CN113927027B

CN113927027B - Virus-like hollow manganese oxide loaded near-infrared two-b-region excited rare earth nanocrystalline and preparation method and application thereof

Info

Publication number: CN113927027B
Application number: CN202111096606.6A
Authority: CN
Inventors: 刘小龙; 王培园; 吴名; 李佳琦; 李阳; 蔺金燕; 罗强
Original assignee: Mengchao Hepatobiliary Hospital Of Fujian Medical University (fuzhou Hospital For Infectious Diseases)
Current assignee: Mengchao Hepatobiliary Hospital Of Fujian Medical University (fuzhou Hospital For Infectious Diseases)
Priority date: 2021-09-16
Filing date: 2021-09-16
Publication date: 2023-06-06
Anticipated expiration: 2041-09-16
Also published as: CN113927027A; WO2023041005A1

Abstract

The invention discloses a virus-like hollow oxide loaded near infrared two b region excited rare earth nanocrystalline and a preparation method and application thereof, wherein the invention firstly synthesizes Er and Ho doped NaErF by a hydrothermal method ₄ :2％Ho@NaYF ₄ The rare earth nano material is then modified on the surface of the viroid hollow manganese oxide poured on the IR1064 through amide reaction, so as to obtain the fluorescent composite probe with good biocompatibility and tumor microenvironment response. After manganese oxide of the fluorescent composite probe is degraded at a tumor, manganese ions are used for chemical dynamic treatment of a metastasis and a nuclear magnetic resonance imaging contrast agent, released rare earth nanocrystals have excitation of 1530nm, emission light of 650nm and emission light of 1180nm, wherein 1180nm can be used for tumor imaging navigation surgery excision with no background interference in a near infrared two region and high resolution and tissue penetrating power, and 650nm can be used for near infrared one-region fluorescent imaging.

Description

Virus-like hollow manganese oxide loaded near-infrared two-b-region excited rare earth nanocrystalline and preparation method and application thereof

Technical Field

The invention belongs to the field of biomedical nano materials, and particularly relates to a virus-like hollow manganese oxide loaded near-infrared two-b region excited rare earth nanocrystal, a preparation method thereof and application thereof in surgical navigation and postoperative chemical power treatment.

Background

Of the current treatments for cancer, complete surgical resection is the most common and desirable method of choice. However, in the conventional preoperative detection means for tumors, the position, edge and micrometastasis of the tumor are difficult to effectively judge and find due to the limitation of the self resolution, and residual tumor cells after surgical excision can cause recurrence of the tumor after the operation of the patient. Accordingly, there is a need for an efficient imaging means for locating the location and boundaries of a tumor in real-time, accurate, objective and precise manner during surgical resection and for finding micrometastases, thereby assisting the clinician in thoroughly resecting the tumor to reduce the postoperative recurrence rate of the patient, which is significant for improving the postoperative survival rate of tumor patients.

Because of the rapid response speed and high detection sensitivity, fluorescence imaging has become one of the most effective approaches for early tumor diagnosis, assisted surgery navigation, detection of recurrent or residual lesions, efficacy monitoring and the like. Current fluorescent surgical navigational probes focus mainly on the first window near infrared range (the first window near-infused region, NIR I, 650-900 nm). Due to limitations of fluorescence tissue penetration, signal-to-noise ratio of NIR I, it is not suitable for fluorescence imaging of tumors located in deep parts of organs, such as brain tumor, ovarian tumor, liver tumor, and lymphatic metastases. Studies have shown that: the second window near infrared light (the second window near-infused region, NIR II, 1000-1700 nm) has deeper tissue penetration depth, higher signal-to-noise ratio, lower photodamage than NIR I fluorescence. Therefore, NIR II fluorescence imaging can be used for precisely positioning tumor positions, clearly observing tumor edges and performing efficient excision.

The intelligent nano carrier is adopted to convey the medicine to the tumor part, and the medicine is specifically and controllably released to the cancerous position under the stimulation of a special microenvironment or a marker of the tumor by utilizing the difference between tumor tissues and normal tissues, so that the curative effect of tumor treatment can be improved, and the toxic and side effects can be reduced. Therefore, the accurate and controllable release of the drug molecules can be realized through the specificity of tumor cells. Manganese oxide is widely applied to tumor treatment, manganese ions released by degradation can generate singlet oxygen in an acidic environment of the tumor for chemo-dynamic treatment, and can also be used for nuclear magnetic resonance imaging. In addition, the viroid silica nanoparticle with a rough surface can greatly increase the uptake rate of cells due to the adhesion of cells. Therefore, development of a new method for constructing viroid metal mesoporous oxide, which is simple in method, is needed to improve tumor cell uptake and biosafety, and has great significance for biological application and clinical transformation.

