CN108727255A - A kind of phenoxy bridge connection double-core copper（II）Complex and its preparation and application - Google Patents
A kind of phenoxy bridge connection double-core copper（II）Complex and its preparation and application Download PDFInfo
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Abstract
The present invention relates to a kind of phenoxy bridges to join double-core copper（II）Complex and its preparation and application.Described phenoxy bridge connection double-core copper (II) complex has efficient antitumor action, and can be used as DNA cutting reagents and applied.The chemical formula of described phenoxy bridge connection double-core copper (II) complex is [Cu2(BHPMP)] Cl, wherein [Cu2(BHPMP)]+With specific structure formula.The preparation method of the complex is to make CuCl2With the H of specific structure3BHPMP carries out complex reaction in acetonitrile solvent, in 40~60 DEG C, and [Cu is obtained after purification2(BHPMP)] Cl solids.Present invention firstly discovers that phenoxy bridge connection copper (II) complex has stronger inhibiting effect to human cervical carcinoma cell (HeLa), non-small cell type lung carcinoma cell (A-549) and human breast cancer cell (MCF-7), significant antitumor activity is shown.
Description
Technical field
The present invention relates to technical field of chemistry, more particularly to a kind of phenoxy bridge connection double-core copper (II) complex and its prepare and
Using.
Background technology
Artificial nucleic acid cutting agent has important potential using value in molecular biotechnology and new drug development field.Transition
Metal complex possesses abundant structure and changeable functional character, is widely used in the research of artificial nucleic acid cutting agent.
In numerous transient metal complexes, copper complex is stronger with suitable oxidation-reduction potential and with DNA due to it
Binding ability receives the favor of researcher.In the presence of oxidant or reducing agent, copper complex, which generates active oxygen species, to be caused
DNA chain is broken or base is modified and shows cleavage activity.In copper complex cuts DNA redox, most generation
One example of table is exactly [Cu (OP)2]2+(OP=1,10- phenanthroline), this is also the Chemistry Nuclease of first case report, it
Higher cutting efficiency is shown to B-form DNA.In general, the active site of most of natural acid enzymes all contain there are two or
The metal ion of multiple synergistic effects, this synergistic effect tend to improve cutting efficiency.Increase cutting efficiency and improve and selects
Selecting property is the center of gravity of artificial nuclease research work, shows multinucleation currently as the metal complex of artificial nuclease research
Trend, it is desirable to using polynuclear complex metal center synergistic effect promote cutting efficiency.In recent years, coordinated with copper
Object emerges one after another as the report of artificial nuclease, and wherein double-core and multinuclear copper complex has attracted researchers more to close
Note.Studies have shown that the synergistic effect at multi-nuclear metal center is convenient for forming reactive intermediate, to promote to aoxidize cutting efficiency and
Enhancing selectivity.
Cancer seriously threaten the life and health of the mankind, according to World Health Organization 2014《Global cancer report》It is aobvious
Show, the newly-increased cancer patient in the whole world in 2012 reaches 14,000,000, while cancer mortality patient numbers, also in huge increasing, cancer has become entirely
Mankind killer the most terrified.Treatment to cancer and its urgent, it is extremely urgent to develop novel efficient anticancer drug.?
In numerous anticancer drugs, platinum medicine use it is the most extensive, it is suitable from the approval of U.S. Food and Drug Administration in 1978
It is significant in efficacy to reproductive system cancers and incidence cancer after platinum enters Clinical practice.Subsequent researcher has developed the again
Two generation platinum-containing anticancer drug carboplatins and third generation platinum-containing anticancer drug oxaliplatin, the platinum-containing anticancer drug clinically used at present
Also Nedaplatin, Lobaplatin and Eptaplatin, in addition kind of platinum complexes are in different clinical experimental stages also more than ten.However,
The strong toxic side effect of platinum medicine and drug resistance and itself with human body in sulfur-bearing biomolecule action and inactivate and generate pair
Effect, this seriously constrains its curative effect and long-time service.Researcher has turned one's attention to non-platinum metals, it is desirable to therefrom screen
Go out can overcome the disadvantages that the new type anticancer reagent of platinum-containing anticancer drug defect.Copper is the activated centre of a variety of large biological molecules, is lived in life
Very important effect is played in dynamic.Copper complex is because of its hypotoxicity and high efficiency, it is considered to be platinum-containing anticancer drug most has uncommon
The replacer of prestige.Currently, some copper complexes show good active anticancer during cytotoxic activity is tested in vivo or in vitro,
It is potential in the future to become antitumor and anticancer agent.
