CN107245085B - Iridium complex with tubulin recognition function and preparation method and application thereof - Google Patents
Iridium complex with tubulin recognition function and preparation method and application thereof Download PDFInfo
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- CN107245085B CN107245085B CN201710559588.8A CN201710559588A CN107245085B CN 107245085 B CN107245085 B CN 107245085B CN 201710559588 A CN201710559588 A CN 201710559588A CN 107245085 B CN107245085 B CN 107245085B
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- tubulin
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- 102000004243 Tubulin Human genes 0.000 title claims abstract description 40
- 108090000704 Tubulin Proteins 0.000 title claims abstract description 40
- GKOZUEZYRPOHIO-UHFFFAOYSA-N iridium atom Chemical compound [Ir] GKOZUEZYRPOHIO-UHFFFAOYSA-N 0.000 title claims abstract description 28
- 229910052741 iridium Inorganic materials 0.000 title claims abstract description 23
- 238000002360 preparation method Methods 0.000 title claims abstract description 13
- 239000003446 ligand Substances 0.000 claims abstract description 8
- DGEZNRSVGBDHLK-UHFFFAOYSA-N [1,10]phenanthroline Chemical compound C1=CN=C2C3=NC=CC=C3C=CC2=C1 DGEZNRSVGBDHLK-UHFFFAOYSA-N 0.000 claims abstract description 5
- OKKJLVBELUTLKV-UHFFFAOYSA-N Methanol Chemical compound OC OKKJLVBELUTLKV-UHFFFAOYSA-N 0.000 claims description 24
- YMWUJEATGCHHMB-UHFFFAOYSA-N Dichloromethane Chemical compound ClCCl YMWUJEATGCHHMB-UHFFFAOYSA-N 0.000 claims description 18
- 239000000523 sample Substances 0.000 claims description 14
- 238000006243 chemical reaction Methods 0.000 claims description 13
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Chemical compound O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims description 11
- 230000015572 biosynthetic process Effects 0.000 claims description 10
- 238000003786 synthesis reaction Methods 0.000 claims description 10
- 239000011888 foil Substances 0.000 claims description 6
- 230000000873 masking effect Effects 0.000 claims description 6
- 239000012046 mixed solvent Substances 0.000 claims description 6
- 239000000047 product Substances 0.000 claims description 6
- 239000007787 solid Substances 0.000 claims description 6
- ZNQVEEAIQZEUHB-UHFFFAOYSA-N 2-ethoxyethanol Chemical compound CCOCCO ZNQVEEAIQZEUHB-UHFFFAOYSA-N 0.000 claims description 5
- 239000012153 distilled water Substances 0.000 claims description 5
- YNPNZTXNASCQKK-UHFFFAOYSA-N Phenanthrene Natural products C1=CC=C2C3=CC=CC=C3C=CC2=C1 YNPNZTXNASCQKK-UHFFFAOYSA-N 0.000 claims description 4
- 238000004440 column chromatography Methods 0.000 claims description 4
- 238000000926 separation method Methods 0.000 claims description 4
- -1 ammonium hexafluorophosphate Chemical compound 0.000 claims description 3
- 239000012043 crude product Substances 0.000 claims description 3
- 238000004821 distillation Methods 0.000 claims description 3
- 239000003480 eluent Substances 0.000 claims description 3
- 238000010438 heat treatment Methods 0.000 claims description 3
- 239000000843 powder Substances 0.000 claims description 3
- 239000002904 solvent Substances 0.000 claims description 3
- 238000003756 stirring Methods 0.000 claims description 3
- MJRFDVWKTFJAPF-UHFFFAOYSA-K trichloroiridium;hydrate Chemical compound O.Cl[Ir](Cl)Cl MJRFDVWKTFJAPF-UHFFFAOYSA-K 0.000 claims description 3
- 238000005406 washing Methods 0.000 claims description 3
- 210000004027 cell Anatomy 0.000 abstract description 26
- IOJUPLGTWVMSFF-UHFFFAOYSA-N benzothiazole Chemical class C1=CC=C2SC=NC2=C1 IOJUPLGTWVMSFF-UHFFFAOYSA-N 0.000 abstract description 6
- 150000001875 compounds Chemical class 0.000 abstract description 5
- 230000011278 mitosis Effects 0.000 abstract description 3
- 238000011160 research Methods 0.000 abstract description 3
- 102000029749 Microtubule Human genes 0.000 abstract 1
- 108091022875 Microtubule Proteins 0.000 abstract 1
