CN105963715A - Double-polypeptide modified europium doped gadolinium oxide nanorod and preparation thereof - Google Patents
Double-polypeptide modified europium doped gadolinium oxide nanorod and preparation thereof Download PDFInfo
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Abstract
The invention relates to a preparation method of a double-polypeptide modified europium doped gadolinium oxide nanorod for in vivo target reinforcement of magnetic resonance-fluorescence dual-mode imaging and restraining of growth of early-stage brain glioma. The preparation method comprises the following steps: dissolving gadolinium nitrate and europium nitrate, conducting reaction on the dissolved gadolinium nitrate and europium nitrate and strong base, and obtaining a europium doped gadolinium hydroxide nanorod through heating growth; sintering at a high temperature to obtain a europium doped gadolinium oxide nanorod; then, through surface hydroxylation, amination and pegylation, connecting an RGD polypeptide and CTX through the sulfhydrylation technology. According to the invention, the technology is simple, the condition is controllable, the operation is easy, the nanorod product is small in size (the nanorod is about 80 nanometers in length and about 25 nanometers in diameter), and low in draw ratio; the nanorod modified by two polypeptides, namely RGD and CTX, can penetrate through double barriers, namely blood brain and hemangioma, of a brain tumor, has excellent nuclear magnetism T1 mode relaxation efficiency, fluorescence characteristic, and early-stage glioma target and restraining characteristics and can be used for clinical diagnosis and treatment of early-stage brain glioma.
Description
Technical field
The present invention relates to technical field of nano material, the preparation method of a kind of double peptide modified europium doping Gadolinia. nanometer rods strengthening magnetic resonance-fluorescent dual module formula imaging and suppression cerebral glioma growth in early days for internal targeting.
Background technology
Cerebral glioma is modal primary pernicious intracranial tumor, and sickness rate is high, postoperative easy recurrence.The most clinical therapeutic modality to cerebral glioma mostly is surgical removal, radiation treatment and chemotherapy etc..But, cerebral glioma be accurately positioned, treat in time, reduce the side effect such as the injury to normal cerebral tissue, to extend survival time after surgical operation be to process the great difficult problem that in early days cerebral glioma faces.Therefore, a kind of medical material that can efficiently diagnose and can effectively treat in early days cerebral glioma is developed significant to clinical cerebroma diagnosis and treatment.At present, conventional clinically diagnosing tumor mode has ultrasonic contrast, x-ray imaging, Computed tomography (CT), Positron emission computed tomography imaging (PET), radionuclide imaging, optical imagery, nuclear magnetic resonance etc..But, single-mode imaging is because of its intrinsic image defects, and can not meet the clinical detection to complicated early stage physical pathological condition；Multi-modal imaging can integrate multiple imaging advantage so that it is mutually learns from other's strong points to offset one's weaknesses, and can detect, for clinical disease, the physiology Pathological Information providing more rich, and magnetic resonance-fluorescent dual module formula imaging is exactly one of them.
At present, the material that can be used as strengthening nuclear magnetic resonance has a lot, such as ferroso-ferric oxide, gamma-iron sesquioxide, the chelate of gadolinium and gadolinio nano-particle.But, ferroso-ferric oxide and gamma-iron sesquioxide are T2Pattern contrast agent, Contrast enhanced efficiency is low and consumption is big；And the chelate of gadolinium is the most easily removed by vascular endothelial cell reticular system, blood circulation time is short, and radiography efficiency is the lowest, repeatedly and a large amount of use brings again toxicity in vivo to increase.Europium doping Gadolinia. nanometer rods is the gadolinio inorganic nanoparticles of cubic structure, has that preparation condition is controlled, particle size is adjustable, stable performance, good dispersion, radiography efficiency is high and advantage that consumption is few.Especially, this nanometer rods can carry out function of surface peptide modification, so that it presents bigger application potential at biomedical sectors such as bio-imaging, cancer target, treatments of cancer.
