CN107417638A - A kind of glutathione and cysteine fluorescence probe based on 7 nitrobenzofurazans and preparation method thereof - Google Patents
A kind of glutathione and cysteine fluorescence probe based on 7 nitrobenzofurazans and preparation method thereof Download PDFInfo
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- CN107417638A CN107417638A CN201710307526.8A CN201710307526A CN107417638A CN 107417638 A CN107417638 A CN 107417638A CN 201710307526 A CN201710307526 A CN 201710307526A CN 107417638 A CN107417638 A CN 107417638A
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- C07—ORGANIC CHEMISTRY
- C07D—HETEROCYCLIC COMPOUNDS
- C07D271/00—Heterocyclic compounds containing five-membered rings having two nitrogen atoms and one oxygen atom as the only ring hetero atoms
- C07D271/12—Heterocyclic compounds containing five-membered rings having two nitrogen atoms and one oxygen atom as the only ring hetero atoms condensed with carbocyclic rings or ring systems
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- 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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- C—CHEMISTRY; METALLURGY
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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
- C09K2211/1044—Heterocyclic compounds characterised by ligands containing two nitrogen atoms as heteroatoms
- C09K2211/1048—Heterocyclic compounds characterised by ligands containing two nitrogen atoms as heteroatoms with oxygen
Abstract
The invention discloses a kind of glutathione based on 7 nitrobenzofurazans and cysteine fluorescence probe and preparation method thereof, the nitro benzofuraxan of 4 amino of intermediate 7 and triethylamine are mixed in dichloromethane solution, the dichloromethane solution containing acryloyl chloride is added dropwise under condition of ice bath, reacted afterwards, reaction product obtains glutathione and cysteine fluorescence probe by subsequent treatment.The fluorescence probe is used for the fluorescence sense analysis of environment or glutathione and cysteine in biological sample.Evident characteristics of the probe MP1 with each amino acid in DMSO/ PBS (10 mM, pH=7.4,1/9, v/v) solution system have studied by sepectrophotofluorometer.As a result show：Probe MP1 is good to the selectivity of glutathione and cysteine, strong antijamming capability, glutathione and cysteine can be quickly distinguished from several amino acids with sensitive, has good application prospect.
Description
Technical field
The present invention relates to a kind of for glutathione and the new detection agent of cysteine fluorescence identifying, and in particular to a kind of
Preparation method and the application of glutathione and cysteine fluorescent probe molecule based on 7- nitrobenzofurazans.
Background technology
In organism, biological thiol such as glutathione (GSH), cysteine (Cys) etc. is in physiology course such as cell generation
Played an important role during journey of apologizing for having done sth. wrong and maintenance redox reaction balance etc..The change of such intracellular mercaptans content
A series of disease, such as AIDS, oedema, coronary heart disease, parkinsonism, cancer and psoriasis can be caused.As containing into the cell
Highest nonprotein mercaptan is measured, glutathione (GSH) plays extremely important in redox homeostasis process is kept
Effect, its content it is too high or too low, have direct relation with many diseases such as cancer.Cysteine in organism
(Cys) shortage can cause a series of illness such as illnesss, such as skin injury, liver oedema.Therefore, such sulphur of fast real-time monitoring
The content of alcohol in vivo has great significance in clinical medicine and biochemistry.
Fluorescence probe have the advantages that the good, high sensitivity of selectivity, it is easy to operate it is quick, detectable substance is damaged it is few wide
Active small molecular etc. in metal cation, anion, organism in the general system applied to detection environmental and biological materials.Fluorescence
The development and utilization research of probe molecule is the crossing domain of the science such as chemistry and biology, medical science, agricultural.Fluorescent probe technique is
The analysis method studied on a molecular scale research object.Fluorescence probe has been widely used in environment and life at present
The detection of the material such as heavy metal, bioactive small molecule in object.
Because it has simple to operate, single-minded efficient selectivity has been widely used in detecting environment and life fluorescence probe method
Active small molecular etc. in metal cation, anion, organism in objects system.Compared to common other in organism
Amino acid, biological thiol such as cysteine (Cys), all contain sulfydryl in glutathione (GSH) molecular structure, therefore to biological sulphur
The detection of alcohol can be realized by the identification to sulfydryl.Sulfydryl possesses stronger nucleophilicity, many mercaptan reported at present
Fluorescence probe mainly realizes the efficient identification to biological thiol using its stronger nucleophilicity.
