst used in the first step of reaction is any one of dimethylaminopyridine, pyridine and triethylamine. The reaction temperature in the first step is-10 to 0 ℃.

In some more specific embodiments: the molar ratio of the compound I to the weakly basic catalyst is 1:1-1: 5.

The second step is that: and reacting the compound II with salicylaldehyde hydrazone in the presence of an acid-binding agent in the presence of potassium iodide serving as a catalyst to obtain a compound III.

In some embodiment aspects: the solvent used in the second step is acetonitrile, ethanol and dichloromethane. The acid-binding agent used in the second step is at least one of potassium carbonate, sodium carbonate and triethylamine. The second reaction temperature is the heating reflux temperature.

In some more specific embodiments: the molar ratio of the compound II to the acid-binding agent is 1:1-1: 10.

The technical scheme of the invention is as follows: the probe is used for detecting zinc ions.

The invention has the beneficial effects that:

the synthetic method is simple, is easy for industrial production, and has low detection limit and high selectivity when being used as a probe molecule for detecting zinc ions.

Drawings

FIG. 1 shows probe molecules hhpa vs. Zn²⁺Selective absorption spectrum identification.

FIG. 2 shows Zn²⁺Absorbance spectrum titration plot for probe molecule hhpa.

FIG. 3 shows probe molecules hhpa vs. Zn²⁺Selective fluorescence spectrum identification.

FIG. 4 is Zn²⁺A fluorescence spectrum titration graph of the probe molecule hhpa.

FIG. 5 shows the selective recognition of Zn for probe hhpa when other coexisting metal ions are present in the solution²⁺Influence graph of (c).

FIG. 6 shows Zn²⁺Graph of the effect of the reaction time with the probe molecule hhpa on the fluorescence intensity of the solution;

FIG. 7 shows probe molecules hhpa in different Zn²⁺Fluorescence intensity in concentration solution.

Detailed Description

The invention is further illustrated by the following examples, without limiting the scope of the invention:

example 1

Adding phenoxazin-3-amine (1.98g, 10mmol), 100m L dichloromethane and 4-dimethylamino pyridine (1.22g, 10mmol) into a 250m L three-neck flask in sequence, fully cooling in an ice salt water bath, slowly dripping a dichloromethane solution of 20m L dissolved with chloroacetyl chloride (1.13g, 10mmol) into the fully stirred three-neck flask by using a constant pressure funnel, controlling the temperature of the reaction solution in the three-neck flask not to exceed 0 ℃, continuing to react in an ice salt water bath for 2h after finishing the dripping, adjusting the pH value of the reaction solution to about 9 by using a 0.1 mol/L NaOH solution after the reaction is finished, extracting the reaction solution by using dichloromethane (3 × 25m L), combining the organic phases, washing by using water (3 × 25m L), and then using anhydrous Na (L)₂SO₄Dry overnight. After filtration, the filtrate was rotary evaporated to remove the organic solvent to give 2.54g of a blue-green solid product ii, yield: 92.7%, purity: 99.36 percent. Elemental analysis: (%) forC14H11N2O2Cl calculated: c61.21; h4.04; n10.20, found: c61.43; h4.18; and (3) N10.07.

IR(KBr),ν,cm^-1:3462,3397,1638,1329,1287,1209,1117,1083,921,848,741,718。

1H NMR(500MHz,CDCl₃,TMS):＝7.73(s,1H),7.45(s,1H),7.24(d,J＝7.2,1H),6.83(t,J＝7.2,1H),6.72-6.68(m,2H),6.61-6.57(m,2H),6.42(s,1H),4.31(s,2H)ppm.

Dissolving a compound II (1.37g, 5mmol), potassium iodide (0.83mg, 0.005mmol) and anhydrous potassium carbonate (1.38g, 5mmol) in acetonitrile 40m L in a flask of 100m L, slowly dropwise adding a solution of salicylaldozone (0.68g, 5mmol) in acetonitrile 15m L by using a constant-pressure funnel under the conditions of reflux and stirring, controlling the dropwise adding to be finished within 1h, continuing refluxing for 20h after the dropwise adding is finished, cooling the reaction liquid to room temperature after the reaction is finished, pouring the reaction liquid into water, extracting by using dichloromethane (3 × 25m L), combining organic phases, washing by using a saturated NaCl solution (3 × 25m L), and using anhydrous Na for an organic phase₂SO₄Dry overnight. After filtration, the filtrate was rotary evaporated to remove the organic solvent to obtain a crude product. The crude product was purified by silica gel column (ethyl acetate: petroleum ether 1:5), and the solvent was evaporated to dryness to give 1.72g of blue-green solid product iii (hhpa), yield: 92.0%, purity: 99.43 percent.