Disclosure of Invention

In order to improve the technical problems, the invention provides a rare earth nanocrystalline, which comprises a carrier and rare earth core-shell nano particles loaded on the surface of the carrier.

According to an embodiment of the invention, the mass ratio of the carrier to the rare earth core-shell nanoparticles is 1 (1-2), preferably 1 (1-1.5), and is exemplified by 1:1, 1:1.2, 1:1.5, 1:2.

According to an embodiment of the invention, the support is connected to the rare earth core-shell nanoparticle by a valence bond, for example by an amide bond.

According to an embodiment of the invention, the support has an amino group which forms an amide bond with the carboxyl group in the rare earth core-shell nanoparticle.

According to an embodiment of the invention, the rare earth nanocrystalline is selectively degradable in a weakly acidic tumor microenvironment, preferably selectively degradable in a weakly acidic tumor microenvironment to release manganese ions (Mn ²⁺ ) Iron ions (Fe) ²⁺ )。

According to an embodiment of the invention, the support is a hollow sphere surface-modified with an amino-functional compound. For example, the carrier is hollow manganese oxide or hollow iron oxide modified by amination.

Preferably, the compound bearing an amino function may be, for example, the aminosilane APTES (3-amino-propyl) -triethoxysilane).

Preferably, the carrier has fluorescent quenching molecules supported in its cavity. For example, the fluorescence quenching molecule may be a near infrared absorber, such as IR1064. For another example, the loading of the fluorescence quenching molecules is 10-15%, and exemplary 10%, 12%, 15% of the total weight of the carrier.

Preferably, the hollow manganese oxide is nano particles, has a virus-like morphology, can be rapidly phagocytosed by tumor cells, and realizes accurate imaging and treatment of tumors.

The viroid hollow manganese oxide can be degraded and collapsed in the tumor acidic environment, and released manganese ions can be used for chemodynamic treatment of postoperative tumor metastasis or for nuclear magnetic resonance imaging of tumors; after the fluorescent quenching molecule IR1064 is released, the fluorescence of the rare earth nano particles is recovered, so that the fluorescent quenching fluorescent nanoparticle can be used for fluorescent imaging surgical navigation excision of tumors.

According to an embodiment of the present invention, the rare earth core-shell nanoparticle comprises a core-layer nanoparticle, at least one shell layer coated on the outside of the core-layer nanoparticle, and a polymer modified on the outside of the shell layer and containing carboxyl functional groups, wherein the core-layer nanoparticle and the shell layer are respectively and independently selected from AREF ₄ Wherein: a is Na or K, RE is at least one of Er, ho, gd and Y.

Preferably, a is Na.

Preferably, the RE element in the core layer nanoparticle and the shell layer are different. Preferably, in the core layer nanoparticle, RE is two of Er and Ho; in the shell nanoparticle, RE is Y.

Preferably, the doping amount of Ho in the core layer nanoparticle is 1-5%, and exemplary is 1%, 2%, 5%.

Preferably, the particle size of the rare earth core-shell nanoparticle is 24-25.5 nm, and exemplary are 24nm, 24.13nm, 25nm, 25.25nm, 25.5nm.

Preferably, the ratio of the particle size of the core layer nanoparticle to the thickness of the shell layer is 1 (1-1.5), and exemplary are 1:1, 1:1.18, and 1:1.5.

Preferably, the particle size of the core layer nanoparticle is 17 to 18nm, and exemplary is 17.09nm, 17.5nm, 17.95nm.

Preferably, the polymer containing carboxyl functional groups canThe polymer is PEG polymer, small molecule acid or carboxyl-containing polymer, such as phospholipid polyethylene glycol (DSEP-PEG) ₂₀₀₀ -COOH), maleamic acid, polyacrylic acid, and the like.

The polymer containing carboxyl functional groups can enhance the biocompatibility of the material and reduce cytotoxicity.

Preferably, the rare earth core-shell nanoparticle has an optical contrast function in a near infrared first region (650-900 nm) and a near infrared second region (1000-1700 nm).

