A flavonoid-based quenching fluorescent probe for specific detection of hypochlorous acid and its preparation method and application

Abstract

The present invention discloses a flavonoid-based quenching fluorescent probe for specific detection of hypochlorous acid, and a preparation method and application thereof. The fluorescent probe is: 3-hydroxy-2-(4-thiomorpholinephenyl)-6-(4-(1,2,2-triphenylvinyl)phenyl)-4H-methene-4-one. The present invention uses 1-(4-hydroxy-4'-(1,2,2-triphenylvinyl)-[1,1'-biphenyl]-3-yl)ethane-1-one as a raw material, and carries out aldol condensation reaction and cyclization reaction with 4-thiomorpholinebenzaldehyde to obtain a target compound. The compound can specifically identify hypochlorous acid and can sensitively detect the content of hypochlorous acid in a solution. As a fluorescent probe for detecting hypochlorous acid, it has the advantages of convenient synthesis, good selectivity, high sensitivity, and a detection limit as low as 5.7×10 ‑8 mol / L and many other advantages, and has good application prospects.

Description

Technical Field

[0001] The invention belongs to the technical field of fine organic synthesis, and relates to a flavonoid-based quenching fluorescent probe for specific detection of hypochlorous acid, and a preparation method and application thereof. Background Art

[0002] As an efficient and safe disinfectant and bactericide, hypochlorous acid is widely used as a food additive in the field of food processing. In addition, hypochlorous acid plays a vital role in the immune system and can resist the invasion of various pathogens and bacteria. However, the accumulation of excessive hypochlorous acid in organisms can cause oxidation of biomolecules, leading to tissue damage and inflammation, cardiovascular disease, rheumatoid arthritis, cancer and other major diseases. Therefore, it is urgent to establish an effective analytical method to monitor the changes in hypochlorous acid content in food and biological systems in real time.

[0003] At present, the main methods for detecting hypochlorous acid include electrochemical method, chromatography, high performance liquid chromatography, etc. Compared with traditional detection methods, fluorescent probes have unique advantages such as simple synthesis, high sensitivity, good selectivity, and good biocompatibility, and have developed into a very important analytical detection tool. There is no relevant report on the synthesis of a flavonoid-based quenching fluorescent probe for the specific detection of hypochlorous acid using 1-(4-hydroxy-4'-(1,2,2-triphenylvinyl)-[1,1'-biphenyl]-3-yl)ethane-1-one as a raw material. Summary of the invention

[0004] In view of the deficiencies in the prior art, the technical problem to be solved by the present invention is to provide a flavonoid-based quenching fluorescent probe for specific detection of hypochlorous acid, which can meet the use requirements of specific detection of hypochlorous acid. Another technical problem to be solved by the present invention is to provide a preparation method of 3-hydroxy-2-(4-thiomorpholinephenyl)-6-(4-(1,2,2-triphenylvinyl)phenyl)-4H-methen-4-one. The present invention also solves a technical problem of providing an application of the above-mentioned 3-hydroxy-2-(4-thiomorpholinephenyl)-6-(4-(1,2,2-triphenylvinyl)phenyl)-4H-methen-4-one in the specific detection of hypochlorous acid.

[0005] In order to solve the above technical problems, the technical solution adopted by the present invention is:

[0006] A flavonoid-based quenching fluorescent probe for specific detection of hypochlorous acid is 3-hydroxy-2-(4-thiomorpholinephenyl)-6-(4-(1,2,2-triphenylvinyl)phenyl)-4H-methen-4-one, and its structural formula is:

[0007]

[0008] The preparation method of the flavonoid-based quenching fluorescent probe for specific detection of hypochlorous acid comprises: using 1-(4-hydroxy-4'-(1,2,2-triphenylvinyl)-[1,1'-biphenyl]-3-yl)ethane-1-one as a raw material, carrying out aldol condensation reaction and cyclization reaction with 4-thiomorpholine benzaldehyde to obtain 3-hydroxy-2-(4-thiomorpholine phenyl)-6-(4-(1,2,2-triphenylvinyl) phenyl)-4H-methen-4-one.

