CN110946999B - Diagnosis and treatment integrated nano platform, preparation method and application thereof - Google Patents

Diagnosis and treatment integrated nano platform, preparation method and application thereof Download PDF

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CN110946999B
CN110946999B CN201910898652.4A CN201910898652A CN110946999B CN 110946999 B CN110946999 B CN 110946999B CN 201910898652 A CN201910898652 A CN 201910898652A CN 110946999 B CN110946999 B CN 110946999B
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孙丽宁
张强
施利毅
王卓
孙松强
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University of Shanghai for Science and Technology
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Abstract

The invention discloses a diagnosis and treatment integrated nano platform and a preparation method thereof, wherein fullerene, doxorubicin hydrochloride and rare earth up-conversion luminescent nano crystal are taken as main bodies, intermolecular bonding is utilized, firstly, photosensitizer fullerene (C60) is modified on the surface of the rare earth up-conversion luminescent nano crystal, secondly, dithioglycolic acid is introduced to improve water solubility and simultaneously have a glutathione response release function, then, chemotherapy drug doxorubicin hydrochloride is connected, and the formed nano platform and folic acid modified beta-cyclodextrin are subjected to main-object self-assembly to enable the nano platform to have specific targeting and further improve the water solubility, so that the diagnosis and treatment integrated nano platform is obtained and can be used for optical imaging guided photodynamic and specific targeting drug release dual-mode treatment. The invention has good stability and strong targeting property, has two modes of cooperative treatment functions, carries out accurate fixed-point administration and meets the requirement of clinical diagnosis and treatment integration.

Description

Diagnosis and treatment integrated nano platform, preparation method and application thereof
Technical Field
The invention relates to the technical field of nano biomaterial manufacturing, in particular to a diagnosis and treatment integrated nano platform based on fullerene, doxorubicin hydrochloride and rare earth up-conversion luminescent nanocrystals as main bodies, and a preparation method and application thereof.
Background
Fullerene: the photosensitizer is zinc phthalocyanine, fullerene, rose bengal, methylene blue or chlorin.
Cancer has been one of the first killers of life threatening human health for over a century. This threat comes primarily from two areas: early cancer is not easy to diagnose and the clinical treatment means are limited. Due to the untimely diagnosis and treatment of early stage cancer, most of the patients receiving cancer treatment are middle and advanced stage patients. Currently, the main approaches for clinical treatment are chemotherapy and radiotherapy. However, both methods have certain disadvantages, for example, the drugs cannot enter cancer cells in a large amount accurately in the chemotherapy process, and the side effects of radiotherapy are large, which both result in the effect that the tumor cannot be completely cured. Therefore, the realization of a rapid and sensitive detection method and accurate and efficient treatment is urgent.
In recent years, fluorescence imaging has become a hot point of research in optical imaging because of the advantages of visualization of fluorescence imaging, no damage to cells and biological tissues, and the like. Among many fluorescent materials, rare earth doped up-conversion luminescent nano materials (UCNPs) have received much attention as a novel fluorescent material. This is because it can absorb low-energy infrared/near-infrared light (980nm) and emit high-energy visible light (red, green, blue, etc.), which has the advantages of eliminating background fluorescence interference, high signal-to-noise ratio, and deep penetration depth.
For cancer treatment, a series of novel nanomaterial-based therapeutic approaches such as photodynamic therapy (PDT), photothermal therapy (PTT), targeted drug therapy, etc. are currently developed in the laboratory. However, these nanomaterials have poor reproducibility, difficult structure control, poor water solubility, etc., which limits their biological applications. At present, C60 is taken as a photosensitizer capable of generating singlet oxygen, and has good biocompatibility; meanwhile, the characteristic of high Glutathione (GSH) content in a tumor microenvironment is utilized, and disulfide bond modified Dox is introduced to enable the Dox to have a specific response release function; the Folic Acid (FA) -modified beta-cyclodextrin (beta-CD) is assembled together on the up-conversion nanocrystal body to form the nanocomposite material, which has the advantages of low toxicity, targeted precise treatment, good dispersibility and the like, so that the nanocomposite material has great potential in cancer treatment.
In the prior art, Chinese invention patent 201410035675.X discloses aAn upconversion nanoparticle and gold nanorod composite nanoparticle, a preparation method and application thereof, belonging to the field of nano biomedicine. The method comprises the steps of reacting up-conversion core-shell nano particles with a photosensitizer and then reacting with SiO2Carrying out coupling reaction on the coated gold nanorods to obtain up-conversion nanoparticles and gold nanorod composite nanoparticles; the SiO2The thickness of the shell layer of the coated gold nanorod is 30-60 nm. According to the method, the photosensitizer is covalently or adsorbed on the surface of the up-conversion nanoparticles, so that the energy transfer efficiency is improved, the singlet oxygen yield is improved, and the photodynamic treatment effect is improved; and meanwhile, the action distance between the gold nanorod and the up-conversion nano particle is regulated and controlled, and the up-conversion fluorescence quenching phenomenon is avoided. The invention combines upconversion with photodynamic therapy with photosensitizers and SiO2The photo-thermal effects of the coated gold nanorods are combined to prepare a cooperative treatment nano platform, and one excitation light source achieves the purpose of two treatment means.
However, in the above patent technical scheme, two kinds of nanoparticles are adopted for compounding, the stability of the material is poor, and the material is suitable for a new photothermal compound therapy, and the reliability, stability and maturity of the material are yet to be verified; compared with the phototherapy and chemotherapy combined therapy, the nanoparticle photothermal therapy adopting gold nanorod combination has obvious differences in the reliability, stability and maturity of the curative effect.
Aiming at the problems of mutual independence of cancer diagnosis and treatment, long treatment period, high cost, single means, large toxic and side effects and the like in the current clinic and the problems of unstable material performance, unreliable curative effect and the like adopted by the existing photothermal therapy, in order to improve the treatment effect, reduce the toxic and side effects and the like, the invention applies and researches a nano particle as a main body, links organic small molecules through chemical bonds, has stronger stability and smaller overall size, is beneficial to treatment and discharge from the body, combines imaging and two modes of treatment, prepares a novel diagnosis and treatment integrated nano composite material with good stability and high curative effect reliability, and becomes a new direction of cancer treatment.
Therefore, the invention provides a diagnosis and treatment integrated nano platform taking fullerene, doxorubicin hydrochloride and rare earth up-conversion luminescent nanocrystals as main bodies, and the development of the diagnosis and treatment integrated nano platform is very necessary for applying the diagnosis and treatment integrated nano platform to phototherapy and chemotherapy compound therapy of cancer.
