CN107290323B - Near-infrared fluorescent probe and preparation method and application technology thereof - Google Patents

Abstract

The technical field of the patent of the present invention belongs to the technical field of fluorescent probe sensing in analytical chemistry, and relates to a novel near-infrared fluorescent probe and its preparation method and application in the detection of polysulfides. The method includes obtaining the probe molecule by reacting a Nile red derivative with 2-fluoro-4-nitrobenzoic acid. The preparation method of the probe is simple, the structure is stable, and the fluorescence spectrophotometer can be used to detect sodium disulfide with high selectivity, high sensitivity and rapidity. The probe provides a novel method for detecting polysulfides, and has good application prospects.

Description

A near-infrared fluorescent probe and its preparation method and application technology

technical field

The invention relates to the technical field of fluorescent probe sensing in analytical chemistry, and relates to a near-infrared fluorescent probe for detecting polysulfides and a preparation method and application technology thereof.

Background technique

With the development of industry and agriculture, polysulfides are widely used as polymerization terminators, depilatory agents in the tanning industry, pesticides in agriculture, and petroleum refining auxiliaries. Its impact on the environment cannot be underestimated. Therefore, the qualitative and quantitative detection of polysulfides is very important.

In addition, active sulfur-containing compounds, including hydrogen sulfide, hydrogen persulfide, and thiol amino acids, play a very important role in cellular metabolism (protein function, redox signaling, etc.). As one of the important signaling molecules, hydrogen sulfide is considered to be the third type of gas transmitter in biological systems. Too high or too low hydrogen sulfide content will affect the central nervous system, and it is associated with many major diseases, such as: Down's syndrome syndrome, Alzheimer's disease, etc. However, recent studies have shown that intracellular signaling may be achieved through the co-action of hydrogen sulfide (produced by the oxidation of hydrogen sulfide with biologically reactive oxygen species) and hydrogen sulfide, and that hydrogen sulfide can activate tumor suppressor more efficiently than hydrogen sulfide. , ion channels, transcription factors and more efficient sulfation. However, the research on the biological function of hydrogen polysulfide is still in its infancy, and there are still many problems that need to be solved urgently. The traditional method for the determination of polysulfide mainly relies on GC-MS. Although this method can determine hydrogen polysulfide, it also has certain defects, such as poor selectivity, complicated sample preparation, complicated operation, expensive instrument, time-consuming and difficult. Universal application. Therefore, it is particularly important to develop an effective, accurate and convenient method for detecting hydrogen polysulfide, which is not only of great significance for environmental monitoring, but also of great significance for the initial clinical diagnosis of some major diseases.

The invention relates to a near-infrared fluorescent probe for detecting polysulfides and a preparation method and application technology thereof. We reacted Nile red derivatives with 2-fluoro-4-nitrobenzoic acid and 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride and 4-dimethylaminopyridine , designed this novel near-infrared fluorescent probe. The probe has the advantages of high selectivity, high sensitivity and rapid detection in the detection of hydrogen polysulfide. In addition, near-infrared fluorescence does little damage to biological organisms when performing biological tissue or cell detection. At the same time, biological tissue has the least absorption of fluorescence in the near-infrared region, and the near-infrared light can penetrate biological tissue in a large distance, and can detect and image deep tissues and organs. Therefore, this probe has potential application prospects.

SUMMARY OF THE INVENTION

The purpose of the present invention is to provide a near-infrared fluorescent probe and a preparation method of the probe and its application in detecting polysulfides.

According to the present invention, to obtain a probe molecule by reacting a Nile red derivative with a hydroxyl group with 2-fluoro-4-nitrobenzoic acid, the synthesis steps specifically include: (1) under an ice bath, 5-diethyl Amino-2-nitrosophenol hydrochloride and 1,6-dihydroxynaphthalene are dissolved in N,N-dimethylformamide solvent, and the reaction is stirred under argon protection, and finally a Nile red with hydroxyl group is obtained. Derivative; (2) combining the Nile Red derivative obtained in step (1) with 2-fluoro-4-nitrobenzoic acid and 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide The amine hydrochloride is dissolved in dichloromethane, an appropriate amount of catalyst is added, the reaction is stirred at room temperature, and the near-infrared fluorescent probe is finally obtained through separation and purification.

Compared with the prior art, the patented technology has the following advantages:

1. Good selectivity and strong anti-interference ability;

2. High sensitivity, capable of detecting trace targets;

3. Quick response, which can realize rapid detection;

3. Simple synthesis and convenient detection;

4. Near-infrared light detection has little damage to organisms and has potential application prospects.

Description of drawings

Figure 1 shows the fluorescence titration diagram of probe detection of sodium disulfide

Figure 2 shows the response time fluorescence spectrum of probe detection of polysulfides

Figure 3 shows the pH-optimized fluorescence spectrum of the probe.