According to an embodiment of the present invention, the rare earth nanocrystals are monodisperse and uniform in size. Preferably, the average particle size of the rare earth nanocrystals is about 24 to 26nm, with 24.13nm, 24.69nm, 25nm, 25.25nm being exemplary.

According to an embodiment of the present invention, the rare earth nanocrystalline may be excited by a near infrared two b region, producing near infrared one region, near infrared two region fluorescence emission.

For example, the rare earth nanocrystals may emit red light in the near infrared region of 660nm upon excitation at 1532 nm. For near infrared one-region fluorescence imaging, and can be used as a reference for near infrared two-region imaging.

For example, the rare earth nanocrystals can emit 1180nm fluorescence upon excitation at 1532 nm. The method is used for near infrared two-region fluorescence imaging navigation tumor surgical excision, solves the problems of low resolution and penetration potential in the current surgical navigation, and can be used for deep tissue background-free fluorescence imaging.

According to embodiments of the invention, the rare earth core-shell nanoparticle may be, for example, beta-NaErF ₄ :Ho@NaYF ₄ 。

The rare earth nanocrystalline particles can realize fluorescence imaging navigation tumor excision, fluorescence imaging guidance of postoperative metastasis, tumor photodynamic enhancement and nuclear magnetic resonance imaging of tumors, greatly improve the tumor removal efficiency and provide guidance for the treatment of clinical malignant tumors.

The invention also provides a preparation method of the rare earth nanocrystalline, which comprises the steps of reacting a carrier with rare earth core-shell nano particles to prepare the rare earth nanocrystalline.

According to an embodiment of the invention, the mass ratio of the rare earth core-shell nanoparticle to the carrier is 2-5:1, and exemplary are 2:1, 3:1, 4:1 and 5:1.

According to an embodiment of the invention, the preparation method of the rare earth core-shell nanoparticle comprises the following steps:

(1) Adding rare earth salt into a mixed solution of oleic acid and octadecene, and then adding an alkali metal fluoride solution and an alkaline solution for reaction to obtain the nuclear layer nano-particles;

(2) And (3) adding RE-OA and A-TFA-OA precursor solutions into the product of the step (1) to react, so as to prepare the rare earth core-shell nanoparticle.

Preferably, step (2) is performed at least once to obtain nanoparticles coated with at least one shell layer. Illustratively, 1, 2, 3 or more times;

wherein A is Na or K, RE is at least one of Er, ho, gd and Y, and preferably Y.

Preferably, the alkali metal fluoride solution is NaF, NH ₄ One of the solutions of F and KF, preferably NH ₄ F solution.

Preferably, in step (1), the molar ratio of rare earth ions to alkali metal fluoride in the rare earth salt is 1 (2-5), and is exemplified by 1:4.

Preferably, the rare earth ions in the rare earth salt are at least one of Er, ho and Y, and preferably two of Er and Ho.

Preferably, the molar ratio of the Er to Ho is (48-49.5): (0.5-2), exemplary 49.5:0.5, 49:1, 48:2.

Preferably, in step (1), the core layer nanoparticle is further dissolved in a solvent, for example, cyclohexane. For example, the core layer nanoparticle dispersion may have a concentration of 0.05 to 0.2mol/L, and exemplary is 0.1mol/L.

Preferably, in step (1), the rare earth salt is a rare earth chloride, rare earth nitrate or rare earth acetate, preferably rare earth acetate.

For example, the rare earth chloridesIs HoCl ₃ 、ErCl ₃ And YCl ₃ At least one of them.

For example, the rare earth nitrate is Ho (NO ₃ ) ₃ 、Er(NO ₃ ) ₃ And Y (NO) ₃ ) ₃ At least one of them.

For example, the rare earth acetate is Ho (CH) ₃ COO) ₃ 、Er(CH ₃ COO) ₃ And Y (CH) ₃ COO) ₃ At least one of them.

Preferably, in step (1), the alkaline solution is a sodium hydroxide solution or a potassium hydroxide solution.

Preferably, the solvent used in the alkali metal fluoride solution and the alkaline solution is methanol.

Preferably, in step (1), the total amount of rare earth salt is 0.5 to 2mmol, and exemplary is 0.5mmol, 1mmol, 2mmol.

Preferably, in step (1), the ratio of oleic acid to octadecene is 1 (2-3), and is exemplified by 1:2, 1:2.5, and 1:3.