Disclosure of Invention
Aiming at the defects in the prior art, the invention provides a diagnosis and treatment integrated nano platform taking fullerene, doxorubicin hydrochloride and rare earth up-conversion luminescent nanocrystals as main bodies, a preparation method and application thereof, so as to realize the integration of diagnosis and treatment, namely the integration of fluorescence imaging, photodynamic therapy and precise chemotherapy.
In order to achieve the purpose, the invention adopts the technical scheme that:
a preparation method of a diagnosis and treatment integrated nano platform is characterized in that a rare earth up-conversion luminescent nano crystal is used as a main body, and a nano composite structure of fullerene and doxorubicin hydrochloride is bonded, and the preparation method comprises the following steps:
(1) dispersing oil-soluble rare earth up-conversion luminescence nanocrystals in cyclohexane, adding an acidic aqueous solution with the pH value of 3-5, reacting at room temperature for 4-5 hours to remove oleic acid ligands, modifying the oil-soluble rare earth up-conversion luminescence nanocrystals into water-soluble nanocrystals, centrifuging and washing, dispersing solids in water, adding citric acid into the solution, fully stirring at room temperature for 12 hours, centrifuging and washing to obtain citric acid modified water-soluble rare earth up-conversion luminescence nanocrystals, dispersing solids in water, adding amino-modified fullerene (C60), stirring while adding 1- (3-dimethylaminopropyl) -3-ethylcarbodiimide hydrochloride (EDC) and N-hydroxysuccinimide (NHS), stirring at room temperature for 12 hours, centrifuging and washing, dispersing solids in water to form a first dispersion liquid;
(2) dissolving dithioglycolic acid, doxorubicin hydrochloride (Dox), EDC and NHS in water, adding the first dispersion, stirring at room temperature for 12 hours, centrifuging, washing, and dispersing the solid in water to form a second dispersion;
(3) dissolving 3A-amino-3A-deoxy- (2AS,3AS) -beta-cyclodextrin hydrate, Folic Acid (FA), EDC and NHS in water, stirring at room temperature for 12 hours, adding the second dispersion, stirring at room temperature for 12 hours, centrifuging and washing to obtain the diagnosis and treatment integrated nano platform taking fullerene, doxorubicin hydrochloride and rare earth up-conversion luminescent nanocrystals AS main bodies.
In the step (1):
the mass ratio of the rare earth up-conversion luminescent nanocrystal to the citric acid is as follows: 10: (500-1000); dissolving C60 (100-200) and sodium hydroxide (NaOH) (3000-4000) in 200 parts of toluene, dissolving 50 parts of ethylenediamine in 100 parts of 75% ethanol solution, mixing the two solutions, and stirring at room temperature for 7-10 days; and finally, drying for 24 hours at 50-60 ℃, washing the product with 75% ethanol, and drying for 24 hours at 60-70 ℃ to obtain the amino modified C60. The mass part ratio of EDC to NHS is as follows: 50: (25-30).
In the step (2):
the mass part ratio of dithioglycollic acid to doxorubicin hydrochloride (Dox), EDC and NHS is as follows: (5-10): (5-10): 50: (25-30).
In the step (3):
the mass part ratio of the 3A-amino-3A-deoxy- (2AS,3AS) -beta-cyclodextrin hydrate to the Folic Acid (FA) to the EDC to the NHS is AS follows: (10-20): (5-10): 50: (25-30).
The oil-soluble up-conversion luminescence nanocrystal in the step (1) comprises: NaYF4:Yb,Tm、NaYF4:Yb,Er,Tm、NaYF4:Yb,Tm@NaYF4、NaYF4:Yb,Er,Tm@NaYF4、NaYF4:Yb,Tm@NaGdF4、NaYF4:Yb,Er,Tm@NaGdF4、NaYF4:Yb,Tm@NaYF4@NaGdF4、NaYF4:Yb,Er,Tm@NaYF4@NaGdF4And other oil-soluble up-conversion luminescent nanocrystals with similar properties and structures.
In the step (1), the water-soluble surface organic ligand comprises: polymaleic anhydride (HPMA), polymethacrylic acid (PMMA), polyacrylic acid (PAA).
The diagnosis and treatment integrated nano platform prepared by the method is characterized in that a rare earth up-conversion luminescent nano crystal is taken as a main body, a nano composite system of bonded fullerene and doxorubicin hydrochloride is taken, an oil-soluble rare earth up-conversion luminescent nano crystal is taken as a main body, organic functional micromolecules are assembled layer by layer on the surface of the nano crystal, and the organic micromolecules are linked through chemical bonds to form the nano composite system; the fullerene C60 has a spherical structure, wherein an amido bond on one surface of the fullerene C60 is combined with a carboxyl group on the surface of the rare earth up-conversion luminescent nanocrystal, and the other surface of the fullerene C60 is simultaneously bonded with doxorubicin hydrochloride modified by a disulfide bond; the double-sided bonding material further performs host-guest action with folic acid modified beta-cyclodextrin to obtain a diagnosis and treatment integrated nano platform taking fullerene, doxorubicin hydrochloride and rare earth up-conversion luminescent nanocrystals as a host, and the diagnosis and treatment integrated nano platform has good water solubility, biocompatibility and targeting property, strong material stability and small overall size, and is beneficial to treatment and in-vivo discharge.
The diagnosis and treatment integrated nano platform is characterized in that the diagnosis and treatment integrated nano platform is used as an auxiliary agent for photodynamic and drug dual-cooperative treatment guided by optical imaging diagnosis, so that diagnosis and treatment integration of clinical phototherapy and chemotherapy is realized.
The application of the diagnosis and treatment integrated nano platform is characterized in that the nano platform is used as an imaging agent for up-conversion fluorescence imaging.
The application of the diagnosis and treatment integrated nano platform is characterized in that the nano platform is used for accurate drug release of Glutathione (GSH) response.
