CN116041979A - Carboxylic acid functionalized coumarin hemicyanine dye, and preparation method and application thereof - Google Patents
Carboxylic acid functionalized coumarin hemicyanine dye, and preparation method and application thereof Download PDFInfo
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- CN116041979A CN116041979A CN202310121084.3A CN202310121084A CN116041979A CN 116041979 A CN116041979 A CN 116041979A CN 202310121084 A CN202310121084 A CN 202310121084A CN 116041979 A CN116041979 A CN 116041979A
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- ZYGHJZDHTFUPRJ-UHFFFAOYSA-N coumarin Chemical compound C1=CC=C2OC(=O)C=CC2=C1 ZYGHJZDHTFUPRJ-UHFFFAOYSA-N 0.000 title claims abstract description 42
- 229960000956 coumarin Drugs 0.000 title claims abstract description 21
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- 238000002360 preparation method Methods 0.000 title claims abstract description 9
- 150000001732 carboxylic acid derivatives Chemical class 0.000 title abstract 3
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- 210000004027 cell Anatomy 0.000 claims description 21
- 150000001735 carboxylic acids Chemical class 0.000 claims description 18
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- 238000000034 method Methods 0.000 claims description 8
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 claims description 6
- NQRYJNQNLNOLGT-UHFFFAOYSA-N Piperidine Chemical compound C1CCNCC1 NQRYJNQNLNOLGT-UHFFFAOYSA-N 0.000 claims description 6
- 229910052757 nitrogen Inorganic materials 0.000 claims description 6
- QUDAPQXQGBIUAR-UHFFFAOYSA-N 2,3,3-trimethylindole-5-carboxylic acid Chemical compound C1=C(C(O)=O)C=C2C(C)(C)C(C)=NC2=C1 QUDAPQXQGBIUAR-UHFFFAOYSA-N 0.000 claims description 5
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- 238000010992 reflux Methods 0.000 claims description 4
- OEDBHOAJDADPMY-UHFFFAOYSA-N 7-(diethylamino)-2-oxochromene-4-carbaldehyde Chemical compound O=CC1=CC(=O)OC2=CC(N(CC)CC)=CC=C21 OEDBHOAJDADPMY-UHFFFAOYSA-N 0.000 claims description 3
- QXAMGWKESXGGNV-UHFFFAOYSA-N 7-(diethylamino)-1-benzopyran-2-one Chemical compound C1=CC(=O)OC2=CC(N(CC)CC)=CC=C21 QXAMGWKESXGGNV-UHFFFAOYSA-N 0.000 claims description 2
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- KZDCMKVLEYCGQX-UDPGNSCCSA-N 2-(diethylamino)ethyl 4-aminobenzoate;(2s,5r,6r)-3,3-dimethyl-7-oxo-6-[(2-phenylacetyl)amino]-4-thia-1-azabicyclo[3.2.0]heptane-2-carboxylic acid;hydrate Chemical compound O.CCN(CC)CCOC(=O)C1=CC=C(N)C=C1.N([C@H]1[C@H]2SC([C@@H](N2C1=O)C(O)=O)(C)C)C(=O)CC1=CC=CC=C1 KZDCMKVLEYCGQX-UDPGNSCCSA-N 0.000 description 2
- CURLTUGMZLYLDI-UHFFFAOYSA-N Carbon dioxide Chemical compound O=C=O CURLTUGMZLYLDI-UHFFFAOYSA-N 0.000 description 2
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- 150000001875 compounds Chemical class 0.000 description 2
- LOKCTEFSRHRXRJ-UHFFFAOYSA-I dipotassium trisodium dihydrogen phosphate hydrogen phosphate dichloride Chemical compound P(=O)(O)(O)[O-].[K+].P(=O)(O)([O-])[O-].[Na+].[Na+].[Cl-].[K+].[Cl-].[Na+] LOKCTEFSRHRXRJ-UHFFFAOYSA-I 0.000 description 2
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- NHBKXEKEPDILRR-UHFFFAOYSA-N 2,3-bis(butanoylsulfanyl)propyl butanoate Chemical compound CCCC(=O)OCC(SC(=O)CCC)CSC(=O)CCC NHBKXEKEPDILRR-UHFFFAOYSA-N 0.000 description 1
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- C—CHEMISTRY; METALLURGY
- C09—DYES; PAINTS; POLISHES; NATURAL RESINS; ADHESIVES; COMPOSITIONS NOT OTHERWISE PROVIDED FOR; APPLICATIONS OF MATERIALS NOT OTHERWISE PROVIDED FOR
- C09B—ORGANIC DYES OR CLOSELY-RELATED COMPOUNDS FOR PRODUCING DYES, e.g. PIGMENTS; MORDANTS; LAKES
- C09B23/00—Methine or polymethine dyes, e.g. cyanine dyes
- C09B23/10—The polymethine chain containing an even number of >CH- groups
- C09B23/105—The polymethine chain containing an even number of >CH- groups two >CH- groups
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- C—CHEMISTRY; METALLURGY
- C09—DYES; PAINTS; POLISHES; NATURAL RESINS; ADHESIVES; COMPOSITIONS NOT OTHERWISE PROVIDED FOR; APPLICATIONS OF MATERIALS NOT OTHERWISE PROVIDED FOR
- C09K—MATERIALS FOR MISCELLANEOUS APPLICATIONS, NOT PROVIDED FOR ELSEWHERE