In conclusion double-core or multinuclear copper (II) complex show wide application prospect in biomedicine field, change
It learns and provides new thinking and technical support with biomedical subject crossing for the biologic applications of copper complex.Therefore, copper
(II) complex causes the great interest of chemist and pharmacy man as the research of DNA cutting reagents and antitumor drug.
In the implementation of the present invention, the inventors discovered that having at least the following problems in the prior art：Develop and grinds
Study carefully new hypotoxicity double-core or multinuclear copper (II) complex, there is efficient antitumor action.
Invention content
In consideration of it, a kind of phenoxy bridge connection double-core copper (II) complex of present invention offer and its preparation and application, the phenol oxygen
Bridged binuclear copper (II) complex has efficient antitumor action, and can be used as DNA cutting reagents and applied.
Specifically, including technical solution below：
According to the first aspect of the invention, the present invention provides a kind of phenoxy bridges to join double-core copper (II) complex, the phenol
The chemical formula of oxygen bridged binuclear copper (II) complex is [Cu2(BHPMP)] Cl, wherein [Cu2(BHPMP)]+Structural formula it is as follows：
According to the second aspect of the invention, the present invention provides the preparation sides that the phenoxy bridge joins double-core copper (II) complex
Method：
Make CuCl2And H3BHPMP carries out complex reaction in acetonitrile solvent, in 40~60 DEG C (such as 50 DEG C), after purification
To [Cu2(BHPMP)] Cl solids, wherein H3The structural formula of BHPMP is as follows：
Specifically, the H3The preparation method of BHPMP is：By (2- hydroxybenzyls) (2- pyridylmethyls) amine tetrahydrochysene furan
It mutters dissolving, under ice-water bath protection, adjusts pH value of solution to 7~8 with triethylamine, 2,6- dichloromethyl -4- first is then added dropwise thereto
The tetrahydrofuran solution of base phenol after being added dropwise, is stirred overnight, commonly filters to get filtrate at room temperature, and vacuum distillation removes molten
Agent obtains yellow oil, is washed successively with water, ethyl alcohol, ether, is dried in vacuo to obtain yellow solid, as H3BHPMP ligands.
The specific method of the purification is that reaction mixture is filtered under diminished pressure distillation and concentration, ether is added, bottle green is precipitated
Precipitation, centrifugation obtain solid, [Cu are obtained after washing, vacuum drying2(BHPMP)] Cl solids.
Further, the washing is that sequentially acetone, ether wash the solid that centrifugation obtains.
Specifically,
By CuCl2·2H2O and H3BHPMP ligands press 2:1 ratio mixing, is dissolved in acetonitrile solvent, 40~60 DEG C of heating
4~6h is stirred, purification is to get to [Cu after stopping reaction2(BHPMP)] Cl solids.The specific reaction time is to react no longer apparent
It, can be tests determined by those skilled in the art subject to progress.
According to the third aspect of the invention we, the present invention also provides phenoxy bridge connection double-core copper (II) complexs to prepare
Application in antitumor drug.
Specifically, the tumour is human cervical carcinoma, non-small cell type lung cancer and human breast carcinoma.
According to the fourth aspect of the invention, the present invention also provides the phenoxy bridges to join double-core copper (II) complex conduct
The application of DNA cutting reagents.
The advantageous effect of technical solution provided in an embodiment of the present invention includes at least：
1, the present invention joins ligand as architecture basics using phenoxy bridge, in acetonitrile as solvent, under 40~60 DEG C of heating conditions, and phenol
Oxygen bridge ligand and CuCl2·2H2O occurs complex reaction and obtains double-core copper (II) complex, and synthetic method is simple, reaction condition
Mildly, easily controllable, obtained target product purity is high.
2, present invention firstly discovers that, phenoxy bridge connection copper (II) complex is to human cervical carcinoma cell (HeLa), non-small thin
Born of the same parents' type lung carcinoma cell (A-549) and human breast cancer cell (MCF-7) have stronger inhibiting effect, show significant antitumor
Activity.
3, phenoxy bridge connection copper (II) complex of the present invention can be combined in a manner of groove contact with DNA；In ascorbic acid
In the presence of can effectively convert super coiled DNA to and incise and linear DNA, show preferably to aoxidize cleavage activity.