The invention has the advantages that:
(1) the diagnosis and treatment integrated nano platform and the preparation method thereof provided by the invention are characterized in that a rare earth up-conversion luminescent nanocrystal is taken as a main body, a nano composite system of bonded fullerene and doxorubicin hydrochloride is taken, an oil-soluble rare earth up-conversion luminescent nanocrystal is taken as a main body, the surface of the nanocrystal is subjected to layer-by-layer organic functional micromolecule assembly, and the organic micromolecules are linked through chemical bonds to form the nano composite system, so that the material stability is strong, the whole size is small, and the treatment and the discharge from the body are facilitated; the preparation method has the advantages of few steps, high efficiency and easy industrialization;
(2) the invention adopts photosensitizer fullerene C60 of a nano platform, has a unique spherical structure, wherein an amido bond on one surface of the photosensitizer is combined with carboxyl on the surface of the rare earth up-conversion luminescent nanocrystal, and the other surface of the photosensitizer is simultaneously bonded with doxorubicin hydrochloride modified by a disulfide bond; the double-sided bonding material further performs host-guest action with folic acid modified beta-cyclodextrin to obtain a diagnosis and treatment integrated nano platform taking fullerene, doxorubicin hydrochloride and rare earth up-conversion luminescent nanocrystals as a host;
(3) according to the preparation method provided by the invention, organic micromolecules such as C60, cyclodextrin and folic acid are adopted, the preparation method of the nanocomposite based on the organic functional micromolecules and the up-conversion nanocrystalline layer-by-layer assembly is mild, the repeatability is high, and the prepared material has stable performance and good consistency; the prepared nano material system has good water solubility, biocompatibility and targeting property;
(4) the nano platform provided by the invention is based on the nano composite material assembled layer by layer of organic functional micromolecules and up-conversion nano crystals, and has the advantages of uniform size, good monodispersity and better biocompatibility.
(5) According to the nano platform and the application thereof provided by the invention, the division of each component in the system is clear, and each component has own function, and the components are mutually cooperated and do not interfere with each other; the nanocomposite material assembled layer by layer based on organic functional micromolecules and up-conversion nanocrystals has the characteristics of accurate drug release of fluorescence imaging, photodynamic therapy and GSH response, and can be applied to the fields of targeted diagnosis and targeted therapy.
(6) The nano platform and the application thereof provided by the invention are suitable for phototherapy and chemotherapy dual-synergistic therapy, are more mature and have more reliable curative effect compared with photothermal therapy suitable for gold nanorods, and particularly, the nano platform can be used for precise fixed-point administration, has small side effect on an organism, and meets various requirements in the fields of targeted diagnosis and targeted therapy of precise therapy.
The present invention will be described in further detail with reference to the accompanying drawings and specific embodiments.
Drawings
FIG. 1 shows the fullerene, doxorubicin hydrochloride and inert NaYF coated on the surface thereof obtained in accordance with embodiment 3 of the present invention4TEM photograph of diagnosis and treatment integrated nano material with rare earth thulium doped up-conversion luminescent nano crystal as main body;
FIG. 2 shows inert NaYF coated with C60 and surface coating prepared in example 6 of the present invention4Rare earth thulium doping of layersAn ultraviolet absorption spectrogram for measuring singlet oxygen of a nano material assembled by up-conversion nanocrystals;
FIG. 3 shows inert NaYF coated with C60 and surface coating prepared according to example 7 of the present invention4A fluorescence spectrum of a nanomaterial assembled by rare earth thulium doped up-conversion nanocrystals of the layer;
FIG. 4 shows inert NaYF coated with C60 and surface coating prepared according to example 8 of the present invention4The rare earth thulium-doped upconversion nanocrystalline self-assembled nanomaterial of the layer is further used for a potential diagram (Zeta) of an intermediate material in drug loading and access targeting processes.
Detailed Description
The technical solutions in the embodiments of the present invention will be clearly and completely described below with reference to the drawings in the embodiments of the present invention, and it is obvious that the described embodiments are only a part of the embodiments of the present invention, and not all of the embodiments. All other embodiments, which can be derived by a person skilled in the art from the embodiments given herein without making any creative effort, shall fall within the protection scope of the present invention.
Example 1
The preparation method of the diagnosis and treatment integrated nano platform using fullerene, doxorubicin hydrochloride and rare earth thulium doped up-conversion luminescent nanocrystals as main bodies, provided by the embodiment, comprises the following steps:
(1) mixing oil soluble NaYF4Yb and Tm are dispersed in cyclohexane, an acidic aqueous solution with the pH value of 3-5 is added, the reaction is carried out for 4-5 hours at room temperature to remove oleic acid ligand, the modified water-soluble nanocrystal is modified into water-soluble nanocrystal, centrifugation and washing are carried out, a solid is dispersed in water, citric acid is added into the solution, the mixture is fully stirred for 12 hours at room temperature and then centrifuged and washed to obtain the citric acid modified water-soluble rare earth up-conversion luminescence nanocrystal, the solid is dispersed in water, amino-modified fullerene (C60) is added, 1- (3-dimethylaminopropyl) -3-ethylcarbodiimide hydrochloride (EDC) and N-hydroxysuccinimide (NHS) are added while stirring, the mixture is stirred for 12 hours at room temperature and then centrifuged and washed to obtain the NaYF4Yb, Tm @ C60, and dispersing the product in water to obtain a first dispersion liquid;
the method specifically comprises the following steps: the mass ratio of the rare earth up-conversion luminescent nanocrystal to the citric acid is as follows: 10 mg: (50-100 mg); dissolving C60 (100-200 mg) and sodium hydroxide (NaOH) (3-4 g) in 200mL of toluene, dissolving 50mL of ethylenediamine in 100mL of 75% ethanol solution, mixing the two solutions, and stirring at room temperature for 7-10 days; and finally, drying for 24 hours at 50-60 ℃, washing the product with 75% ethanol, and drying for 24 hours at 60-70 ℃ to obtain the amino modified C60. The mass part ratio of EDC to NHS is as follows: 50 mg: (25-30 mg).
(2) Dissolving dithioglycolic acid, doxorubicin hydrochloride (Dox), EDC and NHS in water, adding the first dispersion, stirring at room temperature for 12 hours, centrifuging, washing, and dispersing the solid in water to form a second dispersion;
the method specifically comprises the following steps: the mass part ratio of dithioglycollic acid to doxorubicin hydrochloride (Dox), EDC and NHS is as follows: (5-10 mg): (5-10 mg): 50 mg: (25-30 mg).
(3) Dissolving 3A-amino-3A-deoxy- (2AS,3AS) -beta-cyclodextrin hydrate, Folic Acid (FA), EDC and NHS in water, stirring for 12 hours at room temperature, adding the second dispersion, stirring for 12 hours at room temperature, centrifuging and washing to obtain the diagnosis and treatment integrated nano platform taking fullerene, doxorubicin hydrochloride and rare earth up-conversion luminescent nanocrystals AS main bodies.