Figure 4 shows the selective experimental fluorescence spectrum of the probe

Detailed ways

The present invention will be further described below with reference to the embodiments. The embodiments of the present invention are only used to illustrate the technical solutions of the present invention and do not limit the present invention.

Example 1

Under an ice bath, 5-diethylamino-2-nitrosophenol hydrochloride (0.5 g, 2.6 mmol) and 1,6-dihydroxynaphthalene (0.41 g, 2.6 mmol) were dissolved in 8 mL of N, In N-dimethylformamide, the reaction was refluxed at 140°C for 5 hours under the protection of argon. The reaction was completed. After the reaction solution was cooled to room temperature, 100 mL of H ₂ O was added, extracted with dichloromethane, dried over anhydrous sodium sulfate, and spin-dried. , separated and purified by chromatographic column, and finally Nile red derivatives with hydroxyl groups were obtained. The Nile Red derivative (110 mg, 0.33 mmol) was combined with 2-fluoro-4-nitrobenzoic acid (74 mg, 0.4 mmol) and 1-(3-dimethylaminopropyl)-3-ethylcarbodiidene Amine hydrochloride (126 mg, 0.66 mmol) was dissolved in 10 mL of dichloromethane, and the catalyst 4-dimethylaminopyridine (40 mg, 0.33 mmol) was added, and the reaction was stirred at room temperature. After the reaction was completed, the reaction solution was poured into 40 mL. In the water, extract with dichloromethane, dry with anhydrous sodium sulfate, spin dry, and separate and purify through a chromatographic column to finally obtain the final product, that is, the near-infrared fluorescent probe of the present invention.

Example 2

Under an ice bath, 5-diethylamino-2-nitrosophenol hydrochloride (1.15 g, 6.0 mmol) and 1,6-dihydroxynaphthalene (0.82 g, 5.2 mmol) were dissolved in 12 mL of N, N-dimethylformamide, refluxed at 140°C for 8 hours under argon protection, the reaction was completed, after the reaction solution was cooled to room temperature, 100 mL of H ₂ O was added, extracted with dichloromethane, dried over anhydrous sodium sulfate, and spin-dried , separated and purified by chromatographic column, and finally Nile red derivatives with hydroxyl groups were obtained. The Nile red derivative (330 mg, 0.99 mmol) was combined with 2-fluoro-4-nitrobenzoic acid (370 mg, 2.0 mmol) and 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide Amine hydrochloride (530mg, 3.3mmol) was dissolved in 20mL of dichloromethane, then the catalyst 4-dimethylaminopyridine (50mg, 0.39mmol) was added, and the reaction was stirred at room temperature. After the reaction was completed, the reaction solution was poured into 50mL In the water, extract with dichloromethane, dry with anhydrous sodium sulfate, spin dry, and separate and purify through a chromatographic column to finally obtain the final product, that is, the near-infrared fluorescent probe of the present invention.

Example 3

Under an ice bath, 5-diethylamino-2-nitrosophenol hydrochloride (0.5 g, 2.6 mmol) and 1,6-dihydroxynaphthalene (0.41 g, 2.6 mmol) were dissolved in 8 mL of N, In N-dimethylformamide, refluxed at 140 °C for 3 hours under argon protection, the reaction was completed, after the reaction solution was cooled to room temperature, 100 mL of H ₂ O was added, extracted with dichloromethane, dried over anhydrous sodium sulfate, and spin-dried , separated and purified by chromatographic column, and finally Nile red derivatives with hydroxyl groups were obtained. The Nile Red derivative (110 mg, 0.33 mmol) was combined with 2-fluoro-4-nitrobenzoic acid (74 mg, 0.4 mmol) and 1-(3-dimethylaminopropyl)-3-ethylcarbodiidene Amine hydrochloride (126 mg, 0.66 mmol) was dissolved in 10 mL of dichloromethane, and the catalyst 4-dimethylaminopyridine (20 mg, 0.17 mmol) was added, and the reaction was stirred at room temperature. After the reaction was completed, the reaction solution was poured into 40 mL. In the water, extract with dichloromethane, dry with anhydrous sodium sulfate, spin dry, and separate and purify through a chromatographic column to finally obtain the final product, that is, the near-infrared fluorescent probe of the present invention.