Preparation Eu-Gd2O3The method of nano material has many, common one-step method has sol-gal process (CN103539192A), solid sintering technology (CN104926304A), microwave combusion method (CN104973615A) etc., and two-step method is totally divided into preparation and the preparation of later stage europium doping Gadolinia. nanometer rods of europium doped with hydrogen Gadolinia. presoma.The preparation of europium doped with hydrogen Gadolinia. nanometer rods presoma has coprecipitation, sol-gal process, hydro-thermal method etc., later stage europium doping Gadolinia. nanometer rods typically all high-temperature sintering process to prepare.Publication No. CN103638532B, CN103539192B and CN104857532A patent in, Xu Qunyuan and Zhang Huajuan etc. reports the oxidation gadolinio contrast agent preparation method that can be used for nuclear magnetic resonance and magnetic resonance-fluorescent dual module formula imaging respectively.But the preparation technology of these one-step method is too simple, the material thing obtained is mutually uneven, particle size distribution width, impure more；Time prepared by two-step method, europium doped with hydrogen Gadolinia. granular precursor size prepared by coprecipitation and sol-gal process is too big, and crystal property is poor, between europium doping Gadolinia. nanometer rods granule prepared during high-temperature calcination, serious fusion, its bad dispersibility easily occurs.Owing to the cerebral tumor exists blood brain barrier and the dual obstruction of bloodtumorbarrier, and cerebral tumor size is little in early days and presents netted scattered feature, does not therefore also relate to function of surface peptide at present and modifies and energy targeting cerebral tumor Imaging enhanced nano material.Therefore, design that a kind of size is little, good dispersion, can pass through blood brain and angioma double screen barrier multi-functional europium doping Gadolinia. nano-bar preparation method for material the most necessary, and it is the most indispensable that this small size europium doping Gadolinia. nano-bar material is carried out the modification of multifunction surface.
Polyethylene Glycol (PEG) is a kind of hydrophilic high molecular material with excellent biocompatibility, is one of the most frequently used biomaterial surface modifying agent；It can be greatly increased nano material blood circulation time in vivo after modifying, thus improves nano material cancer target probability.Arginine-glycine-aspartic acid sequence polypeptide (RGD), has and sticks with cell surface Laminin lens high affinity, the α to cerebral tumor surface high expressedvβ3Efficient targeting affinity characteristic is have Deng integration.TM-601 (CTX) is a kind of 36 peptide neurotoxins, but it is nontoxic to mammal and can pass blood brain barrier；And it combines metalloproteases-2 and presents selectively targeted combination the film of early stage brain glioblastoma cell surface high expressed, and this protein binding body can also suppress brain tumor growth because blocking the chloride channel of Tumor cells.Therefore RGD peptide and CTX peptide are simultaneously connected with in nano material, will can improve the cerebroma targeting efficiency of material and suppress the growth of cerebroma.
Summary of the invention
It is an object of the invention to provide the preparation method that technique is simple, react controlled, easily operated a kind of double peptide modified europium doping Gadolinia. nanometer rods that can be used for the enhancing magnetic resonance-fluorescent dual module formula imaging of internal targeting and suppression cerebral glioma in early days, with the problem solving to propose in above-mentioned background.
For achieving the above object, the present invention proposes following technical scheme:
The preparation method of a kind of double peptide modified europium doping Gadolinia. nanometer rods that can be used for the enhancing magnetic resonance-fluorescent dual module formula imaging of internal targeting and suppression cerebral glioma in early days, comprises the following steps:
A) gadolinium nitrate hexahydrate and six nitric hydrate europiums being dissolved in deionized water, form the gadolinium europium saline solution that concentration is 0.025 ~ 0.1M, wherein the doping of europium is 10 ~ 40mol%；Then being added dropwise over by the sodium hydrate aqueous solution that concentration is 0.5 ~ 2M, stirring makes its pH value reach 13, reacted after be changed into containing europium hydroxide and the mixed liquor A of Gadolinium trihydroxide white precipitate；
B) mixed liquor A is transferred in reactor, 100 ~ 250oHydro-thermal reaction 12h under C, is washed with deionized after cooling, is vacuum dried and obtains sample, thus europium hydroxide described in step a) and Gadolinium trihydroxide white precipitate nucleating growth under heating are europium doped with hydrogen Gadolinia. nanometer rods；