The method specifically comprises the following steps: the mass part ratio of the 3A-amino-3A-deoxy- (2AS,3AS) -beta-cyclodextrin hydrate to the Folic Acid (FA) to the EDC to the NHS is AS follows: (10-20 mg): (5-10 mg): 50 mg: (25-30 mg).
The diagnosis and treatment integrated nano platform prepared by the method is a nano composite system which takes a rare earth up-conversion luminescent nano crystal as a main body and bonds fullerene and doxorubicin hydrochloride, and is a nano composite system which takes an oil-soluble rare earth up-conversion luminescent nano crystal as a main body, carries out layer-by-layer organic functional micromolecule assembly on the surface of the nano crystal and links the organic micromolecules through chemical bonds to form the nano composite system; the fullerene C60 has a spherical structure, wherein an amido bond on one surface of the fullerene C60 is combined with a carboxyl group on the surface of the rare earth up-conversion luminescent nanocrystal, and the other surface of the fullerene C60 is simultaneously bonded with doxorubicin hydrochloride modified by a disulfide bond; the double-sided bonding material further performs host-guest action with folic acid modified beta-cyclodextrin to obtain a diagnosis and treatment integrated nano platform taking fullerene, doxorubicin hydrochloride and rare earth up-conversion luminescent nanocrystals as a host, and the diagnosis and treatment integrated nano platform has good water solubility, biocompatibility and targeting property, strong material stability and small overall size, and is beneficial to treatment and in-vivo discharge.
In other embodiments, the oil-soluble up-conversion luminescent nanocrystal of step (1) may be one of: NaYF4:Yb,Tm、NaYF4:Yb,Er,Tm、NaYF4:Yb,Tm@NaYF4、NaYF4:Yb,Er,Tm@NaYF4、NaYF4:Yb,Tm@NaGdF4、NaYF4:Yb,Er,Tm@NaGdF4、NaYF4:Yb,Tm@NaYF4@NaGdF4、NaYF4:Yb,Er,Tm@NaYF4@NaGdF4The oil-soluble up-conversion luminescence nano-crystal with other performances and similar structures can be selected to achieve the technical effect of the invention.
Example 2
The diagnosis and treatment integrated nano platform using fullerene, doxorubicin hydrochloride, rare earth thulium and erbium co-doped up-conversion luminescence nanocrystals as a main body and the preparation method thereof provided in this embodiment are basically the same as those in embodiment 1, and have the difference that the diagnosis and treatment integrated nano platform comprises the following steps:
(1) mixing oil soluble NaYF4Yb, Tm and Er are dispersed in cyclohexane, an acidic aqueous solution with the pH value of 3-5 is added, the reaction is carried out for 4-5 hours at room temperature to remove oleic acid ligand, the oleic acid ligand is modified into water-soluble nano-crystal, the water-soluble nano-crystal is centrifuged and washed, solid is dispersed in water, citric acid is added into the solution, the solution is fully stirred at room temperature for 12 hours and then centrifuged and washed, so that the citric acid modified water-soluble rare earth up-conversion luminescence nano-crystal is obtained, the solid is dispersed in water, amino-modified fullerene (C60) is added, 1- (3-dimethylaminopropyl) -3-ethyl carbodiimide hydrochloride (EDC) and N-hydroxysuccinimide (NHS) are added while stirring, the solution is stirred at room temperature for 12 hours and then centrifuged and washed, so that NaYF is obtained4Yb, Tm, Er @ C60, and dispersing the product in water to obtain a first dispersion liquid;
the method specifically comprises the following steps: the mass ratio of the rare earth up-conversion luminescent nanocrystal to the citric acid is as follows: 10 mg: (50-100 mg); dissolving C60 (100-200 mg) and sodium hydroxide (NaOH) (3-4 g) in 200mL of toluene, dissolving 50mL of ethylenediamine in 100mL of 75% ethanol solution, mixing the two solutions, and stirring at room temperature for 7-10 days; and finally, drying for 24 hours at 50-60 ℃, washing the product with 75% ethanol, and drying for 24 hours at 60-70 ℃ to obtain the amino modified C60. The mass part ratio of EDC to NHS is as follows: 50 mg: (25-30 mg);
(2) dissolving dithioglycolic acid, Dox, EDC and NHS in water, adding the first dispersion, stirring at room temperature for 12 hours, centrifuging, washing, and dispersing solids in water to form a second dispersion;
the method specifically comprises the following steps: the mass parts of dithiodiglycolic acid, Dox, EDC and NHS are as follows: (5-10 mg): (5-10 mg): 50 mg: (25-30 mg).
(3) Dissolving 3A-amino-3A-deoxy- (2AS,3AS) -beta-cyclodextrin hydrate, FA, EDC and NHS in water, stirring for 12 hours at room temperature, then adding a second dispersion, stirring for 12 hours at room temperature, centrifuging and washing to obtain the diagnosis and treatment integrated nano platform taking fullerene, doxorubicin hydrochloride and rare earth up-conversion luminescent nanocrystals AS main bodies.
The method specifically comprises the following steps: the mass part ratio of the 3A-amino-3A-deoxy- (2AS,3AS) -beta-cyclodextrin hydrate to the FA, the EDC and the NHS is AS follows: (10-20 mg): (5-10 mg): 50 mg: (25-30 mg).
Example 3
This example provides a fullerene, doxorubicin hydrochloride, and surface-coated inert NaYF4The diagnosis and treatment integrated nano platform taking rare earth thulium-doped up-conversion luminescent nanocrystals as main bodies and the preparation method thereof are basically the same as those of the embodiments 1 and 2, and have the following difference:
(1) mixing oil soluble NaYF4:Yb,Tm@NaYF4Dispersing in cyclohexane, adding an acidic aqueous solution with the pH value of 3-5, reacting at room temperature for 4-5 hours to remove oleic acid ligand, modifying the oleic acid ligand into water-soluble nanocrystal, centrifuging and washing, taking solid to disperse in water, adding citric acid into the solution, fully stirring at room temperature for 12 hours, centrifuging and washing to obtain the citric acid modified water-soluble nanocrystalDispersing the solid in water, adding amino-modified fullerene (C60), adding 1- (3-dimethylaminopropyl) -3-ethylcarbodiimide hydrochloride (EDC) and N-hydroxysuccinimide (NHS) while stirring, stirring at room temperature for 12 hours, centrifuging and washing to obtain NaYF4:Yb,Tm@NaYF4@ C60, dispersing the product in water to obtain a first dispersion;
the method specifically comprises the following steps: the mass ratio of the rare earth up-conversion luminescent nanocrystal to the citric acid is as follows: 10 mg: (50-100 mg); dissolving C60 (100-200 mg) and sodium hydroxide (NaOH) (3-4 g) in 200mL of toluene, dissolving 50mL of ethylenediamine in 100mL of 75% ethanol solution, mixing the two solutions, and stirring at room temperature for 7-10 days; and finally, drying for 24 hours at 50-60 ℃, washing the product with 75% ethanol, and drying for 24 hours at 60-70 ℃ to obtain the amino modified C60. The mass part ratio of EDC to NHS is as follows: 50 mg: (25-30 mg).