The content of the invention
The present invention proposes a kind of glutathione based on 7- nitrobenzofurazans and cysteine fluorescence probe and its preparation
Method, the fluorescence probe of synthesis sensitive can quickly distinguish glutathione and cysteine from several amino acids.
Realize the technical scheme is that：A kind of glutathione and cysteine fluorescence based on 7- nitrobenzofurazans are visited
Pin（It is denoted as MP1）, the structural formula of the fluorescent probe molecule is as follows：
。
The preparation method of described glutathione and cysteine fluorescence probe based on 7- nitrobenzofurazans, step is such as
Under：
Under the conditions of lucifuge, by intermediate 4- amino -7- nitro benzofuraxans（NBD-NH2）It is molten that dichloromethane is mixed in triethylamine
In liquid, the dichloromethane solution containing acryloyl chloride is added dropwise under condition of ice bath, after reaction, reaction solution is washed with saturated nacl aqueous solution
Wash, organic phase anhydrous sodium sulfate drying, filter, removal of solvent under reduced pressure, be eluant, eluent with ethyl acetate and n-hexane, through silica gel
Column chromatography for separation obtains light yellow solid, obtains glutathione and cysteine fluorescence probe.
The step（1）In, lucifuge condition is using masking foil encapsulation reaction container.
Triethylamine is the mol ratio of acid binding agent, intermediate 4- amino -7- nitros benzofuraxan and triethylamine in the step
For 1：（2-10）, dichloromethane used need to be 0 DEG C through Non-aqueous processing, the ice bath reaction temperature, and the reaction time is 3-12 hours.
The mol ratio of the intermediate 4- amino -7- nitros benzofuraxan and acryloyl chloride is 1：（2-5）.
The eluent of the silica gel post separation is ethyl acetate and n-hexane, the volume ratio of the ethyl acetate and n-hexane
For 1：（5-20）.
The preparation method of the intermediate 4- amino -7- nitro benzofuraxans is：Under the conditions of lucifuge, by ammonia spirit dropwise
It is added drop-wise in the methanol solution containing the chloro- 7- nitrobenzofurazans of 4-, the concentration of the methanol solution of the chloro- 7- nitrobenzofurazans of 4-
For 40-70%, stirring reaction 20-30 hours, vacuum distillation at room temperature obtains solid, and the solid is dissolved with ethyl acetate, with second
Acetoacetic ester and petroleum ether are intermediate as eluant, eluent, the isolated Tan solid of silica gel column chromatography, the Tan solid
4- amino -7- nitro benzofuraxans.
The ratio between amount of material of the chloro- 7- nitrobenzofurazans of the 4- and ammonia spirit is 1:（15-30）.
Glutathione and cysteine fluorescence probe based on 7- nitrobenzofurazans are in detection glutathione and cysteine
In application.
The method of the dichloromethane Non-aqueous processing is：Distilled after calcium hydride backflow 2-4 hours are added into dichloromethane,
Produce anhydrous dichloromethane.
Used chemical reagent, solvent, metal salt etc. during fluorescence probe MP1 are prepared in the present invention and is purchased from state
Chemical reagent Co., Ltd of medicine group.Bruke companies DTX-400 is used in fluorescence probe MP1 confirmation and performance test process
Type nuclear magnetic resonance spectrometer, solvent are deuterochloroform, record proton nmr spectra as internal standard using TMS and carbon is composed.It is public using Thermo
The Q-Exactive HR-MS mass spectrographs record high resolution mass spectrum data of department.Using HIT's F-7000 XRFs
Record fluorescence spectrum.
The specific synthetic route of the present invention is as follows：
The beneficial effects of the invention are as follows：Fluorescence probe is obtained by simple single step reaction, synthetic route is short, and method is easy, production
Rate is high.Fluorescence probe is in DMSO/PBS (10 mM, pH=7.4,1/9, v/v) solution system to glutathione and half Guang
The selectivity of propylhomoserin is good, strong antijamming capability, glutathione and half Guang can be quickly distinguished from several amino acids with sensitive
Propylhomoserin.Fluorogen 4- amino -7- nitros benzofuraxan in fluorescence probe structure has longer launch wavelength and well thin
Born of the same parents' permeability, be advantageous to such probe and identify glutathione and cysteine in vivo, thus with preferably application before
Scape.