C) by dry europium doped with hydrogen Gadolinia. nanometer rods in grinding in ball grinder 0.5-6h, then at 300 ~ 900oAfter sintering 3 ~ 12h under C, obtain europium doping Gadolinia. nanometer rods；
D) by 130 ~ 240mg europium doping Gadolinia. nanometer rods ultrasonic disperse 120 ~ 150mL, 2.0 ~ 15wt% tetramethyl ammonium hydroxide solution B in, 40 ~ 250oC reacts 2 ~ 24h, uses deionized water centrifuge washing three times after cooling, obtains surface hydroxylation europium doping Gadolinia. nanometer rods；
E) surface hydroxylation europium doping Gadolinia. nanometer rods is dispersed in the mixed organic solvents C of 100mL dimethyl sulfoxide and carbon tetrachloride, in mixed liquor C, dimethyl sulfoxide is 1:1 ~ 10 with the volume ratio of carbon tetrachloride, then being sequentially added into 1 ~ 3mL aminopropyl triethoxysilane and 30 ~ 90 L deionized waters, gained mixed liquor D is 40 ~ 120oCooling, centrifugation, washing after magnetic agitation reaction 2 ~ 24h under C, the dried europium doping Gadolinia. nanometer rods obtaining surface amination；
F) by 40 ~ 60mg surface amination europium doping Gadolinia. nanometer rods ultrasonic disperse in 10mL dimethyl sulfoxide, it is subsequently adding the Mal-PEG-NHS of difunctionalization, the Mal-PEG-NHS of difunctionalization is the Polyethylene Glycol that two ends are modified by maleimide and butanimide, after gained mixed liquor E at room temperature magnetic agitation, it is centrifuged separating, washing, it is dried, obtains the europium doping Gadolinia. nanometer rods of PEG cladding；
G) PEG of 15 ~ 25mg is coated with europium doping Gadolinia. nanometer rods ultrasonic disperse in 3mL dimethyl sulfoxide, it is subsequently adding the ring-type arginyl ~ glycyl ~ aspartic acid small peptide of sulfhydrylation, i.e. HS-cRGD, under gained mixed liquor F room temperature after magnetic agitation, it is centrifuged separating, washs, is dried, obtain the europium doping Gadolinia. nanometer rods that RGD modifies；
H) the europium doping Gadolinia. nanometer rods ultrasonic disperse modified by the RGD of 15 ~ 25mg is in 3mL dimethyl sulfoxide, it is subsequently adding the TM-601 of sulfhydrylation, i.e. HS-CTX, under gained mixed liquor G room temperature after magnetic agitation, it is centrifuged separating, washs, is dried, obtain the double peptide modified europium doping Gadolinia. nanometer rods of end product RGD and CTX.
As the further scheme of the present invention, in step f), the addition of described difunctional Mal-PEG-NHS is 10 ~ 20mg, and under gained mixed liquor E room temperature, mixing time is 12 ~ 48h.
As the further scheme of the present invention, in step g), the addition of the ring-type arginyl-glycyl-aspartic acid small peptide of described sulfhydrylation is 1 ~ 10mg, and the gained mixed liquor F magnetic agitation time at room temperature is 12 ~ 48h.
As the further scheme of the present invention, in step h), adding of the TM-601 of described sulfhydrylation is 1 ~ 10mg, and the gained mixed liquor G magnetic agitation time at room temperature is 12 ~ 48h.
In the method for the invention, for obtaining pure separation product, step e), f), g), h) in need to carry out separating and washing operation.Can separate by any suitable method, preferably centrifugation, centrifugal speed is preferably 5000 ~ 8000 revs/min, and centrifugation time is preferably 5 ~ 30min.Reagent for washing is preferably dimethyl sulfoxide, secondary water and ethanol.For obtaining purified product, centrifugal and washing times is respectively 6 times, washs for dimethyl sulfoxide for twice, washs for secondary water for twice, and last twice is washing with alcohol.Nano-particle after isolated and purified can store with hygrometric state or dry state.When dry state stores, need to dry separation product at moderate temperatures, dry temperature and be preferably-20 ~-50oC。
Compared with present technology, the beneficial effects of the present invention is:
The present invention prepares gained double peptide modified europium doping Gadolinia. nanometer rods and can be used for the enhancing magnetic resonance-fluorescent dual module formula imaging of internal targeting and suppression cerebral glioma in early days.Present invention process is simple, reaction condition is gentle, easily operated, prepared double peptide modified europium doping Gadolinia. nanometer rods not only size is little, and (rod is about 80 nanometers, diameter about 25 nanometer), draw ratio is low, good dispersion, blood brain and the angioma double barrier of cerebroma can be passed through after RGD and CTX two peptide species is modified, and there is good T1Relaxation effect, fluorescence imaging ability and cerebral glioma target function, moreover it is possible to the effectively growth of suppression cerebral glioma in early days, the diagnosis and treatment field of cerebral glioma has potential clinical value in early days.