(2) Dissolving dithioglycolic acid, Dox, EDC and NHS in water, adding the first dispersion, stirring at room temperature for 12 hours, centrifuging, washing, and dispersing solids in water to form a second dispersion;
the method specifically comprises the following steps: the mass parts of dithiodiglycolic acid, Dox, EDC and NHS are as follows: (5-10 mg): (5-10 mg): 50 mg: (25-30 mg).
(3) Dissolving 3A-amino-3A-deoxy- (2AS,3AS) -beta-cyclodextrin hydrate, FA, EDC and NHS in water, stirring for 12 hours at room temperature, then adding a second dispersion, stirring for 12 hours at room temperature, centrifuging and washing to obtain the diagnosis and treatment integrated nano platform taking fullerene, doxorubicin hydrochloride and rare earth up-conversion luminescent nanocrystals AS main bodies.
The method specifically comprises the following steps: the mass part ratio of the 3A-amino-3A-deoxy- (2AS,3AS) -beta-cyclodextrin hydrate to the FA, the EDC and the NHS is AS follows: (10-20 mg): (5-10 mg): 50 mg: (25-30 mg).
Referring to fig. 1, a fullerene-based doxorubicin hydrochloride and surface-coated inert NaYF prepared according to embodiment 3 of the present invention4TEM photograph of diagnosis and treatment integrated nano material with rare earth thulium doped up-conversion luminescent nano crystal as main body, and nano particles can be seen from the imageThe components are stable and the dispersibility is good, so that the method can be used for preparing the nano material with good appearance and good monodispersity, and the average particle size of the functionalized nano material is 50-60 nm, and the small-sized nano particles are easier to be endocytosed by cells, so that the method is beneficial to circulation in organisms and has important significance for biological imaging and treatment.
Example 4
This example provides a fullerene-based, doxorubicin hydrochloride and surface-coated inert NaGdF4The diagnosis and treatment integrated nano platform taking rare earth thulium-doped up-conversion luminescent nanocrystals as main bodies and the preparation method thereof are basically the same as those in the embodiments 1 to 3, and have the following differences:
(1) mixing oil soluble NaYF4:Yb,Tm@NaGdF4Dispersing in cyclohexane, adding an acidic aqueous solution with the pH value of 3-5, reacting at room temperature for 4-5 hours to remove an oleic acid ligand, modifying the oleic acid ligand into water-soluble nanocrystals, centrifuging and washing, taking a solid to disperse in water, adding citric acid into the solution, fully stirring at room temperature for 12 hours, centrifuging and washing to obtain the citric acid modified water-soluble rare earth up-conversion luminescent nanocrystals, taking the solid to disperse in water, adding amino-modified fullerene (C60), adding 1- (3-dimethylaminopropyl) -3-ethylcarbodiimide hydrochloride (EDC) and N-hydroxysuccinimide (NHS) while stirring, stirring at room temperature for 12 hours, centrifuging and washing to obtain NaYF4:Yb,Tm@NaGdF4@ C60, dispersing the product in water to obtain a first dispersion;
the method specifically comprises the following steps: the mass ratio of the rare earth up-conversion luminescent nanocrystal to the citric acid is as follows: 10 mg: (50-100 mg); dissolving C60 (100-200 mg) and sodium hydroxide (NaOH) (3-4 g) in 200mL of toluene, dissolving 50mL of ethylenediamine in 100mL of 75% ethanol solution, mixing the two solutions, and stirring at room temperature for 7-10 days; and finally, drying for 24 hours at 50-60 ℃, washing the product with 75% ethanol, and drying for 24 hours at 60-70 ℃ to obtain the amino modified C60. The mass part ratio of EDC to NHS is as follows: 50 mg: (25-30 mg).
(2) Dissolving dithioglycolic acid, Dox, EDC and NHS in water, adding the first dispersion, stirring at room temperature for 12 hours, centrifuging, washing, and dispersing solids in water to form a second dispersion;
the method specifically comprises the following steps: the mass parts of dithiodiglycolic acid, Dox, EDC and NHS are as follows: (5-10 mg): (5-10 mg): 50 mg: (25-30 mg).
(3) Dissolving 3A-amino-3A-deoxy- (2AS,3AS) -beta-cyclodextrin hydrate, FA, EDC and NHS in water, stirring for 12 hours at room temperature, then adding a second dispersion, stirring for 12 hours at room temperature, centrifuging and washing to obtain the diagnosis and treatment integrated nano platform taking fullerene, doxorubicin hydrochloride and rare earth up-conversion luminescent nanocrystals AS main bodies.
The method specifically comprises the following steps: the mass part ratio of the 3A-amino-3A-deoxy- (2AS,3AS) -beta-cyclodextrin hydrate to the FA, the EDC and the NHS is AS follows: (10-20 mg): (5-10 mg): 50 mg: (25-30 mg).
Example 5
This example provides a fullerene-based, doxorubicin hydrochloride and surface-coated inert NaYF4The diagnosis and treatment integrated nano platform taking rare earth thulium-doped up-conversion luminescent nanocrystals as main bodies and the preparation method thereof are basically the same as those of the embodiments 1 to 4, and have the following differences:
(1) mixing oil soluble NaYF4:Yb,Tm@NaGdF4Dispersing in cyclohexane, adding an acidic aqueous solution with the pH value of 3-5, reacting for 4-5 hours at room temperature to remove oleic acid ligand, modifying the solution into water-soluble nanocrystals, centrifuging and washing, taking solid to disperse in water, adding polyacrylic acid (PAA) into the solution, fully stirring for 12 hours at room temperature, centrifuging and washing to obtain PAA-modified water-soluble rare earth up-conversion luminescent nanocrystals, taking solid to disperse in water, adding amino-modified fullerene (C60), adding 1- (3-dimethylaminopropyl) -3-ethylcarbodiimide hydrochloride (EDC) and N-hydroxysuccinimide (NHS) while stirring, stirring for 12 hours at room temperature, centrifuging and washing to obtain PAA-NaYF4:Yb,Tm@NaYF4@ C60, dispersing the product in water to obtain a first dispersion;
the method specifically comprises the following steps: the mass ratio of the rare earth up-conversion luminescent nanocrystal to the citric acid is as follows: 10 mg: (50-100 mg); dissolving C60 (100-200 mg) and sodium hydroxide (NaOH) (3-4 g) in 200mL of toluene, dissolving 50mL of ethylenediamine in 100mL of 75% ethanol solution, mixing the two solutions, and stirring at room temperature for 7-10 days; and finally, drying for 24 hours at 50-60 ℃, washing the product with 75% ethanol, and drying for 24 hours at 60-70 ℃ to obtain the amino modified C60. The mass part ratio of EDC to NHS is as follows: 50 mg: (25-30 mg).