Brief description of the drawings
Fig. 1 be the present invention fluorescence probe MP1 fluorescence selectivity figures, the nm of excitation wavelength 380.
Fig. 2 is that the fluorescence probe MP1 of the present invention identifies GSH competitive lab diagram, the nm of excitation wavelength 380, launch wavelength
500 nm。
Fig. 3 is that the fluorescence probe MP1 of the present invention identifies Cys competitive lab diagram, the nm of excitation wavelength 380, launch wavelength
500 nm。
Fig. 4 be the present invention fluorescence probe MP1 fluorescence kinetics lab diagrams, the nm of excitation wavelength 380, launch wavelength 500
nm。
Fig. 5 is that the fluorescence probe MP1 of the present invention identifies GSH fluorescence titration figure, and illustration is minimum detection limit figure, excitation wave
Long 380 nm, the nm of launch wavelength 500.
Fig. 6 is that the fluorescence probe MP1 of the present invention identifies Cys fluorescence titration figure, and illustration is minimum detection limit figure, excitation wave
Long 380 nm, the nm of launch wavelength 500.
Fig. 7 is that the fluorescence probe MP1 of the present invention identifies GSH and Cys acid-base value lab diagram, the nm of excitation wavelength 380, hair
Ejected wave grows 500 nm.
Embodiment
Embodiment 1
The synthesis of fluorescence probe：
（1）Under the conditions of lucifuge, ammonia spirit is added drop-wise to dropwise in the methanol solution containing the chloro- 7- nitrobenzofurazans of 4-, it is described
The ratio between amount of material of the chloro- 7- nitrobenzofurazans of 4- and ammonia spirit is 1:15, the methanol of the chloro- 7- nitrobenzofurazans of 4- is molten
The concentration of liquid is 40%, at room temperature stirring reaction 20 hours, and vacuum distillation obtains solid, and the solid is dissolved with ethyl acetate, with
Ethyl acetate and petroleum ether are centre as eluant, eluent, the isolated Tan solid of silica gel column chromatography, the Tan solid
Body 4- amino -7- nitro benzofuraxans；
（2）Under the conditions of lucifuge, by intermediate NBD-NH2(180 mg, 1 mmol) is mixed with triethylamine (270 μ L, 2 mmol)
Together in the dichloromethane (10 that acryloyl chloride (162 μ L, 2 mmol) in 15 mL dichloromethane solutions, is added dropwise under condition of ice bath
ML) solution, after reacting completely under condition of ice bath (3 hours), reaction solution is washed with saturated nacl aqueous solution (15 mL*3), organic
Phase anhydrous sodium sulfate drying, filtering, removal of solvent under reduced pressure, is eluant, eluent with ethyl acetate and n-hexane（Volume ratio is 1:6）,
Light yellow solid through isolated 105.3 mg of silica gel column chromatography, as fluorescent probe molecule, yield 45%.
Nuclear magnetic resonance hydrogen spectruming determining：1H NMR (400 MHz, DMSO, ppm) δ: 5.97 (dd, 1 H, J =
1.6 Hz), 6.46 (dd, 1 H, J = 1.6 Hz), 6.93 (t, 1 H, J = 9.0 Hz), 8.51 (d, 1 H,J = 8.0 Hz), 8.79 (d, 1 H, J = 8.4 Hz), 11.62 (s, 1 H)。
Carbon-13 nmr spectra determines：13C NMR (100 MHz, DMSO, ppm) δ: 114.7, 130.6, 130.8,
131.0, 135.1, 136.2, 143.9, 146.1, 165.5。
High resolution mass spectrum determines：HR-MS m/z:Theoretical value：C9H7N4O4([M+H+]+) 235.0647, detection value：
235.0649。
Embodiment 2
The preparation method of fluorescence probe is as follows in the present embodiment：
（1）Under the conditions of lucifuge, ammonia spirit is added drop-wise to dropwise in the methanol solution containing the chloro- 7- nitrobenzofurazans of 4-, it is described
The ratio between amount of material of the chloro- 7- nitrobenzofurazans of 4- and ammonia spirit is 1:20, the methanol of the chloro- 7- nitrobenzofurazans of 4- is molten
The concentration of liquid is 60%, at room temperature stirring reaction 25 hours, and vacuum distillation obtains solid, and the solid is dissolved with ethyl acetate, with
Ethyl acetate and petroleum ether are centre as eluant, eluent, the isolated Tan solid of silica gel column chromatography, the Tan solid
Body 4- amino -7- nitro benzofuraxans；
（2）Under the conditions of lucifuge, by intermediate NBD-NH2(180 mg, 1 mmol) is mixed with triethylamine (810 μ L, 6 mmol)
Together in the dichloromethane (10 that acryloyl chloride (324 μ L, 4 mmol) in 15 mL dichloromethane solutions, is added dropwise under condition of ice bath
ML) solution, after reacting completely under condition of ice bath (6 hours), reaction solution is washed with saturated nacl aqueous solution (15 mL*3), organic
Phase anhydrous sodium sulfate drying, filtering, removal of solvent under reduced pressure, is eluant, eluent with ethyl acetate and n-hexane（Volume ratio is 1:8）,
Light yellow solid through isolated 128 mg of silica gel column chromatography, as fluorescent probe molecule, yield 55%.