Accompanying drawing explanation
Fig. 1 is that calcining obtains Eu-Gd2O3The facies analysis of nanometer rods and morphology observation figure.
Fig. 2 is Eu-Gd2O3Transmission electron microscope picture before and after nanometer rods cladding PEG.
Fig. 3 is to Eu-Gd2O3Nanometer rods carries out surface PEG cladding and peptide modified illustrative steps.
Detailed description of the invention
Below in conjunction with the embodiment of the present invention, the technical scheme in the embodiment of the present invention is clearly and completely described, it is clear that described embodiment is only a part of embodiment of the present invention rather than whole embodiments.Based on the embodiment in the present invention, the every other embodiment that those of ordinary skill in the art are obtained under not making creative work premise, broadly fall into the scope of protection of the invention.
Europium doping Gadolinia. nanometer rods that what the present invention provided prepare and the method route that surface is modified due to equipment, reagent and experimental situation etc. without rigors, be thus susceptible to be amplified experiment and easy volume production.The following is the exemplary specific embodiments of the present invention, the above and other advantage of the present invention can be more fully understood by by these embodiments.
Embodiment
1
Weigh 1.806g gadolinium nitrate hexahydrate, 0.446g six nitric hydrate europium is dissolved in 100mL deionized water, is the most under agitation added dropwise over 1M sodium hydroxide solution, and monitors pH of mixed change, when pH value reaches 13, stops dropping sodium hydroxide solution.Continue stirring 5min, reacted after must contain Gadolinium trihydroxide and the mixed liquor A of europium hydroxide white precipitate.Mixed liquor A is transferred in 200mL reactor, is placed in 180oReact 12h under the conditions of C, after cooling, gained precipitate is washed with deionized three times, after vacuum drying, obtains europium doped with hydrogen Gadolinia. presoma.By europium doped with hydrogen Gadolinia. presoma after grinding in ball grinder 2h is disperseed, 400oC air sinters 8h, after cooling, obtains europium doping Gadolinia. nanometer rods (i.e. Eu-Gd2O3
NRs).By the Eu-Gd of 200mg2O3Nanometer rods ultrasonic disperse in the tetramethyl ammonium hydroxide solution B of 120mL, 7.0wt.%, 180oC reacts 6h, is washed with deionized centrifugal three times, obtains surface hydroxylation Eu-Gd after cooling2O3Nanometer rods；By 200mg hydroxylating Eu-Gd2O3Nanometer rods ultrasonic disperse, in the mixed organic solvents C of 100mL dimethyl sulfoxide and carbon tetrachloride, is then sequentially added into 3mL aminopropyl triethoxysilane and 90 L deionized waters, and gained mixed liquor D is 70oCooling, centrifugation, washing three times after magnetic agitation reaction 12h, the dried Eu-Gd obtaining surface amination under C2O3Nanometer rods；By 50mg surface amination Eu-Gd2O3Nanometer rods ultrasonic disperse, in 10mL dimethyl sulfoxide, is subsequently adding the Mal-PEG-NHS of 20mg difunctionalization, centrifugation after gained mixed liquor E at room temperature magnetic agitation, washing three times, the dried europium doping Gadolinia. nanometer rods obtaining PEG cladding；The PEG of 20mg is coated with europium doping Gadolinia. nanometer rods ultrasonic disperse in 3mL dimethyl sulfoxide, it is subsequently adding the ring-type RGD(i.e. HS-cRGD of 5mg sulfhydrylation), centrifugation after magnetic agitation under gained mixed liquor F room temperature, washing three times, the europium doping Gadolinia. nanometer rods that the dried RGD of obtaining modifies；The Eu-Gd that the RGD of 20mg is modified2O3Nanometer rods ultrasonic disperse, in 3mL dimethyl sulfoxide, is subsequently adding the TM-601 (i.e. HS-CTX) of 5mg sulfhydrylation, centrifugation after magnetic agitation under gained mixed liquor G room temperature, washing three times, the double peptide modified Eu-Gd of dried end product RGD and CTX2O3Nanometer rods (i.e. RGD-Eu-Gd2O3-CTX
NRs).Need to be centrifuged separating and washing operation during nanorod surfaces amination, PEG cladding, RGD and CTX are modified.In centrifugal process, centrifugal speed is 8000 revs/min, and centrifugation time is 10min.In washing process, the reagent for washing is respectively dimethyl sulfoxide, secondary water and ethanol.For obtaining purified product, centrifugal and washing times is respectively 6 times, washs for dimethyl sulfoxide for twice, washs for secondary water for twice, and last secondary is washing with alcohol.Nano-particle after isolated and purified can store with hygrometric state or dry state.When dry state stores, need to separate product vacuum drying in a low temperature of-20 ~-50 DEG C and store after 24 hours.