[0009] The method for preparing the flavonoid-based quenching fluorescent probe for specific detection of hypochlorous acid specifically comprises the following steps:

[0010] (1) At 0°C, 0.1 mol of 1-(4-hydroxy-4'-(1,2,2-triphenylvinyl)-[1,1'-biphenyl]-3-yl)ethane-1-one, 0.15-0.2 mol of 4-thiomorpholinebenzaldehyde, 0.3-0.4 mol of sodium hydroxide, and 3200-4000 mL of anhydrous ethanol were added in sequence into a single-necked flask equipped with a stirrer; after reacting at room temperature overnight, 160-200 mL of 20% sodium hydroxide aqueous solution, 80-100 mL of 30% H 2 O 2 , stirring at room temperature for 6 to 8 hours;

[0011] (2) The reaction solution was poured into ice water, neutralized with dilute hydrochloric acid, and filtered to obtain a crude product of 3-hydroxy-2-(4-thiomorpholinophenyl)-6-(4-(1,2,2-triphenylvinyl)phenyl)-4H-methen-4-one;

[0012] (3) The crude product of 3-hydroxy-2-(4-thiomorpholinophenyl)-6-(4-(1,2,2-triphenylvinyl)phenyl)-4H-methen-4-one was chromatographed on a silica gel column (dichloromethane:methanol=100:1) to obtain a yellow powder 3-hydroxy-2-(4-thiomorpholinophenyl)-6-(4-(1,2,2-triphenylvinyl)phenyl)-4H-methen-4-one.

[0013] The compound 3-hydroxy-2-(4-thiomorpholinephenyl)-6-(4-(1,2,2-triphenylvinyl)phenyl)-4H-methen-4-one can selectively identify hypochlorous acid. It can emit very strong orange-yellow fluorescence under ultraviolet light with a wavelength of 365nm. After adding hypochlorous acid, the fluorescence color of the solution changes from orange-yellow to light green. It can be used as a quenching fluorescent probe for the specific detection of hypochlorous acid, and the minimum detection limit can reach 5.7×10 -8 mol / L.

[0014] Beneficial effects: Compared with the prior art, the present invention uses 1-(3-hydroxy-4'-(1,2,2-triphenylvinyl)-[1,1'-biphenyl]-4-yl)ethane-1-one as a raw material, and carries out aldol condensation reaction and cyclization reaction with 4-thiomorpholine benzaldehyde to obtain 3-hydroxy-2-(4-thiomorpholine phenyl)-6-(4-(1,2,2-triphenylvinyl) phenyl)-4H-methen-4-one. The compound can specifically identify hypochlorous acid and can sensitively detect the content of hypochlorous acid in the solution. As a fluorescent probe for detecting hypochlorous acid, it has the advantages of convenient synthesis, good selectivity, high sensitivity, and a detection limit as low as 5.7×10 -8 mol / L and many other advantages, and has good application prospects. BRIEF DESCRIPTION OF THE DRAWINGS

[0015] Figure 1 It is the fluorescence spectrum of 3-hydroxy-2-(4-thiomorpholinophenyl)-6-(4-(1,2,2-triphenylvinyl)phenyl)-4H-methen-4-one reacting with hypochlorous acid of different concentrations;

[0016] Figure 2 It is a fluorescence spectrum of 3-hydroxy-2-(4-thiomorpholinophenyl)-6-(4-(1,2,2-triphenylvinyl)phenyl)-4H-methen-4-one interacting with different analytes such as metal ions, anions, bioactive small molecules and reactive oxygen. DETAILED DESCRIPTION

[0017] The present invention will be further described below in conjunction with specific embodiments.