Nuclear magnetic resonance hydrogen spectruming determining：1H NMR (400 MHz, DMSO, ppm) δ: 5.97 (dd, 1 H, J =
1.6 Hz), 6.46 (dd, 1 H, J = 1.6 Hz), 6.93 (t, 1 H, J = 9.0 Hz), 8.51 (d, 1 H,J = 8.0 Hz), 8.79 (d, 1 H, J = 8.4 Hz), 11.62 (s, 1 H);
Carbon-13 nmr spectra determines：13C NMR (100 MHz, DMSO, ppm) δ: 114.7, 130.6, 130.8,
131.0, 135.1, 136.2, 143.9, 146.1, 165.5；
High resolution mass spectrum determines：HR-MS m/z:Theoretical value：C9H7N4O4 ([M+H+]+) 235.0647, detection value：
235.0649。
Embodiment 3
The preparation method of fluorescence probe is as follows in the present embodiment：
（1）Under the conditions of lucifuge, ammonia spirit is added drop-wise to dropwise in the methanol solution containing the chloro- 7- nitrobenzofurazans of 4-, it is described
The ratio between amount of material of the chloro- 7- nitrobenzofurazans of 4- and ammonia spirit is 1:30, the methanol of the chloro- 7- nitrobenzofurazans of 4- is molten
The concentration of liquid is 70%, at room temperature stirring reaction 30 hours, and vacuum distillation obtains solid, and the solid is dissolved with ethyl acetate, with
Ethyl acetate and petroleum ether are centre as eluant, eluent, the isolated Tan solid of silica gel column chromatography, the Tan solid
Body 4- amino -7- nitro benzofuraxans；
（2）Under the conditions of lucifuge, by intermediate NBD-NH2(180 mg, 1 mmol) and triethylamine (1350 μ L, 6 mmol)
It is mixed in 15 mL dichloromethane solutions, acryloyl chloride (405 μ L, 5 mmol) dichloromethane is added dropwise under condition of ice bath
(10 mL) solution, after reacting completely under condition of ice bath (12 hours), reaction solution is washed with saturated nacl aqueous solution (15 mL*3),
Organic phase anhydrous sodium sulfate drying, filtering, removal of solvent under reduced pressure, is eluant, eluent with ethyl acetate and n-hexane（Volume ratio is
1:8）, the light yellow solid through isolated 128 mg of silica gel column chromatography, as fluorescent probe molecule, yield 55%.
Probe MP1 prepared by embodiment 1 application examples
（1）The preparation of solution：
Calcium chloride, potassium chloride, sodium chloride, natrium nitrosum, hydrogen peroxide and each amino acid, which are purchased from Chinese medicines group chemical reagent, to be had
Limit company.Precise respective metal salt, amino acid, are dissolved in the solution for standby that 10 mM are prepared in high purity water.The dioxygen of purchase
Water is diluted with deionized water, and according to Beer law, the concentration of hydrogen peroxide is demarcated by the nm of ultraviolet spectra 240 absorbance, root
According to formula c=A/Kb（b = 1 cm, K =43.6 M-1 cm-1）Calculate.
1 mM probe solution is prepared：Precise correspondent probe, fluorescence probe is dissolved in DMSO solution and prepares 1
MM solution for standby.
