CN115304572B - Flavonoid fluorescent probe for detecting hydrazine and preparation method and application thereof - Google Patents
Flavonoid fluorescent probe for detecting hydrazine and preparation method and application thereof Download PDFInfo
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- OAKJQQAXSVQMHS-UHFFFAOYSA-N Hydrazine Chemical compound NN OAKJQQAXSVQMHS-UHFFFAOYSA-N 0.000 title claims abstract description 142
- 239000007850 fluorescent dye Substances 0.000 title claims abstract description 88
- 229930003935 flavonoid Natural products 0.000 title claims abstract description 58
- 235000017173 flavonoids Nutrition 0.000 title claims abstract description 58
- 150000002215 flavonoids Chemical class 0.000 title claims abstract description 56
- 238000002360 preparation method Methods 0.000 title claims abstract description 10
- 238000001514 detection method Methods 0.000 claims abstract description 24
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 claims abstract description 18
- JECYUBVRTQDVAT-UHFFFAOYSA-N 2-acetylphenol Chemical compound CC(=O)C1=CC=CC=C1O JECYUBVRTQDVAT-UHFFFAOYSA-N 0.000 claims abstract description 16
- OTXINXDGSUFPNU-UHFFFAOYSA-N 4-tert-butylbenzaldehyde Chemical compound CC(C)(C)C1=CC=C(C=O)C=C1 OTXINXDGSUFPNU-UHFFFAOYSA-N 0.000 claims abstract description 16
- -1 flavonoid compound Chemical class 0.000 claims abstract description 14
- WETWJCDKMRHUPV-UHFFFAOYSA-N acetyl chloride Chemical compound CC(Cl)=O WETWJCDKMRHUPV-UHFFFAOYSA-N 0.000 claims abstract description 7
- 239000012346 acetyl chloride Substances 0.000 claims abstract description 7
- 239000000126 substance Substances 0.000 claims abstract description 5
- 238000005882 aldol condensation reaction Methods 0.000 claims abstract description 3
- 238000005886 esterification reaction Methods 0.000 claims abstract description 3
- 230000003647 oxidation Effects 0.000 claims abstract description 3
- 238000007254 oxidation reaction Methods 0.000 claims abstract description 3
- 238000007363 ring formation reaction Methods 0.000 claims abstract description 3
- YMWUJEATGCHHMB-UHFFFAOYSA-N Dichloromethane Chemical compound ClCCl YMWUJEATGCHHMB-UHFFFAOYSA-N 0.000 claims description 30
- 238000006243 chemical reaction Methods 0.000 claims description 14
- HEMHJVSKTPXQMS-UHFFFAOYSA-M Sodium hydroxide Chemical compound [OH-].[Na+] HEMHJVSKTPXQMS-UHFFFAOYSA-M 0.000 claims description 12
- ZMANZCXQSJIPKH-UHFFFAOYSA-N Triethylamine Chemical compound CCN(CC)CC ZMANZCXQSJIPKH-UHFFFAOYSA-N 0.000 claims description 12
- 241000252212 Danio rerio Species 0.000 claims description 11
- 238000000034 method Methods 0.000 claims description 9
- 230000007613 environmental effect Effects 0.000 claims description 8
- 150000001875 compounds Chemical class 0.000 claims description 7
- HVQAJTFOCKOKIN-UHFFFAOYSA-N flavonol Natural products O1C2=CC=CC=C2C(=O)C(O)=C1C1=CC=CC=C1 HVQAJTFOCKOKIN-UHFFFAOYSA-N 0.000 claims description 7
- 235000011957 flavonols Nutrition 0.000 claims description 7
- 239000000203 mixture Substances 0.000 claims description 6
- 239000007787 solid Substances 0.000 claims description 5
- 238000003756 stirring Methods 0.000 claims description 5
- 238000001816 cooling Methods 0.000 claims description 3
- 238000010438 heat treatment Methods 0.000 claims description 3
- 238000010992 reflux Methods 0.000 claims description 3
- 238000000799 fluorescence microscopy Methods 0.000 claims description 2
- 238000001917 fluorescence detection Methods 0.000 claims 1
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 abstract description 19
- 230000035945 sensitivity Effects 0.000 abstract description 2
- 239000000243 solution Substances 0.000 description 13
- 239000000523 sample Substances 0.000 description 10
- 230000009286 beneficial effect Effects 0.000 description 4
- 230000000694 effects Effects 0.000 description 4
- 241000282414 Homo sapiens Species 0.000 description 3
- 239000012086 standard solution Substances 0.000 description 3