Embodiment
2
Precise 1.355 g gadolinium nitrate hexahydrate, 0.893 g six nitric hydrate europium are dissolved in 100 mL deionized waters, the most under agitation it is added dropwise over 2M sodium hydroxide solution, and monitor pH of mixed change, when pH value reaches 13, stop dropping sodium hydroxide solution.Continue stirring 5min, reacted after must contain Gadolinium trihydroxide and the mixed liquor A of europium hydroxide white depositions.Mixed liquor A is transferred in 200mL reactor, is placed in 200oReact 12h under the conditions of C, after cooling, gained precipitate is washed with deionized three times, after vacuum drying, obtains europium doped with hydrogen Gadolinia. presoma.After europium doped with hydrogen Gadolinia. presoma is ground 3h dispersion, 600oC air atmosphere sinters 6h, after cooling, obtains europium doping Gadolinia. nanometer rods (i.e. Eu-Gd2O3
NRs).By 180mg Eu-Gd2O3Nanometer rods ultrasonic disperse in the tetramethyl ammonium hydroxide solution B of 120mL, 7.0wt.%, 200oC reacts 6h, uses deionized water centrifuge washing three times, obtain surface hydroxylation Eu-Gd after cooling2O3Nanometer rods；By 180mg hydroxylating Eu-Gd2O3Nanometer rods ultrasonic disperse, in the mixed organic solvents C of 100mL dimethyl sulfoxide and carbon tetrachloride, is then sequentially added into 1mL aminopropyl triethoxysilane and 30 L deionized waters, and gained mixed liquor D is 90oCooling, centrifugation, washing three times after magnetic agitation reaction 12h, the dried Eu-Gd obtaining surface amination under C2O3Nanometer rods；By 60mg surface amination Eu-Gd2O3Nanometer rods ultrasonic disperse, in 10mL dimethyl sulfoxide, is subsequently adding the Mal-PEG-NHS of 10mg difunctionalization, centrifugation after gained mixed liquor E at room temperature magnetic agitation, washing three times, the dried europium doping Gadolinia. nanometer rods obtaining PEG cladding；The PEG of 22mg is coated with europium doping Gadolinia. nanometer rods ultrasonic disperse in 3mL dimethyl sulfoxide, it is subsequently adding the ring-type RGD(i.e. HS-cRGD of 6mg sulfhydrylation), centrifugation after magnetic agitation under gained mixed liquor F room temperature, washing three times, the europium doping Gadolinia. nanometer rods that the dried RGD of obtaining modifies；The Eu-Gd that the RGD of 22mg is modified2O3Nanometer rods ultrasonic disperse, in 3mL dimethyl sulfoxide, is subsequently adding the TM-601 (i.e. HS-CTX) of 6mg sulfhydrylation, centrifugation after magnetic agitation under gained mixed liquor G room temperature, washing three times, the double peptide modified Eu-Gd of dried end product RGD and CTX2O3Nanometer rods (i.e. RGD-Eu-Gd2O3-CTX
NRs).Need to be centrifuged separating and washing operation during nanorod surfaces amination, PEG cladding, RGD and CTX are modified.In centrifugal process, centrifugal speed is 6000 revs/min, and centrifugation time is 8min.In washing process, the reagent for washing is respectively dimethyl sulfoxide, secondary water and ethanol.For obtaining purified product, centrifugal and washing times is respectively 6 times, washs for dimethyl sulfoxide for twice, washs for secondary water for twice, and last secondary is washing with alcohol.Nano-particle after isolated and purified can store with hygrometric state or dry state.When dry state stores, need to separate product vacuum drying in a low temperature of-20 ~-50 DEG C and store after 24 hours.