Preferably, in the step (1), the mixed solution of oleic acid and octadecene after adding rare earth salt is heated and stirred to remove water and oxygen in the system. For example, the temperature of the heating and stirring is 130 to 150 ℃, and 130 ℃ and 140 ℃ and 150 ℃ are exemplified; the heating and stirring time is 20-40 min. Exemplary are 20min, 30min, 40min. Further, the heating and stirring are performed under vacuum conditions.

Preferably, in the step (1), the mixed solution after adding the alkali metal fluoride solution and the alkaline solution is further stirred for nucleation. For example, the stirring nucleation adopts a secondary heating mode.

Preferably, in the secondary heating mode:

the temperature of the first stage heating is 40-60 ℃, and is exemplified by 40 ℃, 50 ℃ and 60 ℃; the first stage heating time is 0.5-2 h, and is exemplified by 0.5h, 1h and 2h;

the temperature of the second stage heating is 60-80 ℃, and is 60 ℃ and 70 ℃ for example; the second stage heating time is 0.5-2 h, and exemplary is 0.5h, 1h, 2h.

Preferably, the preparation method further comprises the step of carrying out solid-liquid separation on the nuclear layer nanoparticle mixed liquid prepared in the step (1) to obtain a reaction product. For example, the solid-liquid separation method is to add a precipitant into the mixed solution for precipitation, and centrifuge to obtain a solid product. As another example, the precipitating agent may be ethanol.

Preferably, the preparation method further comprises washing the reaction product obtained by the solid-liquid separation in the step (1). For example, the solvent for washing may be absolute ethanol.

Preferably, in step (2), the core layer nanoparticle dispersion is further added to a mixed solution of oleic acid and octadecene before adding the RE-OA and a-TFA-OA precursor solutions to the product of step (1). For example, the volume ratio of the core layer nanoparticle dispersion to the mixed solution of oleic acid and octadecene is 1 (1-3), and is exemplified by 1:2. As another example, in a mixed solution of oleic acid and octadecene, the volume ratio of oleic acid to octadecene is 1 (1-2), and is exemplified by 1:1.5.

Preferably, in step (2), the concentration of the RE-OA precursor solution is 0.05 to 0.2mol/L, and exemplary is 0.05mol/L, 0.1mol/L, 0.2mol/L.

According to an exemplary embodiment of the present invention, the RE-OA precursor solution is prepared by dissolving a salt containing an RE element in a mixed solvent of oleic acid and octadecene. Preferably, in the mixed solvent, the volume ratio of oleic acid to octadecene is 1 (1-2), and the exemplary ratio is 1:1, 1:1.5 and 1:2. Further, the method also comprises the step of heating and stirring the mixed solution. For example, the temperature of the heating may be 130 to 150 ℃, illustratively 130 ℃, 140 ℃, 150 ℃; the heating time is 0.5-2 h, and exemplary is 0.5h, 1h, 2h. For another example, the salt containing RE element is rare earth chloride, rare earth nitrate or rare earth acetate, preferably chloride.

Preferably, in step (2), the concentration of the A-TFA-OA precursor solution is 0.3 to 0.5mol/L, and exemplified by 0.3mol/L, 0.4mol/L, and 0.5mol/L.

According to an exemplary embodiment of the present invention, the a-TFA-OA precursor solution is prepared by dissolving a trifluoroacetate salt comprising an a element in oleic acid. Preferably, the preparation of the A-TFA-OA precursor solution further comprises stirring the mixed solution.

Preferably, in step (2), the RE-OA precursor solution and the a-TFA-OA precursor solution are added to the core layer nanoparticle dispersion liquid prepared in step (1) in an alternately spaced manner. For example, the RE-OA precursor solution and A-TFA-OA precursor solution may be added alternately at least once each. Illustratively, 1, 2 or more times of each addition are alternated; preferably three times each alternately. For another example, the interval may be 10 to 20min, and exemplary 10min, 15min, and 20min.

Preferably, the molar ratio of the rare earth salt to the rare earth ions and oleic acid in the RE-OA precursor solution in step (2) is 1 (6-8), exemplary 1:6, 1:7.5, 1:8.

Preferably, the reactions of step (1) and step (2) are carried out under an inert atmosphere. For example, the inert atmosphere is nitrogen or argon.

Preferably, in steps (1), (2), the temperatures of the reactions are the same or different, independently of each other, from 200 to 400 ℃, illustratively 200 ℃, 280 ℃, 300 ℃, 400 ℃. Preferably, in step (1), the reaction time is 40-80min, and exemplary is 40min, 60min, 80min.