Elemental analysis: (%) for C21H18N4O3 calculated: c67.37; h4.85; n14.96, found: c67.51; h4.83; n14.78.

IR(KBr),ν,cm^-1:3413,1624,1406,1338,1276,1251,1210,1127,1064,961,917,854,731,662,618。1H NMR(500MHz,CDCl₃,TMS):10.23(s,1H),8.61(s,1H),8.41(t,J＝7.2,1H),7.71(s,1H),7.43(s,1H),7.41-7.35(m,2H),7.23-7.19(m,3H),6.85(t,J＝7.2,1H),6.72-6.67(m,2H),6.62-6.57(m,2H),6.42(s,1H),4.25(d,J＝7.2,2H)ppm.

Example 2

Adding phenoxazin-3-amine (1.98g, 10mmol), 100m L tetrahydrofuran and 3m L pyridine into a 250m L three-neck flask in sequence, fully cooling in an ice salt water bath, slowly dripping a dichloromethane solution of 20m L and chloroacetyl chloride (1.13g, 10mmol) into the fully stirred three-neck flask by using a constant-pressure funnel, controlling the temperature of a reaction solution in the three-neck flask to be not more than 0 ℃, continuing to react in the ice salt water bath for 2h after finishing the dripping, adjusting the pH value of the reaction solution to about 9 by using a 0.1 mol/L NaOH solution after finishing the reaction, extracting the reaction solution by using dichloromethane (3 × 25m L), combining organic phases, washing by using water (3 × 25m L), and then using anhydrous Na₂SO₄Dry overnight. Filtering, rotary evaporating the filtrate, and removing organic solvent to obtainBlue-green solid product II 2.50g, yield: 91.2%, purity: 99.27 percent.

Dissolving a compound II (1.37g, 5mmol), potassium iodide (0.83mg, 0.005mmol) and anhydrous sodium carbonate (2.12g, 10mmol) in 40m L ethanol in a 100m L flask, slowly dropwise adding a 15m L acetonitrile solution dissolved with salicylaldozone (0.68g, 5mmol) by using a constant pressure funnel under the conditions of reflux and stirring, controlling the dropwise adding to be finished within 1h, continuing refluxing reaction for 20h after the dropwise adding is finished, cooling the reaction liquid to room temperature after the reaction is finished, pouring the reaction liquid into water, extracting by using dichloromethane (3 × 25m L), combining organic phases, washing by using a saturated NaCl solution (3 × 25m L), and using anhydrous Na for an organic phase₂SO₄Dry overnight. After filtration, the filtrate was rotary evaporated to remove the organic solvent to obtain a crude product. The crude product was purified by silica gel column (ethyl acetate: petroleum ether 1:5), and the solvent was evaporated to dryness to give 1.69g of blue-green solid product iii (hhpa), yield: 90.4%, purity: 99.31 percent.

Example 3

Adding phenoxazin-3-amine (1.98g, 10mmol), 100m L toluene and 7m L triethylamine into a 250m L three-neck flask in sequence, fully cooling in an ice salt water bath, slowly dripping a dichloromethane solution of 20m L and chloroacetyl chloride (1.13g, 10mmol) into the fully stirred three-neck flask by using a constant-pressure funnel, controlling the temperature of a reaction solution in the three-neck flask to be not more than 0 ℃, continuing to react in the ice salt water bath for 2h after finishing the dripping, adjusting the pH value of the reaction solution to about 9 by using a 0.1 mol/L NaOH solution after finishing the reaction, extracting the reaction solution by using dichloromethane (3 × 25m L), combining organic phases, washing by using water (3 × 25m L), and then using anhydrous Na₂SO₄Dry overnight. After filtration, the filtrate was rotary evaporated to remove the organic solvent to give 2.46g of a blue-green solid product ii, yield: 89.8%, purity: 98.77 percent.

Dissolving compound II (1.37g, 5mmol), potassium iodide (0.83mg, 0.005mmol) and 7m L triethylamine in 40m L dichloromethane in a 100m L flask, slowly dropwise adding a 15m L acetonitrile solution dissolved with salicylaldozone (0.68g, 5mmol) by using a constant pressure funnel under the conditions of reflux and stirring, controlling the dropwise adding to be finished within 1h, continuing the reflux reaction for 20h after the dropwise adding is finished, and after the reaction is finished, continuing the reflux reaction for 20hThe reaction was cooled to room temperature and poured into water, then extracted with dichloromethane (3 × 25m L), the organic phases combined and washed with saturated NaCl solution (3 × 25m L), the organic phase washed with anhydrous Na₂SO₄Dry overnight. After filtration, the filtrate was rotary evaporated to remove the organic solvent to obtain a crude product. The crude product was purified by silica gel column (ethyl acetate: petroleum ether 1:5), and the solvent was evaporated to dryness to give 1.63g of blue-green solid product iii (hhpa), yield: 87.2%, purity: 99.27 percent.