- FULZLIGZKMKICU-UHFFFAOYSA-N N-phenylthiourea Chemical compound NC(=S)NC1=CC=CC=C1 FULZLIGZKMKICU-UHFFFAOYSA-N 0.000 description 2
- PMZURENOXWZQFD-UHFFFAOYSA-L Sodium Sulfate Chemical compound [Na+].[Na+].[O-]S([O-])(=O)=O PMZURENOXWZQFD-UHFFFAOYSA-L 0.000 description 2
- 239000012491 analyte Substances 0.000 description 2
- 239000007864 aqueous solution Substances 0.000 description 2
- IOJUPLGTWVMSFF-UHFFFAOYSA-N benzothiazole Chemical compound C1=CC=C2SC=NC2=C1 IOJUPLGTWVMSFF-UHFFFAOYSA-N 0.000 description 2
- 238000004440 column chromatography Methods 0.000 description 2
- 238000011161 development Methods 0.000 description 2
- 238000000295 emission spectrum Methods 0.000 description 2
- 238000005516 engineering process Methods 0.000 description 2
- 150000002216 flavonol derivatives Chemical class 0.000 description 2
- 238000002189 fluorescence spectrum Methods 0.000 description 2
- 239000012044 organic layer Substances 0.000 description 2
- 239000012074 organic phase Substances 0.000 description 2
- 239000003960 organic solvent Substances 0.000 description 2
- 238000011160 research Methods 0.000 description 2
- 230000004044 response Effects 0.000 description 2
- 238000010898 silica gel chromatography Methods 0.000 description 2
- 239000002904 solvent Substances 0.000 description 2
- 239000011550 stock solution Substances 0.000 description 2
- 239000008399 tap water Substances 0.000 description 2
- 235000020679 tap water Nutrition 0.000 description 2
- GOLORTLGFDVFDW-UHFFFAOYSA-N 3-(1h-benzimidazol-2-yl)-7-(diethylamino)chromen-2-one Chemical compound C1=CC=C2NC(C3=CC4=CC=C(C=C4OC3=O)N(CC)CC)=NC2=C1 GOLORTLGFDVFDW-UHFFFAOYSA-N 0.000 description 1
- FAPWRFPIFSIZLT-UHFFFAOYSA-M Sodium chloride Chemical class [Na+].[Cl-] FAPWRFPIFSIZLT-UHFFFAOYSA-M 0.000 description 1
- GAMYVSCDDLXAQW-AOIWZFSPSA-N Thermopsosid Natural products O(C)c1c(O)ccc(C=2Oc3c(c(O)cc(O[C@H]4[C@H](O)[C@@H](O)[C@H](O)[C@H](CO)O4)c3)C(=O)C=2)c1 GAMYVSCDDLXAQW-AOIWZFSPSA-N 0.000 description 1
- 238000000862 absorption spectrum Methods 0.000 description 1
- 238000012271 agricultural production Methods 0.000 description 1
- XJHABGPPCLHLLV-UHFFFAOYSA-N benzo[de]isoquinoline-1,3-dione Chemical compound C1=CC(C(=O)NC2=O)=C3C2=CC=CC3=C1 XJHABGPPCLHLLV-UHFFFAOYSA-N 0.000 description 1
- 230000015572 biosynthetic process Effects 0.000 description 1
- 230000000711 cancerogenic effect Effects 0.000 description 1
- 231100000315 carcinogenic Toxicity 0.000 description 1
- 210000003169 central nervous system Anatomy 0.000 description 1
- 238000003889 chemical engineering Methods 0.000 description 1
- 239000003153 chemical reaction reagent Substances 0.000 description 1
- 239000012043 crude product Substances 0.000 description 1
- 230000007547 defect Effects 0.000 description 1
- 238000006073 displacement reaction Methods 0.000 description 1
- 239000012153 distilled water Substances 0.000 description 1
- 238000001035 drying Methods 0.000 description 1
- 238000003912 environmental pollution Methods 0.000 description 1
- 238000001704 evaporation Methods 0.000 description 1
- 230000005284 excitation Effects 0.000 description 1
- 230000005281 excited state Effects 0.000 description 1
- 238000001914 filtration Methods 0.000 description 1
- 239000012847 fine chemical Substances 0.000 description 1
- 229930003944 flavone Natural products 0.000 description 1
- 150000002212 flavone derivatives Chemical class 0.000 description 1
- 235000011949 flavones Nutrition 0.000 description 1
- GNBHRKFJIUUOQI-UHFFFAOYSA-N fluorescein Chemical compound O1C(=O)C2=CC=CC=C2C21C1=CC=C(O)C=C1OC1=CC(O)=CC=C21 GNBHRKFJIUUOQI-UHFFFAOYSA-N 0.000 description 1
- 238000002073 fluorescence micrograph Methods 0.000 description 1
- 238000012632 fluorescent imaging Methods 0.000 description 1
- 239000001963 growth medium Substances 0.000 description 1
- 238000009616 inductively coupled plasma Methods 0.000 description 1
- 238000011835 investigation Methods 0.000 description 1
- 210000003734 kidney Anatomy 0.000 description 1
- 238000012417 linear regression Methods 0.000 description 1