Preferably, in steps (1) and (2), the temperature rise rate of the reaction is 5-20 ℃/min, and is exemplified by 5 ℃/min, 10 ℃/min, and 20 ℃/min.

Preferably, the preparation method further comprises step (3): uniformly mixing the nano particles coated with at least one layer of shell layer obtained in the step (2) with a polymer containing carboxyl functional groups in an organic solvent, standing until the organic solvent volatilizes, adding water for dispersion, and centrifugally separating the carboxyl modified rare earth core-shell nano particles.

Preferably, in step (3), the organic solvent is chloroform, cyclohexane, n-hexane, tetrahydrofuran, preferably chloroform.

Preferably, in step (3), the centrifugal separation speed is 8000 to 20000rpm, and exemplified is 8000rpm, 10000rpm, 15000rpm, 17500rpm, 20000rpm. Further, the centrifugation time is 20-40 min, and is exemplified by 20min, 30min, and 40min.

Further, in the step (3), the ratio of the nanoparticles coating at least one shell layer to the polymer containing carboxyl functional groups is 0.1mmol (20-30) mg, and exemplary is 0.1mmol:20mg, 0.1mmol:25mg, 0.1mmol:30mg.

According to an exemplary embodiment of the present invention, the method for preparing rare earth nanocrystals includes the steps of:

(A1)β-NaErF ₄ preparation of 2% Ho core layer

Er (CH) ₃ CO ₂ ) ₃ ·4H ₂ O、Ho(CH ₃ CO ₂ ) ₃ ·4H ₂ Adding O into a mixed solution of oleic acid and octadecene, uniformly mixing, heating and stirring (removing water and oxygen in a system) under a vacuum condition, and finally cooling to room temperature to obtain a clear and transparent solution; then adding methanol solution of ammonium fluoride and sodium hydroxide, maintaining for 0.5-2 h at 40-60 ℃ for full stirring and nucleation, heating to 60-80 ℃, and maintaining for 0.5-2 h (removing redundant methanol, oxygen and water molecules) under vacuum; then, reacting for 50-70 min under the protection of argon at 300 ℃;

(A2) Preparation of the precursor

(1) Synthesis of Y-OA precursor: YCl is combined with ₃ Dissolving in a mixed solvent of oleic acid and octadecene, heating and stirring under the condition of keeping vacuum to obtain a clear and transparent Y-OA precursor solution;

(2) synthesis of Na-TFA-OA precursor: dissolving sodium trifluoroacetate in oleic acid, mixing, vacuumizing, stirring uniformly until the sodium trifluoroacetate is completely dissolved, and obtaining a pale yellow transparent precursor oleic acid solution;

(3)NaYF ₄ is coated by adopting a continuous layer-by-layer growth method

First, naErF is carried out ₄ Adding 2% Ho core layer nanoparticle dispersion into oleic acid and octadecene mixed solution, removing cyclohexane from the system under vacuum, introducing argon, heating, and alternately adding Y-OA and Na-TFA-OA precursor solution to obtainThe luminescent rare earth nanocrystalline is obtained.

According to the embodiment of the invention, the preparation method of the carrier comprises the steps of taking the viroid silicon mesoporous nano-particles as a template, reacting with manganese salt and urotropine, and removing the viroid silicon mesoporous nano-particle template through alkali treatment to obtain the viroid hollow manganese oxide.

According to an exemplary embodiment of the invention, the viroid silicon mesoporous nano-particles are prepared by reacting tetraethyl orthosilicate with cetyltrimethylammonium bromide (CTAB) and triethylamine.

Preferably, both tetraethyl orthosilicate and cetyltrimethylammonium bromide (CTAB) are added to the reaction system in solution. For example, a cyclohexane solution containing tetraethyl orthosilicate and an aqueous solution of cetyltrimethylammonium bromide (CTAB) are prepared separately. Preferably, firstly, mixing an aqueous solution of Cetyl Trimethyl Ammonium Bromide (CTAB) with triethylamine, and then mixing the aqueous solution with a cyclohexane solution of tetraethyl orthosilicate, and reacting to obtain the viroid silicon mesoporous nano-particles.

According to an exemplary embodiment of the invention, the tetraethyl orthosilicate to cetyltrimethylammonium bromide (CTAB), triethylamine are used in a 5.3mL:2g:1mL ratio.