- 238000004020 luminiscence type Methods 0.000 abstract 1
- 210000004688 microtubule Anatomy 0.000 abstract 1
- IAZDPXIOMUYVGZ-WFGJKAKNSA-N Dimethyl sulfoxide Chemical compound [2H]C([2H])([2H])S(=O)C([2H])([2H])[2H] IAZDPXIOMUYVGZ-WFGJKAKNSA-N 0.000 description 4
- 238000004043 dyeing Methods 0.000 description 4
- 239000007789 gas Substances 0.000 description 4
- 238000000034 method Methods 0.000 description 4
- 239000012452 mother liquor Substances 0.000 description 4
- FWBHETKCLVMNFS-UHFFFAOYSA-N 4',6-Diamino-2-phenylindol Chemical compound C1=CC(C(=N)N)=CC=C1C1=CC2=CC=C(C(N)=N)C=C2N1 FWBHETKCLVMNFS-UHFFFAOYSA-N 0.000 description 3
- 230000000694 effects Effects 0.000 description 3
- 238000010166 immunofluorescence Methods 0.000 description 3
- 201000007270 liver cancer Diseases 0.000 description 3
- 208000014018 liver neoplasm Diseases 0.000 description 3
- 230000008685 targeting Effects 0.000 description 3
- 239000002775 capsule Substances 0.000 description 2
- 230000032823 cell division Effects 0.000 description 2
- 239000003795 chemical substances by application Substances 0.000 description 2
- 238000013461 design Methods 0.000 description 2
- 239000000975 dye Substances 0.000 description 2
- 239000000835 fiber Substances 0.000 description 2
- 238000001914 filtration Methods 0.000 description 2
- 239000001963 growth medium Substances 0.000 description 2
- 238000011081 inoculation Methods 0.000 description 2
- 229910052751 metal Inorganic materials 0.000 description 2
- 239000002184 metal Substances 0.000 description 2
- VNWKTOKETHGBQD-UHFFFAOYSA-N methane Chemical compound C VNWKTOKETHGBQD-UHFFFAOYSA-N 0.000 description 2
- 239000003226 mitogen Substances 0.000 description 2
- 238000002156 mixing Methods 0.000 description 2
- 239000002244 precipitate Substances 0.000 description 2
- 239000000126 substance Substances 0.000 description 2
- 238000012549 training Methods 0.000 description 2
- 238000001644 13C nuclear magnetic resonance spectroscopy Methods 0.000 description 1
- 238000005160 1H NMR spectroscopy Methods 0.000 description 1
- 102100021238 Dynamin-2 Human genes 0.000 description 1
- 101000817607 Homo sapiens Dynamin-2 Proteins 0.000 description 1
- 102000006830 Luminescent Proteins Human genes 0.000 description 1
- 108010047357 Luminescent Proteins Proteins 0.000 description 1
- 206010028980 Neoplasm Diseases 0.000 description 1
- 241001597008 Nomeidae Species 0.000 description 1
- 230000010721 Tubulin Interactions Effects 0.000 description 1
- JXLYSJRDGCGARV-WWYNWVTFSA-N Vinblastine Natural products O=C(O[C@H]1[C@](O)(C(=O)OC)[C@@H]2N(C)c3c(cc(c(OC)c3)[C@]3(C(=O)OC)c4[nH]c5c(c4CCN4C[C@](O)(CC)C[C@H](C3)C4)cccc5)[C@@]32[C@H]2[C@@]1(CC)C=CCN2CC3)C JXLYSJRDGCGARV-WWYNWVTFSA-N 0.000 description 1
- 238000009825 accumulation Methods 0.000 description 1
- 150000001450 anions Chemical class 0.000 description 1
- 230000009286 beneficial effect Effects 0.000 description 1
- 201000011510 cancer Diseases 0.000 description 1
- 230000008859 change Effects 0.000 description 1
- NEHMKBQYUWJMIP-UHFFFAOYSA-N chloromethane Chemical class ClC NEHMKBQYUWJMIP-UHFFFAOYSA-N 0.000 description 1
- 210000004292 cytoskeleton Anatomy 0.000 description 1
- 231100000135 cytotoxicity Toxicity 0.000 description 1
- 230000003013 cytotoxicity Effects 0.000 description 1
- 231100000263 cytotoxicity test Toxicity 0.000 description 1
- 230000006378 damage Effects 0.000 description 1
- 238000011161 development Methods 0.000 description 1
- 230000018109 developmental process Effects 0.000 description 1
- 125000003963 dichloro group Chemical group Cl* 0.000 description 1
- 230000004069 differentiation Effects 0.000 description 1
- 239000000539 dimer Substances 0.000 description 1
- 238000002330 electrospray ionisation mass spectrometry Methods 0.000 description 1
- 238000007306 functionalization reaction Methods 0.000 description 1
- 229910052739 hydrogen Inorganic materials 0.000 description 1