- 210000004185 liver Anatomy 0.000 description 1
- 210000001161 mammalian embryo Anatomy 0.000 description 1
- 238000004519 manufacturing process Methods 0.000 description 1
- 238000012544 monitoring process Methods 0.000 description 1
- 150000002894 organic compounds Chemical class 0.000 description 1
- 238000000746 purification Methods 0.000 description 1
- 238000004445 quantitative analysis Methods 0.000 description 1
- 210000002345 respiratory system Anatomy 0.000 description 1
- 230000004043 responsiveness Effects 0.000 description 1
- HPALAKNZSZLMCH-UHFFFAOYSA-M sodium;chloride;hydrate Chemical class O.[Na+].[Cl-] HPALAKNZSZLMCH-UHFFFAOYSA-M 0.000 description 1
- 238000003786 synthesis reaction Methods 0.000 description 1
- 238000012360 testing method Methods 0.000 description 1
- 231100000331 toxic Toxicity 0.000 description 1
- 230000002588 toxic effect Effects 0.000 description 1
- 238000012546 transfer Methods 0.000 description 1
- VHBFFQKBGNRLFZ-UHFFFAOYSA-N vitamin p Natural products O1C2=CC=CC=C2C(=O)C=C1C1=CC=CC=C1 VHBFFQKBGNRLFZ-UHFFFAOYSA-N 0.000 description 1
- 238000005406 washing Methods 0.000 description 1
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- C07—ORGANIC CHEMISTRY
- C07D—HETEROCYCLIC COMPOUNDS
- C07D311/00—Heterocyclic compounds containing six-membered rings having one oxygen atom as the only hetero atom, condensed with other rings
- C07D311/02—Heterocyclic compounds containing six-membered rings having one oxygen atom as the only hetero atom, condensed with other rings ortho- or peri-condensed with carbocyclic rings or ring systems
- C07D311/04—Benzo[b]pyrans, not hydrogenated in the carbocyclic ring
- C07D311/22—Benzo[b]pyrans, not hydrogenated in the carbocyclic ring with oxygen or sulfur atoms directly attached in position 4
- C07D311/26—Benzo[b]pyrans, not hydrogenated in the carbocyclic ring with oxygen or sulfur atoms directly attached in position 4 with aromatic rings attached in position 2 or 3
- C07D311/28—Benzo[b]pyrans, not hydrogenated in the carbocyclic ring with oxygen or sulfur atoms directly attached in position 4 with aromatic rings attached in position 2 or 3 with aromatic rings attached in position 2 only
- C07D311/30—Benzo[b]pyrans, not hydrogenated in the carbocyclic ring with oxygen or sulfur atoms directly attached in position 4 with aromatic rings attached in position 2 or 3 with aromatic rings attached in position 2 only not hydrogenated in the hetero ring, e.g. flavones
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- A61K49/00—Preparations for testing in vivo
- A61K49/001—Preparation for luminescence or biological staining
- A61K49/0013—Luminescence
- A61K49/0017—Fluorescence in vivo
- A61K49/0019—Fluorescence in vivo characterised by the fluorescent group, e.g. oligomeric, polymeric or dendritic molecules
- A61K49/0021—Fluorescence in vivo characterised by the fluorescent group, e.g. oligomeric, polymeric or dendritic molecules the fluorescent group being a small organic molecule
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- 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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Abstract
The invention discloses a flavonoid fluorescent probe for detecting hydrazine, a preparation method and application thereof, wherein the fluorescent probe is a flavonoid compound obtained by aldol condensation and oxidation ring closure reaction of p-tert-butyl benzaldehyde and o-hydroxyacetophenone and esterification reaction of p-tert-butyl benzaldehyde and o-hydroxyacetophenone with acetyl chloride, and the chemical structural formula of the flavonoid compound is shown as a formula (I). The fluorescent probe has unique fluorescence selectivity, extremely high sensitivity and extremely strong anti-interference capability on hydrazine in ethanol/water solution, and the detection limit can be as low as 0.116 mu M. The invention provides a simple and sensitive flavonoid fluorescent probe for detecting hydrazine, which has wide application prospect.