Description of the drawings
To describe the technical solutions in the embodiments of the present invention more clearly, make required in being described below to embodiment
Attached drawing is briefly described, it should be apparent that, drawings in the following description are only some embodiments of the invention, for
For those of ordinary skill in the art, without creative efforts, other are can also be obtained according to these attached drawings
Attached drawing.
Fig. 1 is under the conditions of 25 DEG C, 5mM Tris-HCl buffer solutions (pH=7.4), and phenoxy bridge of the present invention joins double-core
(embedded figure is [DNA]/(ε to copper (II) complex with the ultra-violet absorption spectrum in the presence of various concentration CT-DNAa-εf)vs.
[DNA], wherein [DNA] is CT-DNA concentration).
Fig. 2 is CT-DNA-EB systems ([DNA]=[EB]=1.136 × 10-6M phenoxy bridge connection of the present invention is added in)
Double-core copper (II) complex (1.0 × 10-310 μ L run-downs are often added dropwise in M) after fluorescence spectrum (embedded figure is F0/F vs.
[complex], wherein abscissa [complex] are complex concentration, and F is the fluorescence intensity of solution after complex is added dropwise, F0For
The fluorescence intensity of CT-DNA-EB system solutions before dropwise addition complex), prolong arrows direction, the corresponding complex of curve is dense
Degree gradually increases.
Fig. 3 is that phenoxy bridge of the present invention joins double-core copper (II) complex and the CD of CT-DNA effects schemes, wherein dotted line pair
The DNA (1.48 × 10 of complex is added in Ying Yuwei-4M) CD spectrum (i.e. a concentration of 0) of complex, in the direction of the arrow, complex
Concentration gradually increases.
Fig. 4 is in the presence of Vc, and the phenoxy bridge of the present invention of various concentration joins double-core copper (II) complex to pBR322
The gel electrophoresis figure of Plasmid DNA effect, wherein：
Lane 1,DNA；
lane 2,DNA+1mM Vc；
Lane 3, DNA+200 μM of phenoxy bridge join double-core copper (II) complex；
Lane 4, DNA+200 μM of ligand H3BHPMP；
Lane 5, DNA+200 μM of ligand H3BHPMP+1mM Vc；
Lane 6-18, it is double that DNA distinguishes+5,10,15,20,25,30,35,40,45,50,60,80,100 μM of phenoxy bridge connection
+ 100 times of excessive Vc of core copper (II) complex.
Fig. 5 be there are different activities oxygen killer in the case of, phenoxy bridge of the present invention joins double-core copper (II) complex pair
The gel electrophoresis figure of pBR322 Plasmid DNA effect, wherein：
Lane 1,DNA；
Lane 2, DNA+1mM Vc+ phenoxy bridges join double-core copper (II) complex；
Lane 3, DNA+1mM Vc+ phenoxy bridges join double-core copper (II) complex+1M DMSO (dimethyl sulfoxide (DMSO))；
Lane 4, DNA+1mM Vc+ phenoxy bridges join double-core copper (II) complex+10mM KI；
Lane 5, DNA+1mM Vc+ phenoxy bridges join double-core copper (II) complex+10mM NaN3；
Lane 6, DNA+1mM Vc+ phenoxy bridges join double-core copper (II) complex+D2O (70%)；
Lane 7, DNA+1mM Vc+ phenoxy bridges join double-core copper (II) complex+1M tert-butyl alcohols.
Specific implementation mode
To keep technical scheme of the present invention and advantage clearer, below in conjunction with attached drawing to embodiment of the present invention make into
One step it is described in detail.
According to the first aspect of the invention, the present invention provides a kind of phenoxy bridges to join double-core copper (II) complex, the phenol
The chemical formula of oxygen bridged binuclear copper (II) complex is [Cu2(BHPMP)] Cl, wherein [Cu2(BHPMP)]+Structural formula it is as follows：
According to the second aspect of the invention, the present invention provides the preparation sides that the phenoxy bridge joins double-core copper (II) complex
Method：
Make CuCl2And H3BHPMP carries out complex reaction in acetonitrile solvent, in 40~60 DEG C, and [Cu is obtained after purification2
(BHPMP)] Cl solids, wherein H3The structural formula of BHPMP is as follows：
Specifically, the H3The preparation method of BHPMP is：By (2- hydroxybenzyls) (2- pyridylmethyls) amine tetrahydrochysene furan
It mutters dissolving, under ice-water bath protection, adjusts pH value of solution to 7~8 with triethylamine, 2,6- dichloromethyl -4- first is then added dropwise thereto
The tetrahydrofuran solution of base phenol after being added dropwise, is stirred overnight, commonly filters to get filtrate at room temperature, and vacuum distillation removes molten
Agent obtains yellow oil, is washed successively with water, ethyl alcohol, ether, is dried in vacuo to obtain yellow solid, as H3BHPMP ligands.(2-
Hydroxybenzyl) molar ratio of (2- pyridylmethyls) amine and 2,6- dichloromethyl -4- methylphenols is (2.2~2.0):1.
