A flavonol fluorescent probe for detecting biological amines and a preparation method and application thereof
By preparing CTBA-based cellulose fluorescent films, the problems of complex synthesis and insufficient biocompatibility of existing fluorescent sensors were solved, enabling rapid and non-invasive detection of biogenic amines with good detection performance.
Patent Information
- Authority / Receiving Office
- CN · China
- Patent Type
- Applications(China)
- Current Assignee / Owner
- NANJING FORESTRY UNIV
- Filing Date
- 2025-01-06
- Publication Date
- 2026-07-07
AI Technical Summary
Existing fluorescent sensors for detecting biogenic amines suffer from complex synthesis and are difficult to prepare. Furthermore, existing thin-film fluorescent probes are insufficient in terms of biocompatibility and degradability, making it difficult to achieve rapid and non-invasive food quality monitoring.
4-(2-(2,4,6-trimethoxyphenyl)-3-((4-cyanobenzoyl)oxy)-chromone-7-yl)benzoic acid (CTBA) was used as a flavonol fluorescent probe. A fluorescent film was prepared by binding it with cellulose, and the specific fluorescence signal changes emitted by the film under 365 nm ultraviolet light were used to detect biogenic amines.
An easily prepared fluorescent probe was developed, enabling rapid and non-invasive detection of biogenic amines under 365 nm ultraviolet light, with good selectivity and sensitivity, and a detection limit of 3.6 × 10⁻⁷ M.
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Abstract
Description
Technical Field
[0001] This invention belongs to the field of fluorescence detection technology, and relates to a flavonol fluorescent probe for detecting biogenic amines, its preparation method and application. Background Technology
[0002] Biogenic amines are a class of nitrogen-containing organic compounds, primarily produced from amino acids through decarboxylation or transamination reactions. They play important physiological roles in organisms, such as participating in nerve conduction, cell growth, and immune regulation. However, excessive accumulation or intake of biogenic amines can cause serious harm to human health. Particularly in food, an increase in biogenic amines (cadaverine) usually indicates food spoilage and quality decline. Excessive intake can lead to symptoms such as food poisoning, difficulty breathing, headache, high blood pressure, and vomiting. Furthermore, they may react with nitrites during cooking to form carcinogenic nitrosamines. Therefore, monitoring the content of biogenic amines in food is crucial for ensuring consumer health and food safety.
[0003] Fluorescent sensors exhibit significant advantages in detecting biogenic amines due to their high sensitivity, rapid response, and intuitive signal output. These sensors can quantitatively analyze the concentration of biogenic amines by detecting changes in fluorescence signals, thus providing real-time and accurate monitoring results. Currently, developed fluorescent sensors for monitoring biogenic amines include rhodamine derivatives, Schiff bases, and pyrrolopyrrolocyanine derivatives. However, compared to these relatively complex fluorescent small molecules, the ease of synthesis of flavonol fluorescent small molecules greatly reduces the time and cost of developing high-performance fluorescent sensors. Compared to solution detection, thin-film-based fluorescent probes offer non-invasiveness and rapid response. Cellulose, as a highly biocompatible biopolymer, is not only harmless to humans but also possesses excellent biodegradability. Furthermore, cellulose has superior processing properties and can be easily converted into different material forms, such as printing inks, coatings, flexible films, and nanofiber membranes. Therefore, the preparation of cellulose-based fluorescent sensing films shows great potential for real-time, visual monitoring of food quality. Summary of the Invention
[0004] To address the shortcomings of existing technologies, the technical problem this invention aims to solve is to provide a flavonol-based fluorescent probe for detecting biogenic amines. This fluorescent probe, in DMF / PBS solution, emits a pale blue fluorescence under 365 nm ultraviolet light, but the solution emits a bright yellow fluorescence upon the addition of biogenic amines, thus enabling its use in the detection of biogenic amines. Another technical problem this invention aims to solve is to provide a method for preparing the aforementioned flavonol-based fluorescent probe. A further technical problem this invention aims to solve is to provide an application of the aforementioned flavonol-based fluorescent probe.