According to an exemplary embodiment of the present invention, the concentration of triethylamine may be 10 to 30%, and exemplary 10%, 25%, 30%.

Preferably, the preparation method of the viroid silicon mesoporous nano-particles further comprises the step of carrying out solid-liquid separation on the prepared viroid silicon mesoporous nano-particles. For example, the solid-liquid separation may be by means known in the art, such as filtration, centrifugation.

Preferably, the preparation method further comprises washing the reaction product obtained by solid-liquid separation. For example, the solvent used for washing may be water or ethanol. As another example, the number of times of washing may be one, two, three.

Preferably, the reaction of the viroid silicon mesoporous nano-particles with manganese salt and urotropine is carried out in a solvent system. For example, the viroid silicon mesoporous nano-particles are firstly dispersed in water, and then manganese salt and urotropine are added. As another example, the concentration of the viroid silicon mesoporous nanoparticle dispersion is 1-3 mg/mL, and exemplary is 1mg/mL, 2mg/mL, 3mg/mL.

According to an exemplary embodiment of the invention, the mass ratio of the viroid silicon mesoporous nano-particles to manganese salt and urotropine is 10:9:9. The manganese salt may be, for example, mn (NO ₃ ) ₂ ·6H ₂ O。

According to an exemplary embodiment of the present invention, the reaction temperature of the viroid silicon mesoporous nanoparticle with manganese salt and urotropine is 80-100 ℃, and is exemplary 80 ℃, 90 ℃ and 100 ℃. Further, the reaction time is 3 to 5 hours, and is exemplified by 3 hours, 4 hours, and 5 hours.

According to an exemplary embodiment of the invention, the alkaline solution used for the alkaline treatment is an aqueous NaOH solution. Preferably, the temperature of the alkali treatment is 60 to 80 ℃, and exemplary is 60 ℃, 75 ℃, 80 ℃. Further, the concentration of the alkali liquor is 1 to 3mol/L, and 1mol/L, 2mol/L and 3mol/L are exemplified.

Preferably, the preparation method of the viroid mesoporous manganese oxide further comprises the step of carrying out solid-liquid separation on the prepared viroid mesoporous manganese oxide reaction liquid. For example, the solid-liquid separation may be by means known in the art, such as filtration, centrifugation.

Preferably, the preparation method of the viroid mesoporous manganese oxide further comprises the step of washing a reaction product obtained by solid-liquid separation. For example, the solvent used for washing may be water or ethanol. As another example, the number of times of washing may be one, two, three.

Preferably, the preparation method of the viroid mesoporous manganese oxide further comprises the step of reacting the washed reaction product with a compound containing amino functional groups to prepare the viroid mesoporous manganese oxide with the amino functional groups modified on the surface.

Preferably, the preparation method of the viroid mesoporous manganese oxide further comprises the step of reacting the viroid mesoporous manganese oxide with the amino functional groups modified on the surface with fluorescence quenching molecules to prepare the viroid mesoporous manganese oxide with the amino functional groups modified on the surface of the fluorescence quenching molecules. For example, the mass ratio of the viroid mesoporous manganese oxide with the amino functional group modified on the surface to the fluorescence quenching molecule is (3-5): 1, and exemplary are 3:1, 4:1 and 5:1. As another example, the reaction time is not less than 12 hours, and is exemplified by 24 hours.

Preferably, the preparation method of the viroid mesoporous manganese oxide further comprises the step of carrying out solid-liquid separation on the viroid mesoporous manganese oxide loaded with the fluorescence quenching molecule and modified with amino functional groups on the surface. For example, the solid-liquid separation may employ means known in the art, such as centrifugation.

According to an embodiment of the present invention, the method for preparing the carrier comprises the steps of:

(S1) synthesizing viroid silicon mesoporous nano particles by a two-phase method;

cetyl Trimethyl Ammonium Bromide (CTAB) is dissolved in water, and then mixed solution of triethylamine, cyclohexane and tetraethyl orthosilicate is added to prepare the viroid silicon particles through reaction;

(S2) preparation of viroid hollow manganese oxide

Adding Mn (NO) into the viroid silicon mesoporous nano particle dispersion liquid under the condition of heating and stirring ₃ ) ₂ ·6H ₂ O and urotropine, after reaction, centrifugally washing, and finally etching off the viroid mesoporous silicon template in NaOH aqueous solution to prepare the hollow viroid manganese oxide; then, the viroid hollow manganese oxide reacts with a compound containing amino functional groups to prepare viroid mesoporous manganese oxide with the amino functional groups modified on the surface;

(S3) preparation of viroid hollow manganese oxide with IR1064 loaded and rare earth nanocrystalline connected on surface

And (3) dissolving the viroid hollow manganese oxide particles prepared in the step (S2) in water, then adding IR1064, stirring at room temperature for reaction, and centrifuging to remove redundant IR1064, thereby preparing the viroid hollow manganese oxide particles loaded with the IR1064 surface and connected with rare earth nanocrystals.