- C09K11/00—Luminescent, e.g. electroluminescent, chemiluminescent materials
- C09K11/06—Luminescent, e.g. electroluminescent, chemiluminescent materials containing organic luminescent materials
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N21/00—Investigating or analysing materials by the use of optical means, i.e. using sub-millimetre waves, infrared, visible or ultraviolet light
- G01N21/62—Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light
- G01N21/63—Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light optically excited
- G01N21/64—Fluorescence; Phosphorescence
- G01N21/6428—Measuring fluorescence of fluorescent products of reactions or of fluorochrome labelled reactive substances, e.g. measuring quenching effects, using measuring "optrodes"
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- C09K2211/00—Chemical nature of organic luminescent or tenebrescent compounds
- C09K2211/10—Non-macromolecular compounds
- C09K2211/1018—Heterocyclic compounds
- C09K2211/1025—Heterocyclic compounds characterised by ligands
- C09K2211/1029—Heterocyclic compounds characterised by ligands containing one nitrogen atom as the heteroatom
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- C09—DYES; PAINTS; POLISHES; NATURAL RESINS; ADHESIVES; COMPOSITIONS NOT OTHERWISE PROVIDED FOR; APPLICATIONS OF MATERIALS NOT OTHERWISE PROVIDED FOR
- C09K—MATERIALS FOR MISCELLANEOUS APPLICATIONS, NOT PROVIDED FOR ELSEWHERE
- C09K2211/00—Chemical nature of organic luminescent or tenebrescent compounds
- C09K2211/10—Non-macromolecular compounds
- C09K2211/1018—Heterocyclic compounds
- C09K2211/1025—Heterocyclic compounds characterised by ligands
- C09K2211/1088—Heterocyclic compounds characterised by ligands containing oxygen as the only heteroatom
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Abstract
The invention belongs to the technical field of fluorescent probes, and particularly relates to a carboxylic acid functionalized coumarin hemicyanine dye, and a preparation method and application thereof. In order to overcome a plurality of defects existing in the conventional tumor examination means and provide a means for realizing early and accurate diagnosis and treatment of tumors, the carboxylic acid functionalized coumarin hemicyanine dye developed by the invention is used for distinguishing lung cancer cells (HepG 2) from normal lung cells (LO 2) and distinguishing liver cancer tissues and normal organ tissues of tumor-bearing mice, and has the characteristics of short analysis time, no cleaning and the like.
Description
Technical Field
The invention belongs to the technical field of fluorescent probes, and particularly relates to a carboxylic acid functionalized coumarin hemicyanine dye, and a preparation method and application thereof.
Background
Cancer is a disease with uncontrolled cell growth and cell division, the incidence and death rate are increased year by year in recent years, and early detection, early diagnosis and early treatment of cancer are main means for improving the cure rate of malignant tumor, improving the life quality of patients and prolonging the life cycle. Among them, early diagnosis is the basis of early treatment. Currently, conventional means for tumor diagnosis include X-ray imaging, angiography, CT examination, magnetic resonance imaging, ultrasound examination, endoscopy, biomarker examination, pathology examination, etc., however, due to the lack of sufficient specificity and sensitivity of the conventional means, in addition to the small tissue volume and irregular morphology before the lesion of cancer, the tumor can be accurately diagnosed only when the metastasis and diffusion occur in the middle to late stages of the tumor. Compared with the prior art, the fluorescent probe imaging technology has the advantages of high sensitivity, low cost, short analysis time, simple operation, visualization, no ionizing radiation, real-time multicolor image and the like, can overcome a plurality of defects existing in the conventional tumor examination means, and provides a means for realizing early and accurate diagnosis and treatment of tumors.