(2) Dissolving dithioglycolic acid, Dox, EDC and NHS in water, adding the first dispersion, stirring at room temperature for 12 hours, centrifuging, washing, and dispersing solids in water to form a second dispersion;
the method specifically comprises the following steps: the mass parts of dithiodiglycolic acid, Dox, EDC and NHS are as follows: (5-10 mg): (5-10 mg): 50 mg: (25-30 mg).
(3) Dissolving 3A-amino-3A-deoxy- (2AS,3AS) -beta-cyclodextrin hydrate, FA, EDC and NHS in water, stirring for 12 hours at room temperature, then adding a second dispersion, stirring for 12 hours at room temperature, centrifuging and washing to obtain the diagnosis and treatment integrated nano platform taking fullerene, doxorubicin hydrochloride and rare earth up-conversion luminescent nanocrystals AS main bodies.
The method specifically comprises the following steps: the mass part ratio of the 3A-amino-3A-deoxy- (2AS,3AS) -beta-cyclodextrin hydrate to the FA, the EDC and the NHS is AS follows: (10-20 mg): (5-10 mg): 50 mg: (25-30 mg).
Application example 6
This example provides the fullerene, doxorubicin hydrochloride, and surface-coated inert NaYF prepared in example 34A diagnosis and treatment integrated nano platform taking rare earth thulium doped up-conversion luminescent nanocrystals as main bodies is used for a method for in-vitro singlet oxygen diagnosis, and comprises the following steps:
(1) preparing 1-2 mg of fullerene, doxorubicin hydrochloride and inert NaYF coated on the surface4The diagnosis and treatment integrated nano material taking rare earth thulium-doped up-conversion luminescent nanocrystals as main bodies, a cuvette and a 1,3 diphenyl isobenzofuran (DPBF) ethanol solution (7-10 mgDPBF dissolved in 5-10 mL of ethanol solution);
(2) dissolving the prepared nano material in 2mL of water, performing ultrasonic dispersion for 5 minutes, and adding 30-50 mu LDPBF ethanol solution to form a first dispersion solution;
(3) the first dispersion was added to a cuvette, irradiated with a 980nm laser for different times, and the absorption was measured with an ultraviolet absorption spectrometer.
FIG. 2 shows a C60-based inert NaYF coated substrate prepared in example 3 of the present invention4The ultraviolet absorption spectrogram of the rare earth thulium-doped upconversion nanocrystalline self-assembled nano material for in-vitro singlet oxygen determination can observe that an absorption peak at 420nm is gradually reduced along with the extension of irradiation time, and the structure corresponding to DPBF is damaged by singlet oxygen to cause the reduction of absorption, which indicates that the C60 functionalized upconversion nanocrystalline has good capacity of generating singlet oxygen.
Application example 7
This example provides the fullerene, doxorubicin hydrochloride, and surface-coated inert NaYF prepared in example 34The rare earth thulium doped up-conversion luminescent nano-crystal of the layer is used as a diagnosis and treatment integrated nano-platform of a main body, and the nano-platform is used as an auxiliary agent, an imaging agent of up-conversion fluorescence imaging is used for imaging diagnosis guided photodynamic therapy, and clinical diagnosis and treatment integration is realized; the method is particularly used for in vitro fluorescence imaging, and comprises the following steps:
(1) 1-2 mg of oil-soluble NaYF is prepared respectively4:Yb,Tm@NaYF4(UCN), citric acid modified rare earth up-conversion luminescence nano crystal (cit-UCN), NaYF4:Yb,Tm@NaYF4@C60(cit-UCN-C60)；
(2) Dissolving the prepared nano material in 2mL of deionized water, and performing ultrasonic dispersion for 5 minutes to form a first dispersion liquid;
(3) the first dispersion was added to a cuvette, irradiated with a 980nm laser (power 1W), and the intensity of luminescence was measured by a fluorescence spectrometer.
FIG. 3 is a view showing that the present invention is used in application example 3 to coat C60 with inert NaYF4The rare earth thulium-doped upconversion nanocrystalline self-assembled nano material of the layer has a fluorescence spectrogram under 980nm laser excitation, and emission peaks at 650nm, 475nm and 450nm can be observed from the fluorescence spectrogram, and correspond to Tm3+Is/are as follows1G4-3F4、1G4-3H6And, and1D2-3F4transition, and 980nm is just at the "optical window" of the biological tissue, indicating that based on C60 and surface coated with inert NaYF4The rare earth thulium-doped upconversion nanocrystalline self-assembled nanomaterial of the layer still keeps good upconversion fluorescence performance, and has a certain biological application prospect.
Application example 8
This example provides the fullerene, doxorubicin hydrochloride, and surface-coated inert NaYF prepared in example 34The characterization method of the diagnosis and treatment integrated nano platform taking rare earth thulium-doped up-conversion luminescent nanocrystals as main bodies comprises the following steps:
(1) mixing oil soluble NaYF4:Yb,Tm@NaYF4Dispersing in cyclohexane, adding an acidic aqueous solution with the pH value of 3-5, reacting at room temperature for 4-5 hours to remove an oleic acid ligand, modifying the oleic acid ligand into water-soluble nanocrystals, centrifuging and washing, taking a solid to disperse in water, adding citric acid into the solution, fully stirring at room temperature for 12 hours, centrifuging and washing to obtain the citric acid modified water-soluble rare earth up-conversion luminescent nanocrystals, taking the solid to disperse in water, adding amino-modified fullerene (C60), adding 1- (3-dimethylaminopropyl) -3-ethylcarbodiimide hydrochloride (EDC) and N-hydroxysuccinimide (NHS) while stirring, stirring at room temperature for 12 hours, centrifuging and washing to obtain NaYF4:Yb,Tm@NaYF4@ C60, dispersing the product in water to obtain a first dispersion;
the method specifically comprises the following steps: the mass ratio of the rare earth up-conversion luminescent nanocrystal to the citric acid is as follows: 10 mg: (50-100 mg); dissolving C60 (100-200 mg) and sodium hydroxide (NaOH) (3-4 g) in 200mL of toluene, dissolving 50mL of ethylenediamine in 100mL of 75% ethanol solution, mixing the two solutions, and stirring at room temperature for 7-10 days; and finally, drying for 24 hours at 50-60 ℃, washing the product with 75% ethanol, and drying for 24 hours at 60-70 ℃ to obtain the amino modified C60. The mass part ratio of EDC to NHS is as follows: 50 mg: (25-30 mg).