[0018] Example 1

[0019] Preparation of 3-hydroxy-2-(4-thiomorpholinophenyl)-6-(4-(1,2,2-triphenylvinyl)phenyl)-4H-methen-4-one, the reaction formula is as follows:

[0020]

[0021] The specific steps are as follows:

[0022] Preparation of 3-hydroxy-2-(4-thiomorpholinophenyl)-6-(4-(1,2,2-triphenylvinyl)phenyl)-4H-methen-4-one:

[0023] 0.1 mol of 1-(4-hydroxy-4'-(1,2,2-triphenylvinyl)-[1,1'-biphenyl]-3-yl)ethane-1-one, 0.2 mol of 4-thiomorpholinebenzaldehyde, 0.4 mol of sodium hydroxide, and 4000 mL of anhydrous ethanol were added to a single-necked flask equipped with a stirrer at 0°C. After reacting at room temperature overnight, 200 mL of 20% sodium hydroxide aqueous solution and 100 mL of 30% H 2 O 2 , stirred at room temperature for 8 hours, the reaction solution was poured into ice water, neutralized with dilute hydrochloric acid and filtered to obtain a crude product of 3-hydroxy-2-(4-thiomorpholinophenyl)-6-(4-(1,2,2-triphenylvinyl)phenyl)-4H-methen-4-one, and then chromatographed on a silica gel column (dichloromethane:methanol=100:1) to obtain a yellow powder 3-hydroxy-2-(4-thiomorpholinophenyl)-6-(4-(1,2,2-triphenylvinyl)phenyl)-4H-methen-4-one with a yield of 36% and a purity of 99.7%. 1 H NMR(600MHz,DMSO-d6)δ9.41(s,1H),8.19(d,J=8.6Hz,2H),8.10(d,J=8.3Hz, 1H),8.03(s,1H),7.73(d,J=8.4Hz,1H),7.67(d,J=7.9Hz,2H),7.18(dd,J=12. 4,8.0Hz,5H),7.15(d,J=7.2Hz,5H),7.11(d,J=8.4Hz,3H),7.04(dd,J=11.7,7 .5Hz, 4H), 6.99 (d, J = 7.4Hz, 2H), 3.94 (d, J = 5.3Hz, 4H), 3.16 (t, J = 5.0Hz, 4H). 13 C NMR(151MHz,DMSO-d6)δ172.48,155.27,148.72,146.50,144.51,144.23,143 .56,143.52,143.47,141.67,140.36,138.55,136.42,131.95,131.20,131.1 4,131.10,130.11,129.69,128.46,128.40,128.30,127.27,127.16,127.10,126.98,125.75,123.21,120.73,115.88,114.91,50.43,46.28,29.50,22.56.

[0024] Example 2

[0025] 3-Hydroxy-2-(4-thiomorpholinophenyl)-6-(4-(1,2,2-triphenylvinyl)phenyl)-4H-methen-4-one was prepared into 1×10 -5 M PBS buffer solution (pH = 7.4, 10 mM, 1% DMF), and hypochlorous acid was dissolved in PBS buffer to prepare concentrations of 0, 0.1×10 -5 ,0.2×10 -5 ,0.3×10 -5 ,0.4×10 -5 ,0.5×10 -5 ,0.6×10 -5 ,0.8×10 -5 ,1.0×10 -5 ,1.2×10 -5 ,1.5×10 -5 ,1.8×10 -5 ,2.1×10 -5 ,2.4×10 -5 ,2.7×10 -5 ,3.0×10 -5 ,3.5×10 -5 ,4.0×10 -5 M solution. The fluorescence emission spectrum of 3-hydroxy-2-(4-thiomorpholinophenyl)-6-(4-(1,2,2-triphenylvinyl)phenyl)-4H-methen-4-one in the presence of different concentrations of hypochlorous acid was measured on a fluorescence spectrophotometer by fluorescence spectrum titration method, such as Figure 1 The results show that as the concentration of hypochlorous acid in the solution gradually increases from 0 M to 4.0×10 -5 M, the fluorescence emission intensity of the compound at 589 nm gradually decreased. This shows that the compound can be used as a quenching fluorescent probe for sensitive detection of hypochlorous acid.