Description
Technical Field
The invention belongs to the technical fields of organic compound synthesis, fluorescent probes and fine chemical engineering, and particularly relates to a flavonoid fluorescent probe for detecting hydrazine, and a preparation method and application thereof.
Background
Hydrazine (N) 2 H 4 ) Is an important chemical substance, and has strong alkalinity, reducibility and combustibility, so that the chemical substance is widely applied to the fields of industrial manufacture, agricultural production, aerospace and the like. However, N 2 H 4 Are considered to be extremely toxic and potentially carcinogenic. Because of its wide application and excellent water solubility, N 2 H 4 Can easily enter an environmental system to cause serious environmental pollution. In addition, N 2 H 4 Is easy to enter and accumulate in human body through food chain, thereby being beneficial to human bodyThe respiratory tract, kidneys, liver and central nervous system of the body are severely damaged. Therefore, a rapid, effective, convenient and accurate N is developed and researched 2 H 4 The detection method has important significance for the healthy development of human beings.
Fluorescent probes have been considered to be an effective N 2 H 4 Compared with other detection technologies such as absorption spectrum, atomic emission spectrum and inductively coupled plasma emission spectrum, the fluorescent probe detection technology has the advantages of simplicity in operation, low cost, noninvasive detection, high spatial resolution and the like. Up to now, a considerable number of fluorescent probes based on different fluorophores have been developed to detect N 2 H 4 Such as coumarin, 1, 8-naphthalimide, fluorescein, benzothiazole, and the like. However, these fluorescent probes often have the disadvantages of high detection limit, narrow pH range, poor light stability, and the like. Furthermore, most of these reported fluorescent probes are only applied to N in biological systems 2 H 4 While detection N can be used in environmental systems 2 H 4 Still very limited fluorescent probes are available. Therefore, a novel method for detecting N with good selectivity and sensitivity in biological and environmental systems is designed 2 H 4 Still receiving extensive attention from researchers.
In recent years, some fluorescent dyes having an excited state intramolecular proton transfer (esit) feature have been widely used in the development of various fluorescent probes. The flavone and the derivative thereof have excellent photophysical characteristics and good biocompatibility, and can be used as a very promising fluorophore to construct various fluorescent probes. Inspired by the phenomenon, the invention prepares a novel Huang Tongji fluorescent probe. The fluorescent probe can selectively recognize N 2 H 4 After the reaction with hydrazine, the fluorescence emission intensity is obviously enhanced, and the detection limit is lower. The fluorescent probe can be applied to a wide pH range and quantitative detection. In addition, the probe can also be applied to monitoring N in living zebra fish and environment in real time 2 H 4 The content is of great significance for expanding the application range of the fluorescent probe.
Disclosure of Invention
Aiming at the defects of the prior art, the invention synthesizes the flavonoid fluorescent probe for detecting hydrazine, which has the advantages of good selectivity, strong anti-interference capability, wide pH value application range and low detection limit.
The invention also provides a preparation method of the flavonoid fluorescent probe for detecting hydrazine.
The invention also provides application of the flavonoid fluorescent probe for detecting hydrazine.