[0005] To solve the above-mentioned technical problems, the technical solution adopted by the present invention is as follows:
[0006] A flavonol-based fluorescent probe for detecting biogenic amines is 4-(2-(2,4,6-trimethoxyphenyl)-3-((4-cyanobenzoyl)oxy)-chromone-7-yl)benzoic acid, abbreviated as CTBA, with the following structural formula:
[0007]
[0008] The synthetic route for the flavonol fluorescent probe is as follows:
[0009]
[0010] The method for preparing the flavonol fluorescent probe is characterized by comprising the following steps:
[0011] (1) Using 3′-hydroxy-4′-acetyl-[1,1′-biphenyl]-4-carboxylic acid as a starting material, it undergoes an aldol condensation reaction with 2,4,6-trimethoxybenzaldehyde under the action of an alkaline catalyst to prepare 3′-hydroxy-4′-(3-(2,4,6-trimethoxyphenyl)acryloyloxy)-[1,1′-biphenyl]-4-carboxylic acid, abbreviated as HTC;
[0012] (2) Under alkaline conditions, compound HTC is oxidatively cyclized to obtain 4-(2-(2,4,6-trimethoxyphenyl)-3-hydroxy-chromone-7-yl)benzoic acid, abbreviated as HTBA;
[0013] (3) HTBA and 4-cyanobenzoyl chloride are catalytically esterified to obtain 4-(2.(2,4,6-trimethoxyphenyl)-3-((4-cyanobenzoyl)oxy).chromone-7-yl)benzoic acid, abbreviated as CTBA.
[0014] The preparation steps of compound HTC in step (1) are as follows:
[0015] 1) Add 0.5 g of 3′-hydroxy-4′-acetyl-[1,1′-biphenyl]-4-carboxylic acid, 40-60 mL of tert-butanol and 0.6-1.0 g of potassium tert-butoxide to a 100 mL flask equipped with a stirrer, and stir the mixture at room temperature for 20-24 h.
[0016] 2) After the solvent is removed by distillation, the residue is dissolved in anhydrous ethanol. After adding 3-5 mL of concentrated hydrochloric acid, the precipitated solid is filtered, washed with water, and dried. The obtained solid is recrystallized in ethanol to obtain compound HTC.
[0017] The preparation steps of compound HTBA in step (2) are as follows:
[0018] 1) Add 0.868g HTC, 40-50mL ethanol, 0.8-1.5g potassium hydroxide and 2-3mL hydrogen peroxide to a 100mL flask equipped with a stirrer, and stir the mixture at room temperature for 20h.
[0019] 2) Add 40-60 mL of water to the reaction solution, then add 3-5 mL of concentrated hydrochloric acid. The precipitated solid is filtered, washed with water, and dried. The obtained solid is recrystallized in ethanol to obtain the compound HTBA.
[0020] The preparation steps of compound CTBA in step (3) are as follows:
[0021] 1) Add 0.896g of compound HTBA, 0.33g of 4-cyanobenzoyl chloride, 10-15mL of dichloromethane and 0.3-0.6mL of triethylamine to a 100mL three-necked flask equipped with a stirrer and thermometer, and stir the mixture in an ice bath for 5-10 hours.
[0022] 2) After the solvent was removed by distillation, the residue was purified by column chromatography (DCM:MeOH = 80:1, v / v) to obtain compound CTBA.
[0023] The application of the compound CTBA in the detection of cadaverine.
[0024] In the aforementioned application, under 365nm ultraviolet light irradiation, the compound CTBA emits a light blue fluorescence in a DMF / 10mM PBS buffer system (v / v = 1 / 9). After the addition of biogenic amine, the fluorescence color of the solution changes to bright yellow.
[0025] The application of the compound CTBA as a fluorescent probe in the detection of biogenic amines.
[0026] This invention uses 3′-hydroxy-4′-acetyl-[1,1′-biphenyl]-4-carboxylic acid as a starting material and reacts it with 2,4,6-trimethoxybenzaldehyde via an aldol condensation reaction to obtain compound HTC. Compound HTC is then subjected to oxidative cyclization under alkaline conditions to obtain compound HTBA. HTBA is further esterified with 4-cyanobenzoyl chloride to obtain compound CTBA. Under 365 nm ultraviolet light irradiation, the addition of biogenic amine to a DMF / PBS buffer solution (10 mM, v / v = 1 / 9) of compound CTBA causes the fluorescence color of the solution to change from light blue to bright yellow, indicating a detection limit of 3.6 × 10⁻⁶ for cadaverine. -7 M can be used as a fluorescent probe for the detection of cadaverine.