The probes obtained in Examples 1-3 are the same substance through TLC monitoring, and the successful preparation of the probes is proved by NMR characterization. The characterization data are as follows: ¹ H NMR (400MHz, CDCl ₃ )δ9.10-9.07 (q,1H),8.56-8.52(m,2H),8.42(d,J=8.0Hz,1H),7.60(d,J=8.0Hz,1H),7.54-7.51(q,1H),7.45( t, J=9.2Hz, 1H), 6.70-6.67(q, 1H), 6.49(d, J=2.8Hz, 1H), 6.41(s, 1H), 3.51-3.46(q, 4H), 1.27(t , J=7.2Hz, 6H). ¹³ C NMR (100MHz, CDCl ₃ )δ182.6, 166.9, 164.1, 160.3, 152.5, 151.1, 147.0, 138.8, 133.9, 131.4, 130.5, 130.3, 130.0, 128.7, 127.9, 125.0 123.1, 118.9, 118.7, 116.3, 110.0, 105.6, 96.3, 45.2, 12.6.

Application Example 1: As shown in Figure 1, the fluorescence titration diagram of probe detection of sodium disulfide. The probe (5 μM) was added to the mixed solution of dimethyl sulfoxide and water, and the fluorescence of the probe itself was very weak under the excitation of light with a wavelength of 550 nm. When the probe responds to sodium disulfide, the fluorescence intensity at 655 nm in the near-infrared region increases, and the fluorescence intensity increases with the increase of sodium disulfide concentration, showing a good linear relationship. It is proved that the probe can be well applied to the detection of sodium disulfide.

Application Example 2: As shown in Figure 2, the response time fluorescence spectrum of probe detection of sodium disulfide. The probe (5μM) was added to the mixed solution of dimethyl sulfoxide and water. The fluorescence of the probe itself was very weak under the excitation of light with a wavelength of 550nm, but after the addition of sodium disulfide, the fluorescence intensity at 655nm in the near-infrared region It started to increase gradually and completely responded within 6 minutes with no change in fluorescence intensity. This proves that the probe has the ability to rapidly detect sodium disulfide.

Application Example 3: As shown in Figure 3, the pH-optimized fluorescence spectrum of the probe. Add the probe (5 μM) and the target sodium disulfide to the mixed solution of dimethyl sulfoxide and water (adjust the pH so that the pH value of the solution is 3-10 respectively), and keep other test conditions unchanged. The response of the probe to sodium disulfide at different pH was obtained. It can be seen from the figure that the probe can maintain good stability between pH=5 and 10, and can respond well to sodium disulfide.

Application Example 4: As shown in Figure 4, the selective experimental fluorescence spectrum of the probe. Under the same test system and test conditions, different interfering substances (1-29 are respectively: probe itself, HCO ₃ ^- , Br ^- , HPO ₄ ^2- , I ^- , NO ₃ ^- , NO ₂ ^- , H ₂ PO ₄ ^- , Ac ^- , K ⁺ , Thr, Mg ²⁺ , Zn ²⁺ , Pro, SO ₄ ^2- , Gly, Ca ²⁺ , Na ⁺ , Trp, Arg, Ala, Na ₂ S ₂ O ₃ , Hcy, SO ₃ ²⁻ , GSH, Cys, S ²⁻ , HS ⁻ , Na ₂ S ₂ ). The experimental results show that the selectivity of the probe is very good, the probe does not respond to these interfering substances, and the fluorescence intensity is almost unchanged. Only in the presence of sodium disulfide in the solution, the probe responds rapidly to it, and the fluorescence intensity at 655 nm in the near-infrared region is greatly enhanced. It reflects the high selectivity of the probe for the detection of sodium disulfide.

Claims (6)

1. A near-infrared fluorescent probe is characterized in that the molecular structural formula is as follows:

2. the method for preparing the near-infrared fluorescent probe according to claim 1, characterized in that the preparation process comprises the following steps:

a: dissolving 5-diethylamino-2-nitrosophenol hydrochloride and 1, 6-dihydroxynaphthalene in an N, N-dimethylformamide solvent in an ice bath, carrying out reflux reaction for a certain time under the protection of argon, and separating and purifying to finally obtain a hydroxyl nile red derivative;

b: and (b) dissolving the nile red derivative obtained in the step (a), 2-fluoro-4-nitrobenzoic acid and 1- (3-dimethylaminopropyl) -3-ethylcarbodiimide hydrochloride in dichloromethane, adding a proper amount of catalyst, stirring for reaction at room temperature, and separating and purifying to finally obtain the near-infrared fluorescent probe as claimed in claim 1.

3. The method for preparing a near-infrared fluorescent probe according to claim 2, wherein the catalyst in the step b is 4-dimethylaminopyridine.

4. Use of the near-infrared fluorescent probe according to claim 1 for detecting polysulfides.

5. The use of the near-infrared fluorescent probe according to claim 4 for detecting polysulfides, wherein the excitation light has a wavelength of 520nm to 580nm and the collected emission light has a wavelength of 630nm to 660 nm.

6. The use of the near-infrared fluorescent probe according to claim 4 for detecting polysulfide, wherein the solvent system used for detection is a mixed solution of dimethyl sulfoxide and water.

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