Disclosure of Invention
In view of the above problems, the present invention provides a carboxylic acid functionalized coumarin hemicyanine dye and a preparation method thereof, and is used for distinguishing lung cancer cells (HepG 2) from normal lung cells (LO 2), and distinguishing liver cancer tissues and normal organ tissues of tumor-bearing mice.
In order to achieve the above purpose, the present invention adopts the following technical scheme:
a carboxylic acid functionalized coumarin hemicyanine dye, which has the structural formula:
a method for preparing coumarin hemicyanine dye functionalized by carboxylic acid, comprising the following steps:
(1) Under the protection of nitrogen, 2, 3-trimethyl-3H-indole-5-carboxylic acid (compound 1) and methyl iodide are dissolved in acetonitrile for reaction, after the reaction is finished, the reaction is cooled to room temperature, a large amount of solids are generated, and the obtained solids are filtered, washed and dried to obtain compound 2;
(2) Under the protection of nitrogen, dissolving a compound 2, 7- (diethylamino) coumarin-3-formaldehyde (compound 3) and piperidine in absolute ethyl alcohol for reflux reaction, cooling to room temperature after the reaction is finished, generating a large amount of solids, and filtering, washing and drying the solids to obtain a target product CCHC;
the structural formula of the compound 2 is as follows:
further, the molar ratio of 2, 3-trimethyl-3H-indole-5-carboxylic acid to methyl iodide in the step (1) is 1:1.
Further, the temperature of the reaction in the step (1) is 90 ℃ and the time is 30h.
Further, the molar ratio of compound 2 to 7- (diethylamino) coumarin-3-carbaldehyde in step (2) is 1:1.
Further, the temperature of the reflux reaction in the step (2) is 78.4 ℃ and the time is 12 hours.
The application of the coumarin hemicyanine dye functionalized by carboxylic acid is characterized in that liver cancer cells (HepG 2 cells) and normal liver cells (LO 2 cells) are distinguished, and liver cancer tissues and normal organ tissues of tumor-bearing mice are distinguished.
Compared with the prior art, the invention has the following advantages:
early diagnosis of cancer is the basis of early treatment, whereas traditional means of tumor diagnosis, such as X-ray imaging, CT examination, magnetic resonance imaging, cytological examination in pathological examination, biopsy, and the like, lack accuracy and sensitivity, and can achieve accurate diagnosis only when metastasis and spread occur in the middle to late stages of tumor development. The carboxylic acid functionalized coumarin hemicyanine dye developed by the invention has the characteristics of short analysis time, no need of cleaning and the like, and is expected to provide a visual method for early diagnosis and treatment of tumors.
Drawings
FIG. 1 is an NMR and HRMS plot of compound 2 of the present invention;
FIG. 2 is a NMR and HRMS plot of the dye CCHC of the present invention;
FIG. 3 is a fluorescence imaging diagram after 5 minutes of treatment of HepG2 and LO2 cells with CCHC (2.0. Mu.M), respectively, at a collection wavelength of 570-750nm (λex=561 nm), scale 20 μm;
FIG. 4 is a fluorescence imaging image of tumor tissue sections and normal organ tissue sections of tumor-bearing mice treated with CCHC (2.0. Mu.M) for 5 minutes, with a collection wavelength of 570-750nm (λex=561 nm), scale 20. Mu.m.