The specific method of the purification is that reaction mixture is filtered under diminished pressure distillation and concentration, ether is added, bottle green is precipitated
Precipitation, centrifugation obtain solid, [Cu are obtained after washing, vacuum drying2(BHPMP)] Cl solids.
Further, the washing is to wash the solid that centrifugation obtains with acetone, ether.
Specifically,
By CuCl2·2H2O and H3BHPMP ligands press 2:1 ratio mixing, is dissolved in acetonitrile solvent, 40~60 DEG C of heating
4~6h is stirred, purification is to get to [Cu after stopping reaction2(BHPMP)] Cl solids.
According to the third aspect of the invention we, the present invention also provides phenoxy bridge connection double-core copper (II) complexs to prepare
Application in antitumor drug.
Specifically, the tumour is human cervical carcinoma, non-small cell type lung cancer and human breast carcinoma.
According to the fourth aspect of the invention, the present invention also provides the phenoxy bridges to join double-core copper (II) complex conduct
The application of DNA cutting reagents.
In following embodiment：
Calf thymus DNA (CT-DNA) is purchased from Sigma-Aldrich (Sigma-Aldrich) Chinese companies；Ethidium bromide
(EB) Sigma-Aldrich (Sigma-Aldrich) Chinese companies are purchased from；Super spirial plasmid pBR322DNA is purchased from precious biology
(Takara) company；Chemical reagent involved in synthetic compound process is conventional commercial analytical reagents.
Mass spectral analysis is measured with LCQ electrospray mass spectrometers (ESI-MS, Finnigan)；Uv-visible absorption spectra is used
Hitachi U-3010 ultraviolet-visual spectrometers measure；Fluorescence spectrum is measured with Hitachi F-4500 Fluorescence Spectrometer；Circle two
Color spectrum measures on Jasco-J-810 automatic spectrometers.
Embodiment 1
Synthesize H3BHPMP ligands
(2- hydroxybenzyls) (2- pyridylmethyls) amine (4.28g, 20mmol) is weighed, the dissolving of 25mL tetrahydrofurans is added,
Under ice-water bath, pH value of solution is adjusted to 7-8 with triethylamine, be then added dropwise thereto 2,6- dichloromethyl -4- methylphenols (2.05g,
Tetrahydrofuran solution 10mmol) after being added dropwise, is stirred overnight, commonly filters to get filtrate at room temperature, and vacuum distillation removes molten
Agent obtains yellow oil, is washed successively with water, ethyl alcohol, ether, is dried in vacuo to obtain yellow solid, as H3BHPMP ligands.Detection
As a result as follows：
Yield：82%.
Elemental analysis：C35H36N4O3(%)：found(calc.)C,74.36(75.00)；H,6.36(6.43)；N,11.15
(11.01).
Fourier infrared (KBr tablettings, ν/cm-1)：3429(br,s),2025(m),1603(m),1384(w),1104(s),
621(s).
Positive ionization electrospray mass spectrum ESI-MS (m/z, methanol)：561.33[M+H]+.
Embodiment 2
It synthesizes phenoxy bridge and joins double-core copper (II) complex
Complex [Cu2(BHPMP)] synthesis of Cl：Weigh ligand H3BHPMP (0.28g, 0.5mmol) is dissolved in acetonitrile, is used
2M NaOH solutions adjust its pH to 7-8, and CuCl is then added2·2H2The acetonitrile solution of O (0.21g, 1.0mmol), 50 DEG C of heating
Lower reaction 4h.After stopping reaction, reaction solution vacuum distillation is concentrated into about 2mL, a large amount of ether are added, bottle green precipitation is precipitated,
Solid is centrifuged to obtain, is washed with acetone and ether, is dried in vacuo, obtains green solid powder.Testing result is as follows：
Yield：80%.