The technical scheme is as follows: in order to achieve the above object, the present invention has the technical scheme that: a flavonoid fluorescent probe for detecting hydrazine has a chemical structure shown in a formula (I):
the preparation method of the flavonoid fluorescent probe for detecting hydrazine is characterized by comprising the following experimental steps of;
(1) O-hydroxyacetophenone and p-tert-butylbenzaldehyde are subjected to aldol condensation and oxidation ring closure reaction to obtain a flavonol compound (II);
(2) Under alkaline conditions, acetyl chloride is used for carrying out esterification reaction with the compound (II) obtained in the step (1), and then the flavonoid fluorescent probe compound (I) can be prepared;
the specific synthetic route of the flavonoid fluorescent probe for detecting hydrazine is as follows:
the method comprises the following steps: dissolving p-tert-butylbenzaldehyde, 5.5mmol of o-hydroxyacetophenone and sodium hydroxide solid in ethanol, stirring at room temperature for reaction for 30min, heating and refluxing for 3H, cooling the reaction to room temperature, and adding H into the mixture 2 O 2 Stirring and reacting for 2 hours, after the reaction is finished, evaporating part of solvent in vacuum, extracting for multiple times by using dichloromethane,combining the organic layers, washing with saturated saline solution, drying with anhydrous sodium sulfate, filtering, and separating and purifying by silica gel column chromatography to obtain flavonol compounds (II); the flavonol compound (II) and triethylamine are dissolved in anhydrous dichloromethane and stirred at room temperature for 30min. Subsequently, an anhydrous methylene chloride solution containing acetyl chloride was slowly added to the above mixture, stirred at room temperature for 12 hours, and after completion of the reaction, the reaction solution was poured into water and extracted with methylene chloride. The combined organic phases were washed with water, the organic solvent was evaporated under reduced pressure and purified by column chromatography to give the flavonoid fluorescent probe (I).
The flavonoid fluorescent probe for detecting hydrazine has unique fluorescence response to hydrazine in ethanol/water solution.
Preparing ethanol solution of flavonoid fluorescent probe (I), and adding various analyte solutions such as Ag + 、Ca 2+ 、Cd 2+ 、Cu 2+ 、Mg 2+ 、Ni 2+ 、CO 3 2- 、HCO 3- 、HPO 4 2- 、HSO 3 - 、I - 、SO 3 2- 、SO 4 2- 、H 2 O 2 The fluorescence spectrum test is carried out on GSH and Hcy aqueous solutions to study the selective recognition effect on different analytes, and the result is shown in figure 1, and the intensity change of the fluorescence emission spectrum of the fluorescence probe is shown as the result, the flavonoid fluorescent probe (I) for detecting hydrazine has unique fluorescence responsiveness to hydrazine, and the fluorescence intensity is obviously enhanced after the fluorescence probe (I) is reacted with the hydrazine; in addition, a certain amount of the fluorescent probe solution was taken and hydrazine was gradually added to 1 equivalent, and the fluorescence intensity of the fluorescent probe (I) at 540nm was increased with the increase of the hydrazine concentration, and the result was shown in FIG. 2; drawing a process of dripping hydrazine into a solution containing a fluorescent probe (I), taking the fluorescence intensity as an ordinate, dripping the equivalent as an abscissa, and performing linear fitting to obtain a linear regression curve Y=1014.9940X+9496.0068, wherein the fitting coefficient R 2 0.9947, the result is shown in FIG. 3, which shows that the linear correlation degree is high, and therefore, the fluorescent probe can be used for quantitative analysis and detection of hydrazine molecules.
The flavonoid fluorescent probe for detecting hydrazine has excellent anti-interference performance on different analytes when detecting hydrazine, and the result is shown in figure 4, and the addition of other analytes has little effect on the fluorescence intensity of the fluorescent probe (I), so that the fluorescent probe (I) has unique fluorescence selectivity and stronger anti-interference capability on hydrazine in ethanol/water solution.
The flavonoid fluorescent probe for detecting hydrazine has the characteristic of wide application range of pH value, as shown in figure 5, the probe still has stable fluorescent emission for the identification of hydrazine within the pH value of 5.0-9.0, and the wider application range of pH value is beneficial to improving the practical detection performance of the fluorescent probe.
The invention also comprises a study of fluorescence imaging of the flavonoid fluorescent probe applied to the living zebra fish, as shown in figure 6, which shows that the flavonoid fluorescent probe can be successfully used for detecting hydrazine molecules in a biological system.