Embodiment
3
Precise 1.806g gadolinium nitrate hexahydrate, 0.446g six nitric hydrate europium are dissolved in 100mL deionized water, are the most under agitation added dropwise over 2M sodium hydroxide solution, and monitor pH of mixed change, when pH value reaches 13, stop dropping sodium hydroxide solution.Continue stirring 5min, reacted after must contain Gadolinium trihydroxide and the mixed liquor A of europium hydroxide white depositions.Mixed liquor A is transferred in 200mL reactor, is placed in 250oReact 12h under the conditions of C, after cooling, gained precipitate is washed with deionized three times, after vacuum drying, obtains europium doped with hydrogen Gadolinia. presoma.By europium doped with hydrogen Gadolinia. presoma after grinding in ball grinder 3h is disperseed, 800oC air atmosphere sinters 8h, after cooling, obtains europium doping Gadolinia. nanometer rods (i.e. Eu-Gd2O3
NRs).By 220mg Eu-Gd2O3Nanometer rods ultrasonic disperse in the tetramethyl ammonium hydroxide solution B of 150mL, 7.0wt.%, 250oC reacts 6h, uses deionized water centrifuge washing three times, obtain surface hydroxylation Eu-Gd after cooling2O3Nanometer rods；By 220mg hydroxylating Eu-Gd2O3Nanometer rods ultrasonic disperse, in the mixed organic solvents C of 100mL dimethyl sulfoxide and carbon tetrachloride, is then sequentially added into 2mL aminopropyl triethoxysilane and 60 L deionized waters, and gained mixed liquor D is 90oCooling, centrifugation, washing three times after magnetic agitation reaction 12h, the dried Eu-Gd obtaining surface amination under C2O3Nanometer rods；By 60mg surface amination Eu-Gd2O3Nanometer rods ultrasonic disperse, in 10mL dimethyl sulfoxide, is subsequently adding the Mal-PEG-NHS of 10mg difunctionalization, centrifugation after gained mixed liquor E at room temperature magnetic agitation, washing three times, the dried europium doping Gadolinia. nanometer rods obtaining PEG cladding；The PEG of 25mg is coated with europium doping Gadolinia. nanometer rods ultrasonic disperse in 3mL dimethyl sulfoxide, it is subsequently adding the ring-type RGD(i.e. HS-cRGD of 8mg sulfhydrylation), centrifugation after magnetic agitation under gained mixed liquor F room temperature, washing three times, the europium doping Gadolinia. nanometer rods that the dried RGD of obtaining modifies；The Eu-Gd that the RGD of 25mg is modified2O3Nanometer rods ultrasonic disperse, in 3mL dimethyl sulfoxide, is subsequently adding the TM-601 (i.e. HS-CTX) of 8mg sulfhydrylation, centrifugation after magnetic agitation under gained mixed liquor G room temperature, washing three times, the double peptide modified Eu-Gd of dried end product RGD and CTX2O3Nanometer rods (i.e. RGD-Eu-Gd2O3-CTX
NRs).Need to be centrifuged separating and washing operation during nanorod surfaces amination, PEG cladding, RGD and CTX are modified.In centrifugal process, centrifugal speed is 5000 revs/min, and centrifugation time is 5min.In washing process, the reagent for washing is respectively dimethyl sulfoxide, secondary water and ethanol.For obtaining purified product, centrifugal and washing times is respectively 6 times, washs for dimethyl sulfoxide for twice, washs for secondary water for twice, and last secondary is washing with alcohol.Nano-particle after isolated and purified can store with hygrometric state or dry state.When dry state stores, need to separate product vacuum drying in a low temperature of-20 ~-50 DEG C and store after 24 hours.
The europium doping Gadolinia. nanometer rods that in the present invention, double peptides are modified is to be prepared by multistep processes: continues hydro-thermal reaction under room temperature after co-precipitation and obtains europium doped with hydrogen Gadolinia. nanometer rods presoma；Then calcining presoma obtains europium doping Gadolinia. nanometer rods；Take some reaction treatment arranged that it is carried out surface modification subsequently, the double peptide modified europium doping Gadolinia. nanometer rods of final acquisition RGD and CTX.Fig. 1 is the scanning electron microscope (SEM) photograph that calcining obtains europium doping Gadolinia. nanometer rods, gained europium doping Gadolinia. nanometer rods seen from this figure be the nanometer rods of fine dispersion, and its length is about 80 nanometers, and diameter is about 25 nanometers.Fig. 2 is the transmission electron microscope picture before and after europium doping Gadolinia. nanometer rods cladding PEG, it can be seen that europium doping Gadolinia. is received in fine dispersion nano bar-shape structure, PEG is uniformly coated on nanorod surfaces.Fig. 3 is that europium doping Gadolinia. nanometer rods carries out surface PEG cladding and peptide modified illustrative steps, as can be seen from Figure europium doping Gadolinia. nanorod surfaces modification strict logic, scientific and reasonable, simple to operate, environmental protection.