[0027] Beneficial effects: Compared with the prior art, the advantages of this invention are as follows: Flavonol fluorescent probes are easy to prepare. The cellulose-based fluorescent film prepared by spin-coating the compound CTBA onto cellulose macromolecules exhibits good luminescence performance and structural stability. When cadaverine is added under 365nm ultraviolet light irradiation, the fluorescence color of this cellulose-based flavonol fluorescent film gradually changes from light blue to bright yellow, showing promising application prospects as a fluorescent probe for detecting cadaverine. Attached Figure Description
[0028] Figure 1 These are the infrared spectra of probe cellulose membrane + cadaverine, probe cellulose membrane, ethyl cellulose, and probe compound CTBA;
[0029] Figure 2 The fluorescence spectra of compound CTBA in DMF / 10mM PBS buffer (v / v = 1 / 9) solution before and after the addition of cadaverine are shown.
[0030] Figure 3 The fluorescence spectra of different amine compounds added to a DMF / 10mM PBS buffer solution (v / v = 1 / 9) containing compound CTBA.
[0031] Figure 4 The fluorescence spectra of compound CTBA in DMF / 10mM PBS buffer (v / v = 1 / 9) with different concentrations of cadaverine added are shown. Detailed Implementation
[0032] To make the above-mentioned objects, features and advantages of the present invention more apparent and understandable, the specific embodiments of the present invention will be described in detail below with reference to the examples in the specification.
[0033] Example 1
[0034] Considering the good processability of ethyl cellulose, fluorescent probe cellulose membranes were prepared by spin coating, and the responsiveness of the probe cellulose membranes to cadaverine was investigated. 10 mg CTBA and 490 mg ethyl cellulose were dissolved in 8–12 mL of CH₂Cl₂. Then, 200 μL of the solution was dropped onto the center of a silicon wafer on a spin coater, and the wafer was spin-coated repeatedly at 500 rpm for 10 s and 3000 rpm for 60 s to obtain a transparent fluorescent probe cellulose membrane. The probe cellulose membrane was then vacuum-dried at 45 °C for 30 min.
[0035] Figure 1 These are the FTIR spectra of probe cellulose membrane + cadaverine, probe cellulose membrane, ethyl cellulose, and the probe. The FTIR spectra of probe cellulose membrane + cadaverine, probe cellulose membrane, and ethyl cellulose are at 3475 cm⁻¹. -1The peak at 2975 cm⁻¹ is strong and broad, which is due to the OH stretching vibration of the hydroxyl group; -1 and 2930cm -1 It is the CH stretching vibration of -CH3 and -CH2-; probe cellulose membrane + cadaverine, probe cellulose membrane 1654cm -1 and the 1608cm on the probe -1 The stretching vibration of C=C on the benzene ring skeleton; the probe cellulose membrane reacts with cadaverine at 1704 cm⁻¹. -1 The disappearance of the C=O stretching vibration peak indicates that the ester bonds in the probe cellulose membrane broke after the reaction.
[0036] Example 2
[0037] Compound CTBA was added to a DMF / 10mM PBS buffer solution (v / v = 1 / 9) to prepare a 10μM DMF / 10mM PBS buffer solution (v / v = 1 / 9). The fluorescence emission spectrum in the DMF / 10mM PBS buffer system (v / v = 1 / 9) was measured, as follows: Figure 2 As shown. The results indicate that in the DMF / 10mM PBS buffer (v / v = 1 / 9) system, the fluorescence intensity was weak without the addition of cadaverine, but increased sharply after the addition of cadaverine, with a maximum emission wavelength of 510 nm (excitation wavelength of 365 nm, excitation slit bandwidth of 7.5 nm, and emission slit bandwidth of 5.0 nm).
[0038] Compound CTBA was added to a DMF / 10mM PBS buffer (v / v = 1 / 9) system to prepare a 10μM DMF / 10mM PBS buffer (v / v = 1 / 9) solution. One portion was used as a blank sample, and the other portions were prepared by adding cadaverine, putrescine, hydrazine, spermine, histamine, tryptamine, piperidine, ammonia, and triethylamine, respectively. The fluorescence emission spectra of the solutions were measured, and the results are as follows. Figure 3 As shown. From Figure 3 It was found that the fluorescence intensity of the solution significantly increased after the addition of biogenic amines (cadaverine, putrescine, hydrazine, spermine, histamine, tryptamine). However, the fluorescence intensity of the solution changed very little upon the addition of other amine compounds. This indicates that compound CTBA exhibits good selectivity for biogenic amines.