Elemental analysis：Cu2C35H33N4O3Cl (%)：found(calc.):C,58.56(58.34)；H,4.62(4.58)；
N,7.72(7.78).
Positive ionization electrospray mass spectrum ESI-MS (m/z, methanol)：721.33[M-Cl+2H2O]+；757.13[M+2H+Cl]+；
791.07[M+2H+Cl+CH3OH]+.
Fourier infrared (KBr tablettings, v/cm-1):3446(br,s),3351(br,s),1615(s),1438(m),764
(m).
Molar conductivity ΛM(DMF)=80.8S cm2mol-1.
Embodiment 3
It is thin to human cervical carcinoma that phenoxy bridge of the present embodiment as obtained by experiment detection embodiment 2 joins double-core copper (II) complex
The inhibiting effect that born of the same parents (HeLa), human lung carcinoma cell (A-549) and human breast cancer cell (MCF-7) are proliferated.
Cytotoxic activity is tested：The present embodiment is studied phenoxy bridge connection double-core copper (II) of the present invention using mtt assay and is coordinated
Object is to the external thin of human cervical carcinoma cell (HeLa), non-small cell type lung carcinoma cell (A-549) and human breast cancer cell (MCF-7)
Born of the same parents' cytotoxic activity.Wherein cis-platinum is as positive controls.These tumour cells be added to heat inactivation fetal bovine serum (10%,
V/v), cultivated in the RPMI-1640 culture mediums of glutamine (2mM), penicillin (100U/mL) and streptomysin (100ug/mL),
Cultivate the 95% and 5%CO that atmosphere is high humility at 37 DEG C2.Main process is the cell inoculation that will be acquired from exponential phase of growth
To (every milliliter 2 × 10 in 96- well culture plates5It is a, per wellhole 100ul), make its adherency for 24 hours, is then waited for by concentration gradient addition
The compound of research makes the finally formed concentration of compound from 0.1 μm of ol/l to 10 μm of ol/l.96- well culture plates are incubated again
For 24 hours, the 20 μ l of MTT solution that concentration is about 5mg/ml then are added to each Kong Jingzhong, continuation is incubated 4h at 37 DEG C.Then
100 μ l " three component solutions " (10%SDS-5% isobutanol -12mM HCl) are added.Culture plate temperature at 37 DEG C is incubated overnight, is being divided
The absorbance of solution under 540nm wavelength is measured on light luminosity.Relative to not plus drug control group be calculated the hundred of survivaling cell
Divide ratio.Its IC50Value is obtained with " Logit " method from the semilog diagram of " percentage control-drug concentration ", and is defined as
Cell number can be caused to reduce by 50% drug concentration compared with not treating control group.The average result tested three times obtains to the end
IC50Value.Its result data is as shown in table 1, phenoxy bridge of the present invention connection double-core copper (II) complex to HeLa, A-549 and
The IC of MCF-7 cells50Value is respectively 10.5,9.4,11.2 μM；Under identical condition, anticancer drug cis-platinum is to HeLa, A-549
With the IC of MCF-7 cells50Value is respectively 9.6,9.2,5.6 μM.Find out from result, phenoxy bridge connection double-core copper (II) of the present invention
Complex has with the comparable cytotoxic activity of cis-platinum these three cancer cells, therefore phenoxy bridge of the present invention connection double-core copper (II)
Complex has preferable antitumous effect.