(2) Dissolving dithioglycolic acid, Dox, EDC and NHS in water, adding the first dispersion, stirring at room temperature for 12 hours, centrifuging, washing, and dispersing solids in water to form a second dispersion;
the method specifically comprises the following steps: the mass parts of dithiodiglycolic acid, Dox, EDC and NHS are as follows: (5-10 mg): (5-10 mg): 50 mg: (25-30 mg).
(3) Dissolving 3A-amino-3A-deoxy- (2AS,3AS) -beta-cyclodextrin hydrate, FA, EDC and NHS in water, stirring for 12 hours at room temperature, then adding a second dispersion, stirring for 12 hours at room temperature, centrifuging and washing to obtain the diagnosis and treatment integrated nano platform taking fullerene, doxorubicin hydrochloride and rare earth up-conversion luminescent nanocrystals AS main bodies.
The method specifically comprises the following steps: the mass part ratio of the 3A-amino-3A-deoxy- (2AS,3AS) -beta-cyclodextrin hydrate to the FA, the EDC and the NHS is AS follows: (10-20 mg): (5-10 mg): 50 mg: (25-30 mg).
(4) Respectively taking 1-2 mg of NaYF from which the oleic acid ligand is removed in the step (1)4:Yb,Tm@NaYF4(pure-UCN) and citric acid modified NaYF4:Yb,Tm@NaYF4(cit-UCN), taking 1-2 mg of the solid (UCN-C60) in the step (1), taking 1-2 mg of the solid (UCN-Dox) in the step (2) and 1-2 mg of the solid (UCN-FA) in the step (3), respectively dissolving the solid (UCN-Dox) in 2mL of water, placing the water in a cuvette, respectively measuring the surface potential of the cuvette by using a nanometer particle size and Zeta potential analyzer, and recording and processing the surface potential.
FIG. 4 is based on C60 and surface coated with inert NaYF4The rare earth thulium-doped upconversion nanocrystalline self-assembled nanomaterial of the layer is further used for carrying medicine and accessing a potential diagram (Zeta) of an intermediate material in a targeting process, and after each step of functionalization, the electrical property of the material is changed, so that successful assembly of organic functional micromolecules can be further explained.
Application example 9
This example provides a method of preparing a fullerene, doxorubicin hydrochloride, and surface-coated inert NaYF prepared according to any one of examples 1-54A method for in vitro glutathione response drug release of a diagnosis and treatment integrated nano platform with a rare earth thulium doped up-conversion luminescent nanocrystal as a main body comprises the following steps：
(1) Preparing 10-15 mg of fullerene-based doxorubicin hydrochloride and inert NaYF coated on the surface4The diagnosis and treatment integrated nano material taking rare earth thulium-doped up-conversion luminescent nanocrystals as main bodies, Phosphate Buffer Solution (PBS) with the pH value of 7.4 and glutathione powder;
(2) dissolving prepared glutathione in PBS to prepare a solution of 10 mg/mL;
(3) the final material was added to 2mL of the glutathione solution described above, stirred for 24 hours, centrifuged, and the supernatant was taken, and the drug release amount was calculated by measuring the absorption value of Dox at 480nm in the supernatant.
The key point of the invention is that the invention provides the fullerene, the doxorubicin hydrochloride and the inert NaYF coated on the surface4The diagnosis and treatment integrated nano platform with the rare earth thulium-doped up-conversion luminescent nanocrystals as the main body adopts organic small molecular components and is mutually bonded, so that the stability of the material is improved, the water solubility of the nano platform is improved, the nano platform has the necessary performance of diagnosis and treatment integration applied to up-conversion fluorescence imaging, photodynamic therapy and targeted chemotherapy, and the requirement of clinical diagnosis and treatment integration can be met. The preparation method provided by the invention has the advantages of simple process, low energy, environmental protection, safe operation, good repeatability and the like, and is easy to industrialize.
The invention provides a diagnosis and treatment integrated nano platform taking fullerene, doxorubicin hydrochloride and rare earth up-conversion luminescent nanocrystals as a main body and a preparation method and application thereof, which fully utilize the molecular surface structure and the bonding effect among molecules, firstly, a photosensitizer fullerene (C60) is modified on the surface of rare earth up-conversion luminescent nanocrystals (UCN), secondly, dithioglycolic acid (-S-S-) is introduced to improve the water solubility and simultaneously have the Glutathione (GSH) response release function, then, chemotherapy drug doxorubicin hydrochloride (Dox) is connected, and the formed nano platform and folic acid modified beta-cyclodextrin are subjected to host-object self-assembly to ensure that the nano platform has specific targeting and further improves the water solubility, thus obtaining the diagnosis and treatment integrated nano platform taking the fullerene, the doxorubicin hydrochloride and the rare earth up-conversion luminescent nanocrystals as the main body, can be used for optical imaging guided photodynamic and specific targeting drug release dual-mode treatment.
The present invention is not limited to the above-mentioned embodiments, and other similar methods of obtaining the nanocomposite material using the same or similar method, specific values are specifically selected within the ranges of values of the components described in the examples of the present invention, and up-conversion nanoparticles (NaYF) doped with different rare earth ions are used4:Yb,Tm、NaYF4:Yb,Er,Tm、NaYF4:Yb,Tm@NaYF4、NaYF4:Yb,Er,Tm@NaYF4、NaYF4:Yb,Tm@NaGdF4、NaYF4:Yb,Er,Tm@NaGdF4、NaYF4:Yb,Tm@NaYF4@NaGdF4、NaYF4:Yb,Er,Tm@NaYF4@NaGdF4) Etc., different organic molecules for further improving water solubility and functionalization, etc., are within the scope of the present invention, and the examples of the present invention are not listed.