- 239000001257 hydrogen Substances 0.000 description 1
- 125000004435 hydrogen atom Chemical group [H]* 0.000 description 1
- 238000003384 imaging method Methods 0.000 description 1
- 238000001727 in vivo Methods 0.000 description 1
- 230000003834 intracellular effect Effects 0.000 description 1
- 150000002503 iridium Chemical class 0.000 description 1
- 239000002502 liposome Substances 0.000 description 1
- 238000013332 literature search Methods 0.000 description 1
- 239000000203 mixture Substances 0.000 description 1
- 238000006116 polymerization reaction Methods 0.000 description 1
- 230000008569 process Effects 0.000 description 1
- 239000002994 raw material Substances 0.000 description 1
- 238000005215 recombination Methods 0.000 description 1
- 230000006798 recombination Effects 0.000 description 1
- 210000005084 renal tissue Anatomy 0.000 description 1
- 238000010186 staining Methods 0.000 description 1
- 210000001519 tissue Anatomy 0.000 description 1
- 230000025366 tissue development Effects 0.000 description 1
- 231100000419 toxicity Toxicity 0.000 description 1
- 230000001988 toxicity Effects 0.000 description 1
- 229960003048 vinblastine Drugs 0.000 description 1
- JXLYSJRDGCGARV-XQKSVPLYSA-N vincaleukoblastine Chemical compound C([C@@H](C[C@]1(C(=O)OC)C=2C(=CC3=C([C@]45[C@H]([C@@]([C@H](OC(C)=O)[C@]6(CC)C=CCN([C@H]56)CC4)(O)C(=O)OC)N3C)C=2)OC)C[C@@](C2)(O)CC)N2CCC2=C1NC1=CC=CC=C21 JXLYSJRDGCGARV-XQKSVPLYSA-N 0.000 description 1
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- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07F—ACYCLIC, CARBOCYCLIC OR HETEROCYCLIC COMPOUNDS CONTAINING ELEMENTS OTHER THAN CARBON, HYDROGEN, HALOGEN, OXYGEN, NITROGEN, SULFUR, SELENIUM OR TELLURIUM
- C07F15/00—Compounds containing elements of Groups 8, 9, 10 or 18 of the Periodic Table
- C07F15/0006—Compounds containing elements of Groups 8, 9, 10 or 18 of the Periodic Table compounds of the platinum group
- C07F15/0033—Iridium compounds
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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
- G01N21/6402—Atomic fluorescence; Laser induced fluorescence
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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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- C09—DYES; PAINTS; POLISHES; NATURAL RESINS; ADHESIVES; COMPOSITIONS NOT OTHERWISE PROVIDED FOR; APPLICATIONS OF MATERIALS NOT OTHERWISE PROVIDED FOR
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- C09K2211/00—Chemical nature of organic luminescent or tenebrescent compounds
- C09K2211/18—Metal complexes
- C09K2211/185—Metal complexes of the platinum group, i.e. Os, Ir, Pt, Ru, Rh or Pd
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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"
- G01N2021/6439—Measuring fluorescence of fluorescent products of reactions or of fluorochrome labelled reactive substances, e.g. measuring quenching effects, using measuring "optrodes" with indicators, stains, dyes, tags, labels, marks
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- Peptides Or Proteins (AREA)
Abstract
The invention discloses an iridium complex with a tubulin recognition function, a preparation method and application thereof, wherein the iridium complex with the tubulin recognition function has the following structural formula:according to the invention, a benzothiazole derivative is selected as a main ligand to regulate the luminescence of the complex, and the iridium complex Ir-Tb is designed and synthesized by introducing a phenanthroline to enlarge a conjugated system of the complex. Researches show that the complex has larger Stokes shift and long fluorescence life, the target molecule Ir-Tb can target tubulin in living cells, different cell mitosis periods can be observed by marking a microtubule structure, and the compound has wide application prospects in the field of life science researches.