[0039] Compound CTBA was added to a DMF / 10mM PBS buffer (v / v = 1 / 9) system to prepare a 10μM DMF / 10mM PBS buffer (v / v = 1 / 9) solution. The fluorescence emission spectra of different concentrations of cadaverine were then measured. Figure 4As shown in the figure, the fluorescence signal intensity of compound CTBA at 510 nm gradually increased with increasing cadaverine concentration, indicating that compound CTBA can be used to detect cadaverine concentration in solution, with a detection limit of 3.6 × 10⁻⁶. -7 M.
Claims
1. A flavonol-based fluorescent probe for detecting biogenic amines, characterized in that, The structural formula is:
2. The method for preparing the flavonol fluorescent probe according to claim 1, characterized in that, Includes the following steps: (1) Using 3′-hydroxy-4′-acetyl-[1,1′-biphenyl]-4-carboxylic acid as a starting material, it undergoes an aldol condensation reaction with 2,4,6-trimethoxybenzaldehyde under the action of an alkaline catalyst to prepare 3′-hydroxy-4′-(3-(2,4,6-trimethoxyphenyl)acryloyloxy)-[1,1′-biphenyl]-4-carboxylic acid (HTC); (2) Under alkaline conditions, compound HTC was oxidatively cyclized to prepare 4-(2-(2,4,6-trimethoxyphenyl)-3-hydroxy-chromone-7-yl)benzoic acid (HTBA); (3) HTBA and 4-cyanobenzoyl chloride were catalytically esterified to obtain 4-(2-(2,4,6-trimethoxyphenyl)-3-((4-cyanobenzoyl)oxy)-chromone-7-yl)benzoic acid (CTBA).
3. The method for preparing flavonol fluorescent probes according to claim 2, characterized in that, The preparation steps of compound HTC in step (1) are as follows: 1) Add 0.5 g of 3′-hydroxy-4′-acetyl-[1,1′-biphenyl]-4-carboxylic acid, 40-60 mL of tert-butanol and 0.6-1.0 g of potassium tert-butoxide to a 100 mL flask equipped with a stirrer, and stir the reaction at room temperature for 20-24 h. 2) After the solvent is removed by distillation, the residue is dissolved in anhydrous ethanol. After adding 3-5 mL of concentrated hydrochloric acid, the precipitated solid is filtered, washed with water, and dried. The obtained solid is recrystallized in ethanol to obtain compound HTC.
4. The method for preparing the flavonol fluorescent probe according to claim 2, characterized in that, The preparation steps of compound HTBA in step (2) are as follows: 1) Add 0.868g HTC, 40-50mL ethanol, 0.8-1.5g potassium hydroxide and 2-3mL hydrogen peroxide to a 100mL flask equipped with a stirrer, and stir the mixture at room temperature for 20-24 hours. 2) Add 40-60 mL of water to the reaction solution, then add 3-5 mL of concentrated hydrochloric acid. The precipitated solid is filtered, washed with water, and dried. The obtained solid is recrystallized in ethanol to obtain the compound HTBA.
5. The method for preparing the flavonol fluorescent probe according to claim 2, characterized in that, The preparation steps of compound CTBA in step (3) are as follows: 1) Add 0.896g of compound HTBA, 0.33g of 4-cyanobenzoyl chloride, 10-15mL of dichloromethane and 0.3-0.6mL of triethylamine to a 100mL three-necked flask equipped with a stirrer and thermometer, and stir the mixture in an ice bath for 5-10 hours. 2) After the solvent was removed by distillation, the residue was purified by column chromatography (DCM:MeOH = 80:1, v / v) to obtain compound CTBA.
6. The use of the compound CTBA according to claim 1 in the detection of biogenic amines.
7. The application according to claim 5, characterized in that, Under 365nm ultraviolet light irradiation, the compound CTBA emits a light blue fluorescence in a DMF / 10mM PBS buffer system (v / v = 1 / 9). After the addition of biogenic amine, the fluorescence color of the solution turns bright yellow.
8. The application of the compound CTBA according to claim 1 as a fluorescent probe in the detection of biogenic amines.