Test system compound method：In bottle is tested, 2.5 mL 10 mM PBS buffer (pH 7.4) are added,
The test sample solution of respective volume is added, after being well mixed, 30 μ L 1 mM probe solutions is added, corresponding body is finally added dropwise
Long-pending DMSO and 10 mM PBS buffer (pH 7.4) solution makes the 10 mM PBS buffer (pH that total system is 3 mL
7.4) system containing 10 % DMSO.
（2）Selectivity experiment：
Good selectivity is the whether excellent major criterion of one fluorescent probe molecule of inspection with stronger anti-interference.First
Investigated probe to common active sulfur, amino acid, metal ion selectivity.All of above test substances are added not to surveying
The fluorescent emission intensity of examination system produces obvious change.As a comparison, corresponding glutathione is added under the same terms（GSH）
And cysteine（Cys）When, system fluorescent emission intensity has obvious reduction（Fig. 1）.It is somebody's turn to do test result indicates that probe is to paddy Guang
Sweet peptide（GSH）And cysteine（Cys）Possesses preferable selectivity.In order to test probe to glutathione（GSH）With half Guang ammonia
Acid（Cys）The antijamming capability of identification, test the anti-interference of the probe respectively in fluorescence emission spectrum.Such as Fig. 2 and Fig. 3
It is shown, it is separately added into the various of test in DMSO/PBS (10 mM, pH=7.4,1/9, v/v) solution at (10 μM)
Common amino acid (100 μM) tests its fluorescent emission intensity（500 nm）, then divide again into the solution containing various amino acid
Glutathione that Jia Ru be 100 μM（GSH）And cysteine（Cys）Solution, from Fig. 2 and Fig. 3, deposited in common amino acid
When add glutathione（GSH）And cysteine（Cys）With being individually added into glutathione（GSH）And cysteine（Cys）When
Resulting fluorescent emission intensity (500 nm) is essentially identical, and the result shows probe to glutathione（GSH）And cysteine
（Cys）Detection there is stronger antijamming capability.
（3）Identify dynamic experiment：
Quick response, in real time monitoring are one of important features of fluorescence probe detection.When having investigated reaction by XRF
Between probe in detecting glutathione（GSH）And cysteine（Cys）Influence.As shown in figure 4, DMSO/PBS (10 mM, pH=
7.4,1/9, v/v) in solution system, probe is to glutathione（GSH）And cysteine（Cys）Recognition time is 45 minutes,
And reach balance after 45 mins.As a result show probe to glutathione（GSH）And cysteine（Cys）Recognition time is 45 points
Clock, its reaction product possess stronger stability.
（4）Fluorescence titration and test limit test：
In order to further study probe to glutathione（GSH）And cysteine（Cys）The sensitivity of identification, passes through fluorescence titration
Its minimum detection limit of experiment test.As shown in figure 5, in DMSO/ PBS (10 mM, pH=7.4,1/9, v/v) solution body
In system, control concentration and probe concentration be 10 μM, be continuously increased GSH concentration (0,1,2,3,5,7,10,15,20,
30,40,50,60,70,80,100 μM) its fluorescence emission spectrum is tested, as the increase of GSH concentration, system are glimmering
Light emissive porwer constantly reduces, meanwhile, the reduction of system fluorescent emission intensity is linear between GSH concentration is 1-30 μM
（R 2 =0.991, Fig. 5）, according to IUPAC rules, measure detection of the probe to GSH and be limited to 106 nM.Same method, investigate
Probe and Cys fluorescence titration experiment, measure the detection to Cys and are limited to 103 nM (Fig. 6).
（5）PH values identify glutathione to probe（GSH）And cysteine（Cys）Influence：
Glutathione can be identified in order to test probe under different physiological conditions（GSH）And cysteine（Cys）, investigate
It identifies glutathione in different pH values to it（GSH）And cysteine（Cys）Influence.As shown in Figure 7：In DMSO/PBS
In (10 mM, pH=7.4,1/9, v/v) solution system, change the pH value of buffer system（4-9）, independent probe (10 μ L)
Fluorescence emission spectrum change it is little, but in glutathione（GSH）And cysteine（Cys）（Concentration is respectively 100 μ L）Deposit
Under the conditions, system fluorescent emission intensity has between 7-9 in pH and is significantly quenched, and the result shows that probe can be applied
Glutathione is identified under physiological condition（GSH）And cysteine（Cys）.