Description
Technical field
The present invention relates to a kind of tubulin (tubulin) probes, specifically a kind of that there is tubulin to identify function
The complex of iridium and its preparation method and application of energy.
Background technique
Chief component of the micro-pipe as the cytoskeleton of all eukaryocytes, it is combined by tubulin and micro-pipe
Albumen composition, and tubulin is the dimer being joined together to form by alpha-tubulin and 'beta '-tubulin.Tubulin
As a kind of dynamic cytoskeletal protein, its assembly dynamic makes it have unique effect in cell division.Especially
In cell mitogen period, under the regulation of centerbody, micro-pipe further can polymerize to form spindle, and guide cell therefrom
The differentiation in phase to later period.Since the recombination of micro-pipe net is the necessary process of cell life cycle and cell division function, so micro-
The content exception and normal alignment of tubulin, the destruction of polymerization are all related with the canceration of cell.Targeting tubulin is reported
Fluorescence probe is mostly organic micromolecule compound, and small organic molecule stoke shift and fluorescence lifetime are all shorter, vulnerable to cell
The influence of interior autofluorescence.Therefore, design synthesizes the tubulin probe of big stoke shift and long fluorescence lifetime for into one
The relationship probed between micro-pipe and cancer is walked to be of great significance.
In recent decades, metal organic complex due to its longer fluorescence lifetime and stoke shift it is micro- in living cells
Imaging aspect using more and more extensive, reported in succession with selectively targeted metal organic complex fluorescence probe, example
Such as, the Complex probe with liposome target function [Accounts of chemical research, 2015,48,
2985], have DNA target to [Nature chemistry, 2009,1,662] and the Complex probe with BSA target function
[Chemistry-A European Journal 2009,15,3652].But up to the present, target the triplet state of tubulin
There is not been reported for iridium complexes probe.
Applicant has carried out following literature search to the theme of the application:
1, xueshu.glgoo.com net search result: (2017/07/05)
2, middle National IP Network's search result:
Retrieval mode one:
Piece name-has the complex of iridium of tubulin identification function: without pertinent literature.
A piece a kind of complex of iridium and preparation method thereof with tubulin identification function of name-: without pertinent literature.
Retrieval mode two:
In full-complex of iridium with tubulin identification function: without pertinent literature.
In full-a kind of complex of iridium and preparation method thereof with tubulin identification function: without pertinent literature.
Summary of the invention
The present invention is intended to provide a kind of complex of iridium and its preparation method and application with tubulin identification function.This
Invention selects benzothiazole derivant to shine as main ligand adjusting complex, passes through and introduces being total to for Phen expansion complex
Yoke system, design have synthesized complex of iridium Ir-Tb.The study found that complex has biggish stoke shift and long fluorescence
Service life, and target molecule Ir-Tb can target tubulin in living cells, observe cell mitogen by label micro-tubular structure
Different times, the compound have broad application prospects in life science field.