The invention also comprises the research of the application of the flavonoid fluorescent probe to the detection of the hydrazine content in distilled water, tap water, river water and lake water samples, and the research is shown in the table 1, which shows that the flavonoid fluorescent probe can be successfully used for the detection of the hydrazine in an environmental system.
The beneficial effects of the invention are as follows: (1) The fluorescent probe has the advantages of simple synthetic route, mild reaction condition and simple purification method; (2) The invention realizes the selective rapid detection of hydrazine, and has the advantages of large Stokes displacement, high fluorescence intensity, good selectivity, strong anti-interference capability and detection limit as low as 0.116 mu M. Therefore, the invention is a rapid and sensitive hydrazine detection reagent, and has wide application prospect in the fields of analytical chemistry and environmental detection.
Drawings
FIG. 1 shows a flavonoid fluorescent probe (I) (1X 10) -5 Fluorescence intensity profile after 1 equivalent of different analytes were added to the ethanol solution of M);
FIG. 2 shows flavonoid fluorescent probe (I) (1X 10) -6 M) in ethanol/water solution. In the figure, FL intensity is fluorescence emission intensity, wavelength is Wavelength, and excitation Wavelength is 365nm;
FIG. 3 is a graph of a linear fit of flavonoid fluorescent probe (I) with selected concentrations of hydrazine at different equivalents on the abscissa and fluorescence intensity on the ordinate; the abscissa is the concentration of the dropwise added hydrazine, and the unit is mu M;
FIG. 4 shows a flavonoid fluorescent probe (I) (1X 10) -6 M) bar graph of change in fluorescence intensity after 10 equivalents of other analytes are added to an aqueous solution in the presence of an equivalent amount of hydrazine;
FIG. 5 is a graph showing the effect of different pH values on fluorescence intensity of flavonoid fluorescent probes (I) on the identification of hydrazine;
FIG. 6 is a fluorescence image of flavonoid fluorescent probe (I) applied to living zebra fish containing hydrazine at different concentrations.
Table 1 shows the results of the detection of the concentration of hydrazine in a water sample of an actual environment by using the flavonoid fluorescent probe (I).
Detailed Description
The present invention will be described in further detail with reference to examples and drawings.
Example 1
Preparation of flavonols (II)
Dissolving 5mmol of p-tert-butylbenzaldehyde, 5.5mmol of o-hydroxyacetophenone and 12.5mmol of sodium hydroxide solid in 25mL of ethanol, stirring at room temperature for reaction for 30min, heating and refluxing for 3H, cooling the reaction to room temperature, and adding 1.5mL of H into the mixture 2 O 2 After the reaction was completed, a part of the solvent was distilled off under reduced pressure, extracted three times with 25mL of methylene chloride, and the organic layers were combined, washed with saturated brine, separated by a separating funnel, dried over anhydrous sodium sulfate, filtered, and then separated and purified by silica gel column chromatography to give the flavonol compound (II) as a pale yellow solid in yield: 65%. 1 H NMR(400MHz，CDCl 3 )δ：8.24(m，1H)，8.18(d，J＝6.4Hz，2H)，7.68(m，1H)，7.57(d，J＝6.6Hz，1H)，7.55(d，J＝6.4Hz，2H)，7.40(m，1H)，6.96(s，1H)，1.38(s，9H)；
Example two
Preparation of flavonoid fluorescent probe (I)
0.5mmol of flavonols compoundII) and 1.0mmol of triethylamine are dissolved in 15mL of anhydrous dichloromethane and stirred at room temperature for 30min; subsequently, 5mL of anhydrous dichloromethane containing 1.0mmol of acetyl chloride was slowly added to the above mixture, stirred at room temperature for 12 hours, after the reaction was completed, the reaction solution was poured into water, and extracted with dichloromethane; the combined organic phases were washed with water, the organic solvent was distilled off under reduced pressure, and the crude product was purified by column chromatography to give the flavonoid fluorescent probe (I) as a pale yellow solid, yield: 46%. 1 H NMR(600MHz，CDCl 3 )δ：8.25(d，J＝6Hz，1H)，7.83(d，J＝12Hz，2H)，7.69(t，J＝12Hz，1H)，7.54(d，J＝12Hz，3H)，7.41(t，J＝12Hz，1H)，2.37(s，3H)，1.37(s，9H)； 13 C NMR(150MHz，CDCl 3 )δ：172.27，168.20，156.46，155.70，154.99，133.96，133.61，128.17，127.22，126.16，125.81，125.20，123.70，118.15，35.11，31.21，20.76；HRMS(m/z)：calcd for C 21 H 21 O 4 [M+H] + ：337.1440；found：337.1472。
Example III
Selective investigation of flavonoid fluorescent probes for different analytes
Preparation of fluorescent Probe (I) (1×10) -5 M) with 1 equivalent of standard solutions of different analytes, such as Ag, respectively + 、Ca 2+ 、Cd 2+ 、Cu 2+ 、Mg 2+ 、Ni 2+ 、CO 3 2- 、HCO 3- 、HPO 4 2- 、HSO 3 - 、I - 、SO 3 2- 、SO 4 2- 、H 2 O 2 As can be seen from FIG. 1, the solution of fluorescent probe (I) emits bright green fluorescence with a significant fluorescence enhancement at 540nm when hydrazine is added; and after other analytes are added, no fluorescence change is caused, so that the fluorescent probe (I) has a good specific recognition effect on hydrazine, and can be used as a specific fluorescent probe for detecting the hydrazine.