According to the embodiment of the invention, in the preparation method of the rare earth nanocrystalline, the reaction of the carrier and the rare earth core-shell nanoparticle also comprises the addition of an activating agent. For example, the activator may be at least one of carbodiimide (EDC), N-hydroxysuccinimide (NHS), and Dimethylacetamide (DMAC); illustratively, the activators are EDC and NHS.

Preferably, the process of activator activation is performed in a solution at ph=8.0 to 9.0 (exemplary ph=8.5). Further, the time of activation is not less than 10 hours, and is exemplified by 12 hours.

Preferably, the mass ratio of the rare earth core-shell nano particles to the activator is 1:3-5.

Preferably, the mass ratio of the rare earth core-shell nano particles to the carrier is 2-5:1.

The invention also provides a composite probe which contains the rare earth nanocrystalline.

The invention also provides application of the rare earth nanocrystalline or the composite probe in preparing and/or serving as a contrast agent in the fields of surgical navigation (such as fluorescence imaging navigation tumor resection), postoperative chemo-dynamic treatment, fluorescence guidance of postoperative metastasis, nuclear magnetic resonance imaging of tumors and the like.

For example, the rare earth nanocrystals find use in the preparation and/or as near infrared two-region optical/nuclear magnetic resonance contrast agents.

Preferably, the near infrared two-region optical contrast agent is applied in vascular imaging and lymph node visualization.

Preferably, when used as a near infrared two-region optical contrast agent, the emission wavelength is 1000-1700nm and the excitation wavelength is 700-1100nm.

The invention has the beneficial effects that:

(1) The invention utilizes hydrothermal solvent to thermally synthesize rare earth nanocrystals excited by the near infrared two b regions and emitted by the near infrared two regions; and the viroid mesoporous silica is used as a hard template to synthesize the viroid hollow manganese oxide. And further loading a fluorescence quenching molecule IR1064 in the cavity of the viroid hollow manganese oxide. Then modifying the rare earth nanocrystal through carboxylation reaction, modifying the viroid hollow manganese oxide through amination reaction, and finally modifying the rare earth nanocrystal to the viroid hollow manganese oxide surface through condensation reaction between amino-carboxyl to construct the composite probe which can be used for surgical navigation and postoperative photodynamic therapy. Because manganese oxide has acid environment responsiveness, chemical power treatment of a transfer range can be realized, and meanwhile manganese ions can be used for nuclear magnetic resonance imaging and the fluorescence of the released rare earth nanocrystalline can be recovered. The material has the advantages of high tissue penetrability, no background self-fluorescence interference, high resolution and the like, so that the material can be used for early diagnosis of clinical tumors, fluorescence imaging of near infrared two areas of the tumors, navigation tumor excision and chemodynamic therapy for guiding metastasis.

(2) The invention synthesizes the rare earth nanocrystalline which is excited by the near infrared two-b region, emitted by the near infrared two region and the red light, monodisperse and uniform in size by utilizing the hydrothermal solvothermal method through doping adjustment, thereby realizing high-penetration and high-resolution imaging of tumor tissues and photodynamic therapy of tumor metastasis.

(3) The viroid manganese oxide nano-particles synthesized by the invention can be phagocytized by tumor cells in a large quantity and degrade in the slightly acidic environment of the tumor cells to release singlet oxygen and manganese ions, so that the viroid manganese oxide nano-particles can be used for chemo-dynamic treatment and nuclear magnetic resonance imaging of tumors to realize accurate treatment of the tumors.

(4) The synthesized viroid hollow manganese oxide loaded near infrared two-b-region excited rare earth nanocrystalline composite probe can realize fluorescence imaging of tumor microenvironment response, navigation of tumor excision and photodynamic therapy of metastasis so as to thoroughly remove the tumor, thereby improving the postoperative survival rate of patients and providing reference for clinical treatment of malignant tumor.