[0026] Example 3

[0027] 3-Hydroxy-2-(4-thiomorpholinophenyl)-6-(4-(1,2,2-triphenylvinyl)phenyl)-4H-methen-4-one was prepared into 1×10 -5 M PBS buffer solution (pH = 7.4, 10 mM, 1% DMF), and different analytes such as metal ions, anions, biothiols and reactive oxygen species were dissolved in PBS buffer to a concentration of 2.0×10 -4M solution. The fluorescence emission spectrum of 3-hydroxy-2-(4-thiomorpholinophenyl)-6-(4-(1,2,2-triphenylvinyl)phenyl)-4H-methen-4-one in the presence of different analytes such as metal ions, anions, bioactive small molecules and reactive oxygen species was measured on a fluorescence spectrophotometer by fluorescence spectrometry, such as Figure 2 As shown. After the compound reacted with hypochlorous acid, the fluorescence emission intensity at 589nm decreased significantly, while the addition of metal ions (Na + ,Fe 2+ ,Ag + ,Hg 2+ ,Al 3+ ), anions (Cl - ,Br - ,HPO 4 2- ,CO 3 2- ,SO 3 2- ), biothiols (Cys, Hcy, GSH) and reactive oxygen species (H 2 O 2 ,·OH, 1 O 2 ,ONOO - ,O 2 - ) and other different analytes, the fluorescence spectrum of the compound did not change significantly. This shows that the compound can be used as a quenching fluorescent probe for selective detection of hypochlorous acid.

Claims

1. A flavonoid-quenched fluorescent probe for specific detection of hypochlorous acid, characterized in that: The structural formula is:

2. The method for preparing the flavonoid-based quenching fluorescent probe for specific detection of hypochlorous acid according to claim 1, characterized in that: 1-(4-Hydroxy-4'-(1,2,2-triphenylvinyl)-[1,1'-biphenyl]-3-yl)ethane-1-one undergoes aldol condensation and cyclization with 4-thiomorpholinobenzaldehyde to give 3-hydroxy-2-(4-thiomorpholinophenyl)-6-(4-(1,2,2-triphenylvinyl)phenyl)-4H-methen-4-one.

3. The method for preparing a flavonoid-based quenching fluorescent probe for specific detection of hypochlorous acid according to claim 2, characterized in that: The specific steps include: (1) At 0°C, 0.1 mol of 1-(4-hydroxy-4'-(1,2,2-triphenylvinyl)-[1,1'-biphenyl]-3-yl)ethane-1-one, 0.15-0.2 mol of 4-thiomorpholine benzaldehyde, 0.3-0.4 mol of sodium hydroxide, and 3200-4000 mL of anhydrous ethanol were added in sequence into a single-necked flask equipped with a stirrer; after reacting at room temperature overnight, 160-200 mL of 20% sodium hydroxide aqueous solution and 80-100 mL of 30% H2O2 were added dropwise at 0°C, and stirred at room temperature for 6-8 hours; (2) The reaction solution was poured into ice water, neutralized with dilute hydrochloric acid, and filtered to obtain a crude product of 3-hydroxy-2-(4-thiomorpholinophenyl)-6-(4-(1,2,2-triphenylvinyl)phenyl)-4H-methen-4-one; (3) The crude product of 3-hydroxy-2-(4-thiomorpholinophenyl)-6-(4-(1,2,2-triphenylvinyl)phenyl)-4H-methen-4-one was purified by silica gel column chromatography to obtain yellow powder 3-hydroxy-2-(4-thiomorpholinophenyl)-6-(4-(1,2,2-triphenylvinyl)phenyl)-4H-methen-4-one.

4. Use of the flavonoid-quenched fluorescent probe 3-hydroxy-2-(4-thiomorpholinephenyl)-6-(4-(1,2,2-triphenylvinyl)phenyl)-4H-methen-4-one according to claim 1 in detecting hypochlorous acid.

5. The use according to claim 4, characterized in that: 3-Hydroxy-2-(4-thiomorpholinophenyl)-6-(4-(1,2,2-triphenylvinyl)phenyl)-4H-methen-4-one selectively recognized hypochlorous acid. Under irradiation with 365nm ultraviolet light, the fluorescence color of the probe solution changed from orange-yellow to light green.

6. The use according to claim 4, characterized in that: The content of hypochlorous acid in the solution was detected by 3-hydroxy-2-(4-thiomorpholinophenyl)-6-(4-(1,2,2-triphenylvinyl)phenyl)-4H-methen-4-one, and the detection limit was 5.7×10 - 8 mol / L.

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