Benzothiazole is similar to a kind of vinblastine (known tubulin probe) structure, can pass through functionalization benzo
Thiazole end group increases the binding site of compound and tubulin interaction (hydrogen bond, pi-pi accumulation effect etc.), improves
The specificity of targeting compounds tubulin.
The present invention has the structural formula of the complex of iridium (Ir-Tb) of tubulin identification function are as follows:
The present invention has the preparation method of the complex of iridium of tubulin identification function, includes the following steps:
Step 1: the synthesis of intermediate M
Logical N2Under gas shielded, ligand L 0.60g (2.2mmol) is dissolved in 20mL ethylene glycol monoethyl ether and distilled water mixing is molten
In agent (3:1, v/v), it is added to the 100mL Shrek bottle of masking foil package after to be dissolved, adds and is dissolved in the three of 10mL water
Chloride hydrate iridium solution 0.33g (1.0mmol), back flow reaction is for 24 hours at 110 DEG C；It is cooled to room temperature after reaction, filtering is used in combination
A small amount of distillation water washing precipitates, and dry 12h, obtains orange solids powder, as intermediate M in vacuum oven.
Ligand L synthesis reference literature method (Bioorganic&Medicinal Chemistry Letters, 2011,
21(8),2445-2449)。
Step 2: the synthesis of target product Ir-Tb
Masking foil wraps up 100mL Shrek reaction flask, leads to N2Under gas shielded, weigh intermediate M0.68g (0.50mmol) and
Phen 0.36g (2mmol) is dissolved in 50mL methylene chloride and methanol mixed solvent (1:1, v/v), then anti-at 70 DEG C
It should for 24 hours；It is cooled to room temperature after reaction, is added excess ammonium hexafluorophosphate 0.10g (0.50mmol), stirring at normal temperature 12h is obtained
Red solution, heating boil off most of solvent, filter, obtain red crude product 1.80g, (eluent is dichloro to column chromatography for separation
100:1 is mixed to get by volume for methane and methanol) after obtain Orange red solid 0.81g, as target product Ir-Tb, yield
76.20%.
The synthetic route of complex of iridium of the present invention is as follows:
The present invention has the purposes of the complex of iridium of tubulin identification function, is as tubulin in identification living cells
Fluorescence probe application.
The biological study of Ir-Tb of the present invention is as follows:
By liver cancer cells (HepG2) kind on laser co-focusing capsule, each hole inoculation 104A cell, with the training of 1.5mL
Support base culture 24-36h.Ir-Tb is made into 10-3The mother liquor of M the Ir-Tb mother liquor of 15 μ L is added into culture medium, in cell incubator
Middle dyeing 30min is washed three times with PBS, be can be directly used for developing.
The beneficial effects of the present invention are embodied in:
1, the raw material of Ir-Tb of the present invention is cheap and easy to get, and synthesis is simple and efficient, and makes it possible that it is commercialized.
2, the toxicity of Ir-Tb of the present invention is low (Fig. 2), has good biocompatibility, convenient for being applied to organism.
3, Ir-Tb of the present invention can the intracellular tubulin of specific recognition (Fig. 4), Ir-Tb and tubulin- is immunized glimmering
Common location effect is good in the cell for photoprotein (Tb-Ab), and Pearson's coefficient can reach 89.72%.
4, Ir-Tb of the present invention can observe cell difference mitosis period spindle fiber, spindle by marking tubulin
Structure change, to judge the division period (Fig. 5) of cell.
5, Ir-Tb of the present invention acts not only as targeting living cells tubulin probe, additionally it is possible to for tissue development (figure
6).There is no similar available commercial biological fluorescence probe, commercial values with higher.
Detailed description of the invention
Fig. 1 is the monocrystalline ellipsoid figure (omitting anion and H atom) of Ir-Tb, show synthesis complex of iridium be there is not yet
Report and the specific novel substance of structure.
Fig. 2 is the complex Ir-Tb under various concentration and the cytotoxicity test after liver cancer cells culture for 24 hours, illustrates Ir-
The cytotoxicity of Tb is low, has good biocompatibility.