Example IV
Variation of fluorescence intensity of flavonoid fluorescent probe along with increase of hydrazine addition
1mL of the stock solution prepared in the third embodiment is taken out and added into a 10mL volumetric flask, different equivalents of hydrazine molecule standard solution are added, the volume is fixed to 10mL, the fluorescence intensity change of the emission wavelength at 540nm is measured, and the result is shown in figure 2, the fluorescence intensity of the flavonoid fluorescent probe solution gradually increases along with the increase of the adding amount of hydrazine, and the flavonoid fluorescent probe has high sensitive fluorescence response to the change of the hydrazine concentration. The result of linear fitting with the hydrazine concentration on the abscissa and the fluorescence intensity on the ordinate at the different equivalents selected is shown in fig. 3, and the fluorescence intensity and the hydrazine concentration change have a good linear relationship.
Example five
Interference resistance of flavonoid fluorescent probe to different analytes in detection of hydrazine molecules
1mL of the stock solution of the fluorescent probe in example three was taken out and added to a 10mL volumetric flask, an equivalent amount of hydrazine standard solution was added, and then 10 equivalents of the other analytes were added, respectively, and the volume was fixed to 10mL. The competing analyte comprises Ag + 、Ca 2+ 、Cd 2+ 、Cu 2+ 、Mg 2+ 、Ni 2+ 、CO 3 2- 、HCO 3- 、HPO 4 2- 、HSO 3 - 、I - 、SO 3 2- 、SO 4 2- 、H 2 O 2 The fluorescence intensity changes at 540nm of fluorescence emission are observed, and the results are shown in fig. 4, and it can be found that when other analytes are added, the fluorescence intensity of the flavonoid fluorescent probe is hardly influenced, so that the flavonoid fluorescent probe has good anti-interference performance on other analytes when hydrazine molecules are detected.
Example six
Influence of different pH values on identification of hydrazine by flavonoid fluorescent probes:
in order to study the optimal pH value range for detecting hydrazine, the change of the fluorescence intensity of the flavonoid fluorescent probe (I) under different pH values in the presence and absence of hydrazine is studied, and as can be seen from FIG. 5, the fluorescence intensity of the fluorescent probe (I) without hydrazine is still weak in the pH value range of 3-9; in contrast, in the presence of hydrazine, the fluorescence intensity of the fluorescent probe (I) is significantly enhanced and kept stable at pH values in the range of 5-9. The flavonoid fluorescent probe (I) has a wide applicable pH value range, and is beneficial to improving the actual detection performance of the fluorescent probe.
Example seven
The flavonoid fluorescent probe is applied to living zebra fish hydrazine identification fluorescent imaging.