Detailed Description
Example 1
A carboxylic acid functionalized coumarin hemicyanine dye, which has the structural formula:
a method for preparing coumarin hemicyanine dye functionalized by carboxylic acid, comprising the following steps:
(1) 2, 3-trimethyl-3H-indole-5-carboxylic acid (Compound 1,1.014g,4.99 mmol) and methyl iodide (1.014 g,4.99 mmol) were dissolved in acetonitrile (20 mL) under nitrogen protection, reacted at 90℃for 30 hours, cooled to room temperature after the completion of the reaction, and a large amount of solid was formed, and the obtained solid was filtered, washed and dried to give Compound 2 (0.96 g, yield 56%), FIG. 1 being NMR and HRMS images of Compound 2;
1 H NMR(600MHz,DMSO-d 3 )δ8.38(s,1H),8.20(d,J＝1.2Hz,1H),8.03(d,J＝8.4Hz,1H),3.99(s,3H),2.81(s,3H),1.57(s,6H)；ESI-MS[M] + :calculated for218.1176,found 218.1173.
(2) Compound 2 (1.0 mmol), 7- (diethylamino) coumarin-3-carbaldehyde (compound 3,1.0 mmol) and a few drops of piperidine were dissolved in absolute ethanol (30 mL) under nitrogen protection, and the mixture was refluxed at 78.4 ℃ for 12 hours, cooled to room temperature after the reaction was completed, and a large amount of solid was formed, and the solid was filtered, washed (ethanol) and dried to give the objective CCHC (0.376 g, yield 49.2%) as shown in fig. 2, which is NMR and HRMS chart of CCHC.
1 H NMR(600MHz,CD 3 OD)δ8.61(s,1H),8.40(d,J＝15.6Hz,1H),8.32(s,1H),8.28(d,J＝23.4Hz,1H),8.03(d,J＝15.6Hz,1H),7.81(d,J＝8.4Hz,1H),7.64(d,J＝9.0Hz,1H),6.97(m,1H),6.68(s,1H),4.06(s,3H),3.67(m,4H),1.88(s,6H),1.32(t,J＝7.2Hz,J＝7.2Hz,6H)； 13 C NMR(150MHz,CD 3 OD)δ183.25,167.35,159.99,158.41,155.13,151.97,145.20,143.04,132.64,131.35,130.85,123.49,113.57,112.45,111.48,110.38,109.53,99.53,51.53,45.19,32.70,25.50,11.42；ESI-MS[M + ]:calculated for 445.2122,found 445.2115.
Example 2
1. Preparation of test solutions
Dye CCHC was made up with DMSO (containing 2% acetic acid) into a 2mM stock solution, which was then diluted to the test concentration with the solvent to be tested.
2. Cell imaging studies
All cell lines were purchased from GeneFull biotechnology limited (china). HepG2 cells were cultured in DMEM medium containing 10% FBS (fetal bovine serum), 100U/mL penicillin G sodium and 100. Mu.g/mL streptomycin, and LO2 cells were cultured in RPMI1640 medium containing 10% FBS (fetal bovine serum), 100U/mL penicillin G sodium and 100. Mu.g/mL streptomycin, at 37℃in a 5% carbon dioxide humidified environment. Before performing a cell image experiment, different cells were placed on a 30mm glass-bottomed cell culture dish in advance, allowed to stand for 12 hours until the cells adhere, washed 3 times with Phosphate Buffered Saline (PBS), and then incubated for 5 minutes, and fluorescence imaging was performed using a Ceiss LMS 710 confocal microscope. As shown in fig. 3, hepG2 cells treated with CCHC exhibited strong fluorescent signals on the cell membrane; while the LO2 cells treated with CCHC hardly had any fluorescence signal, which indicates that the liver cancer cells can be distinguished from normal liver cells according to the intensity of fluorescence signal on the cell membrane.
3. Tissue slice imaging studies
All animal experiments were performed according to the relevant laws and guidelines issued by the university of Shanxi ethical Committee, BALB/c male nude mice (6-8 weeks old) were purchased from Beijing Vitolihua laboratory animal technologies Co. HepG2 cells (1X 10) 6 Individual cells) were subcutaneously injected into the root of left leg of nude mice, 15 days after inoculation, tumor-bearing mice were obtained, the tumor-bearing mice were sacrificed by mild anesthesia, five viscera and tumors were taken, and sections with a thickness of 10 μm were prepared on a frozen microtome, sections of different tissues were treated with probes for 5 minutes, and their fluorescence was compared on laser confocalDensity. As shown in FIG. 4, the fluorescence intensity of cancer tissue is significantly higher than that of heart, liver, spleen, lung and kidney tissues, and the above results indicate that CCHC can distinguish tumor tissue from normal tissue of tumor-bearing mice.