Fig. 3 (a) is that Ir-Tb of the present invention, quotient contaminate DAPI and tubulin- immunofluorescence albumen Tb-Ab being total in living cells
Locating development figure, as can be seen from the figure the dyeing coincidence factor of Ir-Tb and Tb-Ab is higher, and Ir-Tb and nucleus dyestuff DAPI
Dyeing coincidence factor it is lower；It (b) is Ir-Tb and tubulin- immunofluorescence albumen Tb-Ab common location related coefficient figure, Pearson
Coefficient is up to 0.8972, illustrates that Ir-Tb can target the tubulin in living cells.
Fig. 4 is to observe Ir-Tb and core quotient dye in the staining conditions of each division stage: (a) in different division stages using cell
Early period, (b) prometaphase, (c) mid-term, (d) latter stage.Illustrate that Ir-Tb can be as tubulin in label spindle and spindle fiber
Feature probes, can be used for observe be in mitosis each period cell.
Fig. 5 is that Ir-Tb of the present invention, quotient contaminate DAPI and tubulin- immunofluorescence albumen Tb-Ab in Vivo of Renal tissue
Common location develop figure, illustrate that Ir-Tb acts not only as cell-targeting tubulin probe, additionally it is possible to for tissue develop.
Specific embodiment
The present invention has the structural formula of the complex of iridium (Ir-Tb) of tubulin identification function are as follows:
The present invention has the preparation method of the complex of iridium of tubulin identification function, includes the following steps:
Step 1: the synthesis of intermediate M
Logical N2Under gas shielded, ligand L 0.60g (2.2mmol) is dissolved in 20mL ethylene glycol monoethyl ether and distilled water mixing is molten
In agent (3:1, v/v), it is added to the 100mL Shrek bottle of masking foil package after to be dissolved, adds and is dissolved in the three of 10mL water
Chloride hydrate iridium solution 0.33g (1.0mmol), back flow reaction is for 24 hours at 110 DEG C；It is cooled to room temperature after reaction, filtering is used in combination
A small amount of distillation water washing precipitates, and dry 12h, obtains orange solids powder, as intermediate M in vacuum oven.
Ligand L synthesis reference literature method (Bioorganic&Medicinal Chemistry Letters, 2011,
21(8),2445-2449)。
Step 2: the synthesis of target product Ir-Tb
Masking foil wraps up 100mL Shrek reaction flask, leads to N2Under gas shielded, intermediate M 0.68g (0.50mmol) is weighed
It is dissolved in 50mL methylene chloride and methanol mixed solvent (1:1, v/v) with Phen 0.36g (2mmol), then at 70 DEG C
Reaction is for 24 hours；It is cooled to room temperature after reaction, is added excess ammonium hexafluorophosphate 0.10g (0.50mmol), stirring at normal temperature 12h is obtained
To red solution, heating boils off most of solvent, filters, obtains red crude product 1.80g, column chromatography for separation (eluent two
100:1 is mixed to get by volume for chloromethanes and methanol) after obtain Orange red solid 0.81g, as target product Ir-Tb, receive
Rate 76.20%.
1H NMR(400MHz,DMSO-d6), δ (ppm): 9.52 (s, 2H), 8.92 (d, J=8.3Hz, 2H), 8.49 (d, J
=5.1Hz, 2H), 8.32 (s, 2H), 8.14 (dd, J=8.2,5.1Hz, 2H), 8.04 (d, J=8.1Hz, 2H), 7.52 (s,
2H), 7.17 (t, J=7.7Hz, 2H), 6.82 (t, J=7.9Hz, 2H), 5.89 (s, 2H), 5.60 (d, J=8.4Hz, 2H),
4.17 (m, 4H), 1.36 (t, J=6.9Hz, 6H)；13C NMR(100MHz,DMSO-d6)δ(ppm):180.41,151.34,
151.15,148.78,146.92,144.43,143.71,139.16,130.60,130.50,130.19,128.26,127.30,
124.68,124.01,119.37,115.42,111.98,64.24,14.83；IR(KBr,cm-1):3053.63(w),2359.43
(w),2341.56(w),1602.77(s),1519.55(s),1480.79(s),1434.58(s),1408.33(m),1314.93
(m),1227.40(s),1155.98(s),1097.13(m),967.47(s),835.99(s),807.76(m),755.37(s),
728.819(s),696.45(m),622.35(s),550.68(m),522.19(m),498.99(m)；ESI-MS:
cal.913.13,found 913.33.