Property part

1. Absorption spectrum experiment

Probe molecules hhpa vs Zn²⁺Identification of absorption spectra

As shown in FIG. 1, 10. mu. L of a metal ion solution (Ag, 5 times the molar weight) having a concentration of 0.5 mol/L was added to a solution of 10m L of 0.1 mmol/L of the probe molecule hhpa⁺、Na⁺、K⁺、Ca²⁺、Cd²⁺、Mn²⁺、Mg²⁺、Co²⁺、Cu²⁺、Ni²⁺、Pb² ⁺、Hg²⁺、Al³⁺、Zn²⁺). The solution system used in the experiment was a mixed solution of acetonitrile/water (v/v-1/2), and the absorption spectrum was measured by shimadzu UV-2450 ultraviolet spectrophotometer. As can be seen from FIG. 1, the self-absorption of probe molecules in the mixed solution of acetonitrile/water (v/v-1/2) is around 580nm, and when we add excessive metal ions into the probe molecule solution, we find that only after adding Zn²⁺Then, the original absorption peak at 580nm disappears, a new absorption peak appears at 460nm, the blue color is shifted by 120nm, the color of the solution is changed from blue to yellow, and when other metal ions are added into the probe molecule solution, the phenomenon does not occur, which shows that the absorption spectrum of the probe molecule to Zn²⁺Has unique response.

As shown in FIG. 2, 0.05, 0.1, 0.15, 0.2, 0.25, 0.3, 0.35, 0.4, 0.45, 0.5, 0.55, 0.6, 0.65, 0.7, 0.75, 0.8, 0.85, 0.9, 0.95, 1.0, 1.2, 1.5 times the molar amount of Zn was added to a solution of 10m L, 0.1 mmol/L probe hhpa in that order²⁺. Experiment ofThe solution system used in (1) was a mixed solution of acetonitrile/water (v/v ═ 1/2), and the absorption spectrum was measured by shimadzu UV-2450 ultraviolet spectrophotometer. As can be seen from FIG. 2, with Zn²⁺The absorption wavelength of the solution is gradually blue-shifted from 580nm to 460nm when Zn is added²⁺After the addition amount reaches 1 time of the molar amount of the probe molecules, the absorption wavelength of the solution does not move any more, and the intensity of the absorption peak is basically kept unchanged. This indicates that the probe molecules hhpa and Zn²⁺Is 1:1 coordinated.

2. Fluorescence spectrum experiment

Probe molecules hhpa vs Zn²⁺Fluorescent identification of

As shown in FIG. 3, the probe molecules ddpb were dissolved in a mixed solution of acetonitrile/water (v/v. 1/2) to prepare a solution having a concentration of 10. mu. mol/L, and metal ions (Ag) were added to the solution in an amount of 5 times the molar amount of each of the solutions⁺、Na⁺、K⁺、Ca²⁺、Cd²⁺、Mn² ⁺、Mg²⁺、Co²⁺、Cu²⁺、Ni²⁺、Pb²⁺、Hg²⁺、Al³⁺、Zn²⁺). The excitation wavelength was 460nm, and the fluorescence spectrum of the solution was measured. As can be seen from FIG. 3, the probe molecule solution has a weak fluorescence emission peak at 665nm when excess Zn is added²⁺Then, the weak fluorescence emission peak of the solution at 665nm disappears, a strong fluorescence emission peak appears at 585nm, and the phenomenon is not caused by adding other metal ions, which indicates that the probe molecule pair Zn²⁺Exhibits very strong fluorescent selective recognition. The solution system used in the experiment was a mixed solution of acetonitrile/water (v/v-1/2), and the fluorescence spectrum was measured on an AMINCO Bowman Series 2 fluorescence spectrometer.

As shown in fig. 4, 0.05, 0.1, 0.15, 0.2, 0.25, 0.3, 0.35, 0.4, 0.45, 0.5, 0.55, 0.6, 0.65, 0.7, 0.75, 0.8, 0.85, 0.9, 0.95, 1.0, 1.2, 1.5 times the molar amount of Zn was added to a mixed solution of 10 μmol/L of probe molecule hhpa in acetonitrile/water (v/v: 1/2)²⁺. Excitation at 460nm, the emission spectrum of the solution was measured, as shown with Zn²⁺Increased concentration of (2), weak fluorescence at 665nmThe light emission peak gradually decreases and finally disappears, and a new fluorescence emission peak appears at 585nm, and Zn is formed²⁺The intensity of the emission peak at 585nm did not substantially increase after the addition amount reached 1-fold molar amount.