Table 1
Embodiment 4
The present embodiment joins double-core copper as uv-vis spectra to study the phenoxy bridge of the present invention obtained by embodiment 2
(II) interaction of complex and calf thymus DNA (CT-DNA).CT-DNA is dissolved in 5mM Tris-HCl/50mM NaCl
Aqueous solution in, pH=7.40 is preserved at 4 DEG C and is used in 4 days.The concentration of CT-DNA surveys its 260nm using ultraviolet spectrometer
The absorbance at place come determine (molar extinction coefficients of the CT-DNA at 260nm be 6600M-1cm-1, and the suction of 260nm and 280nm
The ratio between luminous intensity is 1.8-1.9, shows that the CT-DNA samples are free of protein).Phenoxy bridge connection double-core copper (II) of the present invention
The initial concentration of complex uses the buffer solution (5mM Tris-HCl, 50mM NaCl, pH 7.4) containing 10% methanol to prepare.Dense
Degree for 25 μM phenoxy bridge join double-core copper (II) complex solution in be separately added into a series of not same amounts (0,2.5,7.5,10.0,
12.5,15.0,20.0,22.5,25.0 μM) CT-DNA, and measure the variation of their ultraviolet-visible absorption spectroscopy, such as Fig. 1
Shown, phenoxy bridge of the present invention joins increase of absorbance of double-core copper (II) complex at 238nm with CT-DNA concentration
And increase, and with apparent red shift.This illustrates that phenoxy bridge connection double-core copper (II) complex of the present invention is by non-intrusive work
It is interacted with DNA.Phenoxy bridge connection double-core copper (II) complex and CT- of the present invention are calculated using formula 1
The binding constant K of DNA interactionsb：
[DNA]/(εa-εf)=[DNA]/(εb-εf)+1/[Kb(εb-εf)] (1)
Wherein, εaFor reaction system specified wavelength apparent molar extinction coefficient；εfIt is to dissociate complex in the wavelength
Molar extinction coefficient；εbIn the molar extinction coefficient of the wavelength after being combined with DNA for complex.By [DNA]/(εa-εf) right
[DNA] mapping (as Fig. 1 is embedded shown in figure), can obtain phenoxy bridge connection double-core copper (II) complex-bound constant difference of the present invention
For 7.2 (± 0.02) × 104M-1。
Embodiment 5
The present embodiment using fluorescence spectrum experiments study be the 2 gained phenoxy bridge of embodiment join double-core copper (II) complex
It is tested with the competitiveness of ethidium bromide (EB).A concentration of 1.136 × 10-6Concentration is gradually added dropwise in the CT-DNA-EB system solutions of M
It is 1 × 10-3M phenoxy bridge of the present invention connection double-core copper (II) complex solution (10 μ L/ times), with sepectrophotofluorometer come
Record each launching light spectrogram (Fig. 2).Excitation wavelength is 526nm.Binding constants are calculated using formula 2：
KEB[EB]=Kapp[complex] (2)
Wherein KEB=1.0 × 107M-1, [EB]=3.78 × 10-6M, [complex] are that fluorescence intensity weakens to original
Complex ultimate density when 50%.Calculate the binding constants (K for acquiring complex 1 and DNAapp) be respectively 1.71 ×
105M-1, between 104~105M-1, show that the interaction of the complex and DNA are groove binding.
Embodiment 6
The present embodiment utilizes circular dichroism experimental study phenoxy bridge connection double-core copper (II) complex of the present invention and DNA
In conjunction with the influence to DNA conformations.DNA (1.48 × 10 is recorded with circular dichroism instrument-4M CD spectrum) and [complex]/[DNA]
The CD spectrum that (0.2 and 0.4) is combined under different molar concentration ratios.As shown in figure 3, as phenoxy bridge of the present invention connection is double
The increase of the concentration of core copper (II) complex, the conformation of CT-DNA are changed.
Embodiment 7
PBR322 Plasmid DNA is used as the substrate of cutting experiment, and DNA cutting experiments are carried out by agarose gel electrophoresis.It will
PBR322 Plasmid DNA (0.02 μ g/ μ L) is mixed with 2 gained phenoxy bridge of a certain amount of embodiment connection double-core copper (II) complex, so
100 times ascorbic acid (Vc) initiation reaction that mole is complex is added afterwards, with 50mM Tris-HCl/50mM NaCl
(pH=7.40) buffer solution is settled to 10 μ L.Hatch 30min in 37 DEG C of water-baths, 1.5 μ L are added and terminate buffer (0.25% bromine
Phenol is blue, 25% glycerine, 1mM EDTA) terminate reaction, 1%EB dyeing, gel electrophoresis analysis result.Electrophoretic determination is in TAE (40mM
Tris acetate/1mM EDTA) it carries out in buffer solution, it is powered 1.5 hours under 60V voltages, with UV gel imaging systems to electricity
Swimming figure is taken pictures, to observe cutting situation of the complex to DNA.As shown in figure 4, phenoxy bridge of the present invention joins double-core copper
(II) pBR322 plasmid super coiled DNAs can be cut into the presence of Vc and incise type DNA (Form II) and linear DNA by complex
(Form III)。
Embodiment 8
Join double-core copper (II) complex to the cutting mechanism of DNA to study phenoxy bridge of the present invention, we are in system
Middle addition different activities oxygen killer.To the 2 gained phenol oxygen of embodiment of pBR322 Plasmid DNA (0.02 μ g/ μ L) and respective concentration
DMSO (1M), KI (10mM), NaN are separately added into bridged binuclear copper (II) complex mixing sample3(10mM), D2O (70%) and
The tert-butyl alcohol (1M) hatches 10min in 37 DEG C of water-baths, and 100 times of Vc (1 μ L) initiation reactions are then added, continue to incubate in 37 DEG C of water-baths
Reaction is terminated after changing 30min, subsequent step is carried out according to the step of DNA cutting experiments.As shown in figure 5, DMSO and the tert-butyl alcohol (hydroxyl
Base free radical scavenger), KI (H2O2Agent for capturing), NaN3(singlet oxygen agent for capturing) does not all have the cutting of pBR322 Plasmid DNA
Apparent inhibiting effect.However D2O (singlet oxygen accelerating agent) significantly improves cleavage activity.The experimental results showed that OH,
H2O2, and1O2It is not the active specy in cleavage reaction, it would be possible that is played a major role is the object that copper ion is formed with oxygen
Kind.