The synthetic route of complex of iridium of the present invention is as follows:
The biological study of Ir-Tb of the present invention is as follows:
By liver cancer cells (HepG2) kind on laser co-focusing capsule, each hole inoculation 104A cell, with the training of 1.5mL
Support base culture 24-36h.Ir-Tb is made into 10-3The mother liquor of M the Ir-Tb mother liquor of 15 μ L is added into culture medium, in cell incubator
Middle dyeing 30min is washed three times with PBS, be can be directly used for developing.
Claims (6)
1. a kind of complex of iridium with tubulin identification function, it is characterised in that its structural formula are as follows:
2. a kind of preparation method of the complex of iridium described in claim 1 with tubulin identification function, it is characterised in that
Include the following steps:
Step 1: the synthesis of intermediate M
Logical N2Under protection, ligand L 0.60g is dissolved in 20mL ethylene glycol monoethyl ether and distilled water in the mixed solvent, is added after to be dissolved
Enter the 100mL Shrek bottle wrapped up to masking foil, adds the three chloride hydrate iridium solution 0.33g for being dissolved in 10mL water, 110 DEG C
Lower back flow reaction is for 24 hours；It is cooled to room temperature, filters and is precipitated with a small amount of distillation water washing after reaction, done in vacuum oven
Dry 12h obtains orange solids powder, as intermediate M；
The structural formula of the ligand L are as follows:
Step 2: the synthesis of target product
Masking foil wraps up 100mL Shrek reaction flask, leads to N2Under protection, weighs intermediate M 0.68g and Phen 0.36g is molten
In 50mL methylene chloride and methanol mixed solvent, then reacted for 24 hours at 70 DEG C；It is cooled to room temperature after reaction, is added
Excessive ammonium hexafluorophosphate 0.10g, stirring at normal temperature 12h obtain red solution, and heating boils off most of solvent, filter, obtain red
Crude product 1.80g obtains Orange red solid, as target product after column chromatography for separation.
3. preparation method according to claim 2, it is characterised in that:
In step 1, the volume ratio of ethylene glycol monoethyl ether and distilled water in the mixed solvent ethylene glycol monoethyl ether and distilled water is 3:1.
4. preparation method according to claim 2, it is characterised in that:
In step 2, the volume ratio of methylene chloride and methanol is 1:1 in methylene chloride and methanol mixed solvent.
5. preparation method according to claim 2, it is characterised in that:
In step 2, eluent when column chromatography for separation is that 100:1 is mixed to get by volume for methylene chloride and methanol.
6. a kind of purposes of the complex of iridium described in claim 1 with tubulin identification function, it is characterised in that: be to make
For the application of the fluorescence probe of tubulin in identification living cells.
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| CN105061515A (en) * | 2015-08-08 | 2015-11-18 | 赣南师范学院 | Synthesis of phosphorescent iridium complex and application of phosphorescent iridium complex for fluorescence labeling of schistosome cercaria |
| CN106117270A (en) * | 2016-05-27 | 2016-11-16 | 苏州科技学院 | A kind of annular metal iridium complex and preparation method thereof and the application as protein staining agent |
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| CN105061515A (en) * | 2015-08-08 | 2015-11-18 | 赣南师范学院 | Synthesis of phosphorescent iridium complex and application of phosphorescent iridium complex for fluorescence labeling of schistosome cercaria |
| CN106117270A (en) * | 2016-05-27 | 2016-11-16 | 苏州科技学院 | A kind of annular metal iridium complex and preparation method thereof and the application as protein staining agent |
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| Protein staining agents from cationic and neutral luminescent iridium(Ⅲ) complexes;Yuyang Zhou等;《Chemisty A European Journal Communication》;20161231;第22卷;第16796-16800页 * |
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