In summary, the invention develops a carboxylic acid functionalized coumarin hemicyanine dye (CCHC), which can distinguish liver cancer cells (HepG 2 cells) from normal liver cells (LO 2 cells), and can distinguish liver cancer tissues and normal organ tissues of tumor-bearing mice, so the dye CCHC has potential biological application value.
The foregoing description of the preferred embodiments of the invention is not intended to be limiting, but rather is intended to cover all modifications, equivalents, alternatives, and improvements that fall within the spirit and scope of the invention.
Claims (8)
1. A carboxylic acid functionalized coumarin hemicyanine dye, characterized in that the dye has the structural formula:
2. a process for the preparation of a carboxylic acid functionalized coumarin hemicyanine dye as claimed in claim 1, comprising the steps of:
(1) Under the protection of nitrogen, 2, 3-trimethyl-3H-indole-5-carboxylic acid and methyl iodide are dissolved in acetonitrile for reaction, the reaction is cooled to room temperature after the reaction is finished, a large amount of solids are generated, and the obtained solids are filtered, washed and dried to obtain a compound 2;
(2) Under the protection of nitrogen, dissolving a compound 2, 7- (diethylamino) coumarin-3-formaldehyde and piperidine in absolute ethyl alcohol for reflux reaction, cooling to room temperature after the reaction is finished, generating a large amount of solids, filtering, washing and drying the solids to obtain a target product;
the structural formula of the compound 2 is as follows:
3. the method for preparing a carboxylic acid functionalized coumarin hemicyanine dye according to claim 2, wherein the molar ratio of 2, 3-trimethyl-3H-indole-5-carboxylic acid to methyl iodide in step (1) is 1:1.
4. The method for preparing the coumarin hemicyanine dye functionalized by carboxylic acid according to claim 2, wherein the reaction temperature in the step (1) is 90 ℃ and the reaction time is 30h.
5. The method for preparing a carboxylic acid functionalized coumarin hemicyanine dye according to claim 2, wherein the molar ratio of compound 2 to 7- (diethylamino) coumarin-3-carbaldehyde in step (2) is 1:1.
6. The method for preparing the coumarin hemicyanine dye functionalized by carboxylic acid according to claim 2, wherein the reflux reaction in the step (2) is carried out at 78.4 ℃ for 12 hours.
7. Use of a carboxylic acid functionalized coumarin hemicyanine dye as claimed in claim 1, for the preparation of a reagent for differentiating liver cancer cells from normal liver cells.
8. Use of a carboxylic acid functionalized coumarin hemicyanine dye as claimed in claim 1, for the preparation of a reagent for differentiating liver cancer tissue from normal organ tissue of tumor-bearing mice.
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Citations (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US5223382A (en) * | 1991-12-02 | 1993-06-29 | Fuji Photo Film Co., Ltd. | Silver halide photographic material |
| CN112239462A (en) * | 2019-07-18 | 2021-01-19 | 南京理工大学 | Cyanide receptor compound based on coumarin and carboxylic acid indole, preparation method and application thereof |
| CN114940685A (en) * | 2022-05-05 | 2022-08-26 | 山东第一医科大学(山东省医学科学院) | Visual fluorescent anti-tumor drug and application thereof in anti-tumor field |
| CN115650960A (en) * | 2022-02-22 | 2023-01-31 | 大连理工大学 | Carboxylesterase 1 specific near-infrared fluorescent probe for pesticide residue detection and application thereof |
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2023
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| US5223382A (en) * | 1991-12-02 | 1993-06-29 | Fuji Photo Film Co., Ltd. | Silver halide photographic material |
| CN112239462A (en) * | 2019-07-18 | 2021-01-19 | 南京理工大学 | Cyanide receptor compound based on coumarin and carboxylic acid indole, preparation method and application thereof |
| CN115650960A (en) * | 2022-02-22 | 2023-01-31 | 大连理工大学 | Carboxylesterase 1 specific near-infrared fluorescent probe for pesticide residue detection and application thereof |
| CN114940685A (en) * | 2022-05-05 | 2022-08-26 | 山东第一医科大学(山东省医学科学院) | Visual fluorescent anti-tumor drug and application thereof in anti-tumor field |
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