It is obvious to a person skilled in the art that the invention is not restricted to the details of above-mentioned one exemplary embodiment, and without departing from the spirit or essential characteristics of the present invention, it is possible to realize the present invention in other specific forms.Therefore, no matter from the point of view of which point, embodiment all should be regarded as exemplary, and be nonrestrictive, the scope of the present invention is limited by claims rather than described above, it is intended that all changes fallen in the implication of equivalency and scope of claim included in the present invention.
Claims (10)
1. one kind can be used for the preparation method that internal targeting strengthens the double peptide modified europium doping Gadolinia. nanometer rods of magnetic resonance-fluorescent dual module formula imaging and suppression cerebral glioma in early days, it is characterized in that comprising the steps of and a) gadolinium nitrate hexahydrate and six nitric hydrate europiums are dissolved in deionized water, forming the gadolinium europium saline solution that concentration is 0.025 ~ 0.1M, wherein the doping of europium is 10 ~ 40mol%；Then being added dropwise over by the sodium hydrate aqueous solution that concentration is 0.5 ~ 2M, stirring makes its pH value reach 13, reacted after be changed into containing europium hydroxide and the mixed liquor A of Gadolinium trihydroxide white precipitate；B) mixed liquor A is transferred in reactor, 100 ~ 250oHydro-thermal reaction 12h under C, is washed with deionized after cooling, is vacuum dried and obtains sample, thus europium hydroxide described in step a) and Gadolinium trihydroxide white precipitate nucleating growth under heating are europium doped with hydrogen Gadolinia. nanometer rods；C), after europium doped with hydrogen Gadolinia. nanometer rods drying and grinding being disperseed, after sintering certain time in high temperature furnace, europium doping Gadolinia. nanometer rods is obtained；D) the Gadolinia. nanometer rods ultrasonic disperse that europium adulterated in tetramethyl ammonium hydroxide solution B, temperature reaction certain time, centrifugal with deionized water after cooling, washing three times, obtain surface hydroxylation europium doping Gadolinia. nanometer rods；E) by hydroxylating europium doping Gadolinia. nanometer rods ultrasonic disperse in the mixed organic solvents C of 100mL dimethyl sulfoxide and carbon tetrachloride, then 1 ~ 3mL aminopropyl triethoxysilane and 30 ~ 90 L deionized waters it are sequentially added into, cooling, centrifugation, washing after the reaction of gained mixed liquor D magnetic agitation under heating, the dried europium doping Gadolinia. nanometer rods obtaining surface amination；F) europium of surface amination is adulterated Gadolinia. nanometer rods ultrasonic disperse in 10mL dimethyl sulfoxide, it is subsequently adding the Polyethylene Glycol (Mal-PEG-NHS) that two ends are modified by maleimide and butanimide respectively, the i.e. Polyethylene Glycol of difunctionalization, centrifugation, washing after gained mixed liquor E at room temperature magnetic agitation, the dried europium doping Gadolinia. nanometer rods obtaining PEG cladding；G) the europium doping Gadolinia. nanometer rods ultrasonic disperse being coated with by PEG is in 3mL dimethyl sulfoxide, it is subsequently adding the ring-type arginyl-glycyl-aspartic acid small peptide of sulfhydrylation, i.e. HS-cRGD, under gained mixed liquor F room temperature after magnetic agitation, it is centrifuged separation, washing, the dried europium doping Gadolinia. nanometer rods obtaining RGD modification；H) the europium doping Gadolinia. nanometer rods ultrasonic disperse modified by RGD is in 3mL dimethyl sulfoxide, it is subsequently adding the TM-601 of sulfhydrylation, i.e. HS-CTX, after gained mixed liquor G at room temperature magnetic agitation certain time, it is centrifuged, separates, washs, obtains the double peptide modified europium doping Gadolinia. nanometer rods of end product RGD and CTX after drying.