As shown in FIG. 5, 10 mol/L of metal ions (Ag) dissolved in 10 times the molar amount of each of the probe molecules hhpa in a mixed solution of acetonitrile/water (v/v. 1/2) was added⁺、Na⁺、K⁺、Ca²⁺、Cd²⁺、Mn²⁺、Mg²⁺、Co²⁺、Cu²⁺、Ni²⁺、Pb²⁺、Hg²⁺、Al³⁺) Measuring the fluorescence intensity of the solution at an excitation wavelength of 460nm and an emission wavelength of 585nm, and adding Zn in an amount of 10 times the molar amount of the solution²⁺The fluorescence intensity of the solution was measured at an excitation wavelength of 460nm and an emission wavelength of 585nm, and it can be seen from FIG. 5 that when other metal ions are present in the solution in large amounts, the probe molecules hhpa are coupled to Zn²⁺Is not affected.

As shown in FIG. 6, 2 times the molar amount of Zn was added to a 10. mu. mol/L mixed solution of probe molecule hhpa in acetonitrile/water (v/v. 1/2)²⁺. The fluorescence intensity of the solution was recorded at 0, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10 minutes at an excitation wavelength of 460nm and an emission wavelength of 585nm, respectively. As shown in FIG. 6, Zn was added to the probe molecule hhpa solution²⁺After 2 minutes, the fluorescence emission intensity reaches a maximum and remains substantially constant over time.

As shown in FIG. 7, Zn concentrations of 0.2 to 2. mu. mol/L (in 0.05. mu. mol/L) at 10m L, respectively²⁺A mixed solution of acetonitrile/water (v/v-1/2) was added with a 0.1m solution of L concentration of 1 mmol/L of hhpa, and the fluorescence intensity of the solution was measured at an excitation wavelength of 460nm and an emission wavelength of 585nm²⁺The concentration showed a good linear relationship (R2 ═ 0.9988) in the range of 0.2-1.1. mu. mol/L, with a detection limit of 1.26 × 10 calculated using the 3. sigma. IUPAC standard^-7mol/L。

Claims

1. A preparation method of a probe for detecting zinc ions is characterized by comprising the following steps: the method comprises the following specific steps:

adding 1.98g, 10mmol,100m L dichloromethane and 1.22g, 10mmol of 4-dimethylamino pyridine into a 250m L three-neck flask in sequence, fully cooling in an ice salt water bath, slowly dripping a dichloromethane solution of 20m L dissolved with chloroacetyl chloride of 1.13g and 10mmol into the fully stirred three-neck flask by using a constant pressure funnel, controlling the temperature of the reaction solution in the three-neck flask not to exceed 0 ℃, continuing to react in the ice salt water bath for 2 hours after finishing the reaction, adjusting the pH value of the reaction solution to 9 by using a 0.1 mol/L NaOH solution, extracting the reaction solution by using dichloromethane 3 × 25m L, combining organic phases, washing by using water of 3 × 25m L, and then washing by using anhydrous Na₂SO₄Drying overnight; after filtration, the filtrate was rotary evaporated to remove the organic solvent to give 2.54g of a blue-green solid product ii, yield: 92.7%, purity: 99.36 percent;

dissolving 1.37g and 5mmol of compound II, 0.83mg and 0.005mmol of potassium iodide and 1.38g and 5mmol of anhydrous potassium carbonate in 40m L acetonitrile in a 100m L flask, slowly dropwise adding a 15m L acetonitrile solution containing 0.68g and 5mmol of salicylaldehyde hydrazone under the conditions of reflux and stirring, controlling the dropwise adding within 1h, continuously carrying out reflux reaction for 20h after the dropwise adding is finished, cooling the reaction liquid to room temperature after the reaction is finished, pouring the reaction liquid into water, extracting by using dichloromethane 3 × 25m L, combining organic phases, washing by using a saturated NaCl solution 3 × 25m L, and washing an organic phase by using anhydrous Na₂SO₄Drying overnight, filtering, and then carrying out rotary evaporation on the filtrate to remove the organic solvent to obtain a crude product; the crude product was purified through a silica gel column, ethyl acetate: and (3) evaporating the solvent to dryness to obtain 1.72g of a blue-green solid product III, wherein the yield is as follows: 92.0%, purity: 99.43 percent.