Zebra fish were grown in embryo culture medium containing 1-phenyl-2-thiourea (PTU) and incubated at 28℃for 72h. Treating zebra fish with 10 μm flavonoid fluorescent probe (I) for 0.5h; after three washes with PBS buffer, zebra fish were further incubated with different concentrations of hydrazine (50. Mu.M and 100. Mu.M). As shown in FIG. 6, zebra fish stained with 10. Mu.M of flavonoid fluorescent probe (I) did not show a significant fluorescent signal; after 50 mu M of hydrazine is added, the fluorescence emission of the living zebra fish pretreated by the flavonoid fluorescent probe (I) is obviously increased, and the fluorescence emission is continuously enhanced along with the increase of the concentration of the hydrazine; these results show that the probe has good biocompatibility and is suitable for detecting the hydrazine content in the living zebra fish in real time.
Example eight
The flavonoid fluorescent probe detects the concentration of hydrazine in an environmental water sample.
And (3) detecting the concentration of hydrazine in the water samples of tap water, river water and lake water environments by adopting a flavonoid fluorescent probe (I). As shown in table 1, the concentration of hydrazine detected by probe (I) is very close to the actual concentration added in the water sample (table 1); the results show that the flavonoid fluorescent probe (I) can be used for accurately measuring the concentration of hydrazine in an environmental water sample.
TABLE 1
Claims (5)
1. The use of a flavonoid fluorescent probe is characterized by being used for detecting hydrazine;
the method is used for detecting the content of hydrazine molecules in the environment, wherein the detection is fluorescence detection, the minimum detection limit is 0.116 mu M, and the method can be applied to the detection of the hydrazine molecules in biological and environmental systems;
the linear relationship between fluorescence intensity and hydrazine concentration is: y=1014.9940x+9496.0068, r 2 0.9947;
the pH value range is 5.0-9.0;
the method is used for identifying fluorescence imaging of the living zebra fish hydrazine;
the chemical structural formula of the flavonoid fluorescent probe is shown as a formula (I),
2. the use of a flavonoid fluorescent probe according to claim 1, wherein the preparation method of the flavonoid fluorescent probe comprises the following steps:
(1) O-hydroxyacetophenone and p-tert-butylbenzaldehyde are subjected to aldol condensation and oxidation cyclization reaction to obtain a flavonol compound (II), wherein the structural formula is as follows:
(2) Under alkaline conditions, acetyl chloride is used for carrying out esterification reaction with the compound (II) obtained in the step (1), and the flavonoid fluorescent probe compound (I) for detecting hydrazine can be obtained, wherein the structural formula is as follows:
the step (1) is completed by adopting the following method: dissolving p-tert-butylbenzaldehyde, o-hydroxyacetophenone and sodium hydroxide solid in ethanol, stirring at room temperature for 30min, heating and refluxing for 3H, cooling to room temperature, and adding H into the mixture 2 O 2 The reaction was stirred for 2 hours to give compound (II).
3. Use of a flavonoid fluorescent probe according to claim 2, characterized in that p-tert-butylbenzaldehyde: ortho-hydroxyacetophenone: the molar ratio of sodium hydroxide is 1:1.1:2.5.
4. the use of a flavonoid fluorescent probe according to claim 2, wherein the step (2) is accomplished by the following method: dissolving the compound (II) and triethylamine in anhydrous dichloromethane, and stirring at room temperature for 30min; subsequently, an anhydrous methylene chloride solution containing acetyl chloride was slowly added to the above mixture, and the reaction was stirred at room temperature for 12 hours to give compound (I).
5. The use of a flavonoid fluorescent probe according to claim 4, wherein the molar ratio of triethylamine to acetyl chloride is 1:1.
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| CN103865962A (en) * | 2014-01-24 | 2014-06-18 | 潍坊医学院 | Enzymatic preparation method and application of quercetin-3-O-fatty acid ester |
| CN111484470A (en) * | 2020-03-28 | 2020-08-04 | 齐鲁工业大学 | Fluorescent probe for detecting hydrazine, preparation method and application thereof |
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| Publication number | Priority date | Publication date | Assignee | Title |
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| CN103865962A (en) * | 2014-01-24 | 2014-06-18 | 潍坊医学院 | Enzymatic preparation method and application of quercetin-3-O-fatty acid ester |
| CN111484470A (en) * | 2020-03-28 | 2020-08-04 | 齐鲁工业大学 | Fluorescent probe for detecting hydrazine, preparation method and application thereof |
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| "819835-67-1";STN Registry数据库;STN Registry数据库;第1页 * |
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| STN Registry数据库."819835-67-1".STN Registry数据库.2005,第1页. * |
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