Claims (9)

1. A preparation method of a diagnosis and treatment integrated nano platform is characterized by comprising the following steps:
(1) dispersing oil-soluble rare earth up-conversion luminescence nanocrystals in cyclohexane, adding an acidic aqueous solution with the pH value of 3-5, reacting at room temperature for 4-5 hours to remove oleic acid ligands, modifying the oleic acid ligands into water-soluble nanocrystals, centrifuging and washing, dispersing solids in water, adding citric acid into the solution, fully stirring at room temperature for 12 hours, centrifuging and washing to obtain citric acid modified water-soluble rare earth up-conversion luminescence nanocrystals, dispersing the solids in water, and adding amino-modified fullerene C60Adding 1- (3-dimethylaminopropyl) -3-ethylcarbodiimide hydrochloride (EDC) and N-hydroxysuccinimide (NHS) while stirring, stirring at room temperature for 12 hours, centrifuging, washing, and dispersing the solid in water to form a first dispersion liquid;
(2) dissolving dithioglycolic acid, doxorubicin hydrochloride (Dox), EDC and NHS in water, adding the first dispersion, stirring at room temperature for 12 hours, centrifuging, washing, and dispersing the solid in water to form a second dispersion;
(3) dissolving 3A-amino-3A-deoxy- (2AS,3AS) -beta-cyclodextrin hydrate, Folic Acid (FA), EDC and NHS in water, stirring at room temperature for 12 hours, adding the second dispersion, stirring at room temperature for 12 hours, centrifuging and washing to obtain the diagnosis and treatment integrated nano platform taking fullerene, doxorubicin hydrochloride and rare earth up-conversion luminescent nanocrystals AS main bodies.
2. The preparation method of the diagnosis and treatment integrated nano platform according to claim 1, wherein in the step (1):
the mass ratio of the rare earth up-conversion luminescent nanocrystal to the citric acid is as follows: 10: (500-1000); c is to be60Dissolving 100-200 mg of NaOH and 3000-4000 mg of NaOH in 200mL of methylbenzene, dissolving 50mL of ethylenediamine in 100mL of 75% ethanol solution, mixing the two solutions, and stirring at room temperature for 7-10 days; finally, drying for 24 hours at 50-60 ℃, cleaning the product with 75% ethanol, and drying for 24 hours at 60-70 ℃ to obtain amino modified C60(ii) a The mass part ratio of EDC to NHS is as follows: 50: (25-30).
3. The method for preparing the diagnosis and treatment integrated nano platform according to claim 1, wherein in the step (2):
the mass part ratio of dithioglycollic acid to doxorubicin hydrochloride (Dox), EDC and NHS is as follows: (5-10): (5-10): 50: (25-30).
4. The preparation method of the diagnosis and treatment integrated nano platform according to claim 1, wherein in the step (3):
the mass part ratio of the 3A-amino-3A-deoxy- (2AS,3AS) -beta-cyclodextrin hydrate to the Folic Acid (FA) to the EDC to the NHS is AS follows: (10-20): (5-10): 50: (25-30).
5. The preparation method of the diagnosis and treatment integrated nano platform according to claim 1, wherein the oil-soluble up-conversion luminescence nano crystal in the step (1) comprises: NaYF4:Yb,Tm、NaYF4:Yb,Er,Tm、NaYF4:Yb,Tm@NaYF4、NaYF4:Yb,Er,Tm@NaYF4、NaYF4:Yb,Tm@NaGdF4、NaYF4:Yb,Er,Tm@NaGdF4、NaYF4:Yb,Tm@NaYF4@NaGdF4、NaYF4:Yb,Er,Tm@NaYF4@NaGdF4。
6. The diagnosis and treatment integrated nano platform prepared by the method according to any one of claims 1 to 5, which is a nano composite system mainly comprising rare earth up-conversion luminescent nanocrystals and bonded with fullerene and doxorubicin hydrochloride, wherein the nano composite system mainly comprises the rare earth up-conversion luminescent nanocrystals, is formed by assembling organic functional small molecules layer by layer on the surface of the nanocrystals and linking the organic small molecules through chemical bonds; the fullerene C60The rare earth upconversion luminescent nanocrystal is of a spherical structure, wherein an amido bond on one surface of the nanocrystal is combined with a carboxyl on the surface of the rare earth upconversion luminescent nanocrystal, and the other surface of the nanocrystal is simultaneously bonded with doxorubicin hydrochloride modified by a disulfide bond; the double-sided bonding material further performs host-guest action with folic acid modified beta-cyclodextrin to obtain the diagnosis and treatment integrated nano platform taking fullerene, doxorubicin hydrochloride and rare earth up-conversion luminescent nanocrystals as a host.
7. The application of the diagnosis and treatment integrated nano platform according to claim 6 is characterized in that the nano platform is used as a raw material to prepare a diagnosis and treatment auxiliary agent for photodynamic and drug dual-cooperative treatment guided by optical imaging diagnosis and for realizing diagnosis and treatment integration of clinical phototherapy composite chemotherapy.
8. The application of the integrated diagnosis and treatment nano platform as claimed in claim 6, which is characterized in that the integrated diagnosis and treatment nano platform is used as a raw material for preparing an imaging agent for up-conversion fluorescence imaging.
9. The use of the integrated nanoplatform of diagnosis and treatment according to claim 6, characterized in that it is used for the preparation of a carrier material for precise drug release of Glutathione (GSH) response.
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Citations (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
EP2075304A1 (en) * 2007-12-04 2009-07-01 Sony Corporation A device for modifying the wavelenght range of a spectrum of light
CN106833644A (en) * 2017-01-17 2017-06-13 上海大学 A kind of nano material based on post aromatic hydrocarbons and the crystalline substance self assembly of upper conversion nano and preparation method and application

Patent Citations (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
EP2075304A1 (en) * 2007-12-04 2009-07-01 Sony Corporation A device for modifying the wavelenght range of a spectrum of light
CN106833644A (en) * 2017-01-17 2017-06-13 上海大学 A kind of nano material based on post aromatic hydrocarbons and the crystalline substance self assembly of upper conversion nano and preparation method and application

Non-Patent Citations (2)

* Cited by examiner, † Cited by third party
Title
"Multifunctional upconversion–nanoparticles-trismethylpyridylporphyrin–fullerene nanocomposite:a near-infrared light-triggered theranostic platform for imaging-guided photodynamic therapy";Mirong Guan等;《NPG Asia Materials》;20150525(第7期);第1-11页 *
"靶向上转换发光纳米探针用于生物传感、成像及治疗";王旭;《南开大学博士学位论文》;20140530;第1-115页 *

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