Claims (9)
1. a kind of glutathione and cysteine fluorescence probe based on 7- nitrobenzofurazans, it is characterised in that the fluorescence is visited
The structural formula of pin molecule is as follows：
。
2. the preparation side of the glutathione based on 7- nitrobenzofurazans and cysteine fluorescence probe as described in requiring right 1
Method, it is characterised in that step is as follows：
Under the conditions of lucifuge, intermediate 4- amino -7- nitros benzofuraxan and triethylamine are mixed in dichloromethane solution, ice bath
Under the conditions of be added dropwise acryloyl chloride dichloromethane solution, after reaction, reaction solution is washed with saturated nacl aqueous solution, organic phase nothing
Aqueous sodium persulfate is dried, and filtering, removal of solvent under reduced pressure, is eluant, eluent with ethyl acetate and n-hexane, is separated through silica gel column chromatography
To light yellow solid, glutathione and cysteine fluorescence probe are obtained.
3. the preparation side of the glutathione and cysteine fluorescence probe as claimed in claim 2 based on 7- nitrobenzofurazans
Method, it is characterised in that：The step（1）In, lucifuge condition is using masking foil encapsulation reaction container.
4. the preparation side of the glutathione and cysteine fluorescence probe as claimed in claim 2 based on 7- nitrobenzofurazans
Method, it is characterised in that：The mol ratio of the intermediate 4- amino -7- nitros benzofuraxan and triethylamine is 1：（2-10）, it is used
Dichloromethane need to be 0 DEG C through Non-aqueous processing, the ice bath reaction temperature, and the reaction time is 3-12 hours.
5. the preparation side of the glutathione and cysteine fluorescence probe as claimed in claim 2 based on 7- nitrobenzofurazans
Method, it is characterised in that：The mol ratio of the intermediate 4- amino -7- nitros benzofuraxan and acryloyl chloride is 1：（2-5）.
6. the preparation side of the glutathione and cysteine fluorescence probe as claimed in claim 2 based on 7- nitrobenzofurazans
Method, it is characterised in that：The eluent of the silica gel post separation is ethyl acetate and n-hexane, the ethyl acetate and n-hexane
Volume ratio is 1：（5-20）.
7. the glutathione and cysteine fluorescence probe based on 7- nitrobenzofurazans as described in claim any one of 1-6
Preparation method, it is characterised in that the preparation method of the intermediate 4- amino -7- nitro benzofuraxans is：, will under the conditions of lucifuge
Ammonia spirit is added drop-wise in the methanol solution containing the chloro- 7- nitrobenzofurazans of 4- dropwise, the first of the chloro- 7- nitrobenzofurazans of 4-
The concentration of alcoholic solution is 40-70%, at room temperature stirring reaction 20-30 hours, and vacuum distillation obtains solid, the solid acetic acid
Ethyl ester is dissolved, and eluant, eluent, the isolated Tan solid of silica gel column chromatography, the yellowish-brown are used as using ethyl acetate and petroleum ether
Solid is intermediate 4- amino -7- nitro benzofuraxans.
8. the preparation side of the glutathione and cysteine fluorescence probe as claimed in claim 7 based on 7- nitrobenzofurazans
Method, it is characterised in that：The ratio between amount of material of the chloro- 7- nitrobenzofurazans of the 4- and ammonia spirit is 1:（15-30）.
9. glutathione and cysteine fluorescence probe based on 7- nitrobenzofurazans are in detection glutathione and cysteine
Application.
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| CN108484523A (en) * | 2018-03-26 | 2018-09-04 | 江苏省农用激素工程技术研究中心有限公司 | Benzo oxa- oxadiazole derivatives and its preparation method and application |
| CN113979973A (en) * | 2021-10-29 | 2022-01-28 | 南京碳硅人工智能生物医药技术研究院有限公司 | Design, synthesis and activity research of in-situ cysteine detection fluorescent probe |
| CN115160381A (en) * | 2022-08-18 | 2022-10-11 | 安徽工业大学 | Glutathione-responsive fluorescent probe, preparation method and application thereof |
| CN115160381B (en) * | 2022-08-18 | 2023-12-05 | 安徽工业大学 | Glutathione-responsive fluorescent probe, preparation method and application thereof |
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