The above is merely for convenience of it will be understood by those skilled in the art that technical scheme of the present invention, not limiting
The present invention.All within the spirits and principles of the present invention, any modification, equivalent replacement, improvement and so on should be included in this
Within the protection domain of invention.
Claims (10)
1. a kind of phenoxy bridge joins double-core copper (II) complex, which is characterized in that described phenoxy bridge connection double-core copper (II) complex
Chemical formula is [Cu2(BHPMP)] Cl, wherein [Cu2(BHPMP)]+Structural formula it is as follows：
2. the preparation method of phenoxy bridge connection double-core copper (II) complex described in claim 1, which is characterized in that
Make CuCl2And H3BHPMP carries out complex reaction in acetonitrile solvent, in 40~60 DEG C, and [Cu is obtained after purification2(BHPMP)]
Cl solids, wherein H3The structural formula of BHPMP is as follows：
3. preparation method as claimed in claim 2, which is characterized in that
H3The preparation method of BHPMP is：(2- hydroxybenzyls) (2- pyridylmethyls) amine is dissolved with tetrahydrofuran, ice-water bath is protected
Under shield, pH value of solution is adjusted to 7~8 with triethylamine, the tetrahydrofuran of 2,6- dichloromethyl -4- methylphenols is then added dropwise thereto
Solution after being added dropwise, is stirred overnight, commonly filters to get filtrate at room temperature, and vacuum distillation removes solvent and obtains yellow oil, uses
Water, ethyl alcohol, ether wash successively, are dried in vacuo to obtain yellow solid, as H3BHPMP ligands.
4. preparation method as claimed in claim 3, which is characterized in that (2- hydroxybenzyls) (2- pyridylmethyls) amine with
The molar ratio of the 2,6- dichloromethyls -4- methylphenols is (2.2~2.0):1.
5. preparation method as claimed in claim 2, which is characterized in that
The specific method of the purification is that reaction mixture is filtered under diminished pressure distillation and concentration, ether is added, it is heavy that bottle green is precipitated
It forms sediment, centrifugation obtains solid, and [Cu is obtained after washing, vacuum drying2(BHPMP)] Cl solids.
6. preparation method as claimed in claim 5, which is characterized in that
The washing is that sequentially acetone, ether wash the solid that centrifugation obtains.
7. preparation method as claimed in claim 2, which is characterized in that
By CuCl2·2H2O and H3BHPMP ligands press 2:1 ratio mixing, is dissolved in acetonitrile solvent, 40~60 DEG C of heating stirrings 4
~6h, purification is to get to [Cu after stopping reaction2(BHPMP)] Cl solids.
8. phenoxy bridge described in claim 1 joins double-core copper (II) complex application in preparation of anti-tumor drugs.
9. application as claimed in claim 8, which is characterized in that the tumour is human cervical carcinoma, non-small cell type lung cancer and people
Breast cancer.
10. application of phenoxy bridge connection double-core copper (II) complex as DNA cutting reagents described in claim 1.
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| CN113024449B (en) * | 2021-03-18 | 2023-11-28 | 中国科学技术大学 | Metal-containing high-efficiency cationic antitumor drug and preparation method and application thereof |
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