Can be used for the preparation method that internal targeting strengthens the double peptide modified europium doping Gadolinia. nanometer rods of magnetic resonance-fluorescent dual module formula imaging and suppression cerebral glioma in early days the most as claimed in claim 1, it is characterized in that, in step c), by dry europium doped with hydrogen Gadolinia. nanometer rods in grinding in ball grinder 0.5-6h, then at 300 ~ 900o3 ~ 12h is sintered under C.
Can be used for the preparation method that internal targeting strengthens the double peptide modified europium doping Gadolinia. nanometer rods of magnetic resonance-fluorescent dual module formula imaging and suppression cerebral glioma in early days the most as claimed in claim 1, it is characterized in that, in step d), by 130 ~ 240mg europium doping Gadolinia. nanometer rods ultrasonic disperse 120 ~ 150mL, 2.0 ~ 15wt% tetramethyl ammonium hydroxide solution B in, 40 ~ 250oC reacts 2 ~ 24 h.
Can be used for the preparation method that internal targeting strengthens the double peptide modified europium doping Gadolinia. nanometer rods of magnetic resonance-fluorescent dual module formula imaging and suppression cerebral glioma in early days the most as claimed in claim 1, it is characterized in that, in step e), by 130 ~ 240mg europium doping Gadolinia. nanometer rods ultrasonic disperse in the mixed liquor C of 100mL dimethyl sulfoxide and carbon tetrachloride, in 40 ~ 120oMagnetic agitation reaction 2 ~ 24h under C, wherein dimethyl sulfoxide is 1:1 ~ 10 with the volume ratio of carbon tetrachloride.
Can be used for the preparation method that internal targeting strengthens the double peptide modified europium doping Gadolinia. nanometer rods of magnetic resonance-fluorescent dual module formula imaging and suppression cerebral glioma in early days the most as claimed in claim 1, it is characterized in that, in step f), the addition of the europium doping Gadolinia. nanometer rods of described surface amination is 40 ~ 60mg, the addition of Mal-PEG-NHS is 10 ~ 20mg, and gained mixed liquor E mixing time at room temperature is 12 ~ 48h.
Can be used for the preparation method that internal targeting strengthens the double peptide modified europium doping Gadolinia. nanometer rods of magnetic resonance-fluorescent dual module formula imaging and suppression cerebral glioma in early days the most as claimed in claim 1, it is characterized in that, in step g), the addition of the europium doping Gadolinia. nanometer rods of described PEG cladding is 15 ~ 25mg, the addition of the ring-type arginyl-glycyl-aspartic acid small peptide of sulfhydrylation is 1 ~ 10mg, and under gained mixed liquor F room temperature, the magnetic agitation time is 12 ~ 48h.
Can be used for the preparation method that internal targeting strengthens the double peptide modified europium doping Gadolinia. nanometer rods of magnetic resonance-fluorescent dual module formula imaging and suppression cerebral glioma in early days the most as claimed in claim 1, it is characterized in that, in step h), the addition of the europium doping Gadolinia. nanometer rods that described RGD modifies is 15 ~ 25mg, the addition of the TM-601 of sulfhydrylation is 1 ~ 10mg, and the gained mixed liquor G magnetic agitation time at room temperature is 12 ~ 48h.
Can be used for the preparation method that internal targeting strengthens the double peptide modified europium doping Gadolinia. nanometer rods of magnetic resonance-fluorescent dual module formula imaging and suppression cerebral glioma in early days the most as claimed in claim 1, it is characterized in that, step e), f), g), h) in, the centrifugal rotational speed of centrifugation is 5000 ~ 8000 revs/min, and centrifugation time is 5 ~ 30min.
Can be used for the preparation method that internal targeting strengthens the double peptide modified europium doping Gadolinia. nanometer rods of magnetic resonance-fluorescent dual module formula imaging and suppression cerebral glioma in early days the most as claimed in claim 1, it is characterized in that, step e), f), g), h) in, reagent for washing is dimethyl sulfoxide, secondary water and ethanol, every kind of reagent washes twice, altogether washing 6 times.
Can be used for the preparation method that internal targeting strengthens the double peptide modified europium doping Gadolinia. nanometer rods of magnetic resonance-fluorescent dual module formula imaging and suppression cerebral glioma in early days the most as claimed in claim 1, it is characterized in that, step e), f), g), h) in, baking temperature is-5 ~
-50oC。
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