Drawings

FIG. 1 shows A, B as the cores (. Beta. -NaErF) of near infrared two-b region-excited rare earth nanocrystals obtained in example 1 ₄ 2% Ho) and beta-NaErF ₄ :2％Ho@NaYF ₄ Transmission electron microscopy of nanocrystal core shell structures.

In fig. 2, A, B is a transmission electron microscope image of the virus silicon nanoparticle and the viroid hollow manganese oxide prepared in example 2, respectively.

A, B in FIG. 3 are prepared according to example 3Is prepared by modifying the surface of the viroid hollow manganese oxide loaded with IR1064 with near infrared two b-zone excited rare earth nanocrystalline (MnO) ₂ -IR1064@β-NaErF ₄ :2％Ho@NaYF ₄ ) Is excited by the near infrared two b regions (beta-NaErF) ₄ :2％Ho@NaYF ₄ Core-shell structure and composite probe (MnO) ₂ -IR1064@β-NaErF ₄ :2％Ho@NaYF ₄ ) Is a fluorescent spectrum of (3).

FIG. 4 shows a virus-like manganese oxide particle-loaded nanocrystalline composite probe (MnO) obtained in example 3 ₂ -IR1064@β-NaErF ₄ :2％Ho@NaYF ₄ ) Transmission electron microscopy images after incubation for different times at ph=5.5 and 7.0, respectively.

FIG. 5 shows a rare earth nanocrystalline composite probe (MnO) loaded with viroid manganese oxide particles prepared in example 3 ₂ -IR1064@β-NaErF ₄ :2％Ho@NaYF ₄ ) And a comparison of the fluorescence imaging of clinically approved ICG penetration depths under different laser shots.

Detailed Description

The technical scheme of the invention will be further described in detail below with reference to specific embodiments. It is to be understood that the following examples are illustrative only and are not to be construed as limiting the scope of the invention. All techniques implemented based on the above description of the invention are intended to be included within the scope of the invention.

Unless otherwise indicated, the starting materials and reagents used in the following examples were either commercially available or may be prepared by known methods.

Example 1

β-NaErF ₄ :2％Ho@NaYF ₄ Preparing core-shell structure nanocrystals:

first, er (CH) in a molar ratio of 49:1 was added ₃ CO ₂ ) ₃ ·4H ₂ O(408.123mg)，Ho(CH ₃ CO ₂ ) ₃ ·4H ₂ O (9.000 mg), total 1mmol of rare earth raw material, 6mL of oleic acid and 15mL of octadecene are sequentially added into a 100mL three-necked flask and uniformly mixed, the mixture is heated to 140 ℃ under vacuum condition, and the mixture is stirred and maintained for 30min, so that water and oxygen in the system are removed. Finally coolCooled to room temperature to give a clear and transparent solution. 4mmol of ammonium fluoride and 2.5mmol of sodium hydroxide are respectively dissolved in 5mL of methanol, the clear and transparent solution is added after rapid mixing, the solution is kept for 1h at 50 ℃ for full stirring and nucleation, then the temperature is raised to 70 ℃, and the solution is kept for 60min under vacuum (redundant methanol, oxygen and water molecules are removed). Subsequently, high purity argon was introduced into the solution system while heating to 300 ℃ (heating rate 10 ℃/min) and holding for 60min. (magnetic stirring was maintained throughout the preparation). And finally, cooling the reacted system to room temperature, adding 15mL of absolute ethyl alcohol for precipitation, and centrifuging at a high speed to obtain a product. The NaErF obtained was then subjected to ₄ Washing 2% Ho nanocrystalline with absolute ethanol, centrifuging at 3000rpm for 10min, dispersing the separated solid product in cyclohexane, and freeze preserving at concentration of 0.1mol/L.

Preparation of beta-NaErF by continuous layer-by-layer protocol (one-pot successive layer) ₄ :2％Ho@NaYF ₄ Nanocrystalline with core-shell structure:

synthesis of Y-OA (0.1M) precursor: 5.0mmol of YCl ₃ Oleic acid (20.0 mL), octadecene (30.0 mL) was added to a 100mL three-necked flask and mixed, the temperature was raised to 140℃under vacuum, and stirring was maintained for 60min, keeping the system in a highly anhydrous oxygen environment. Then cooling the synthesized ligand to obtain clear and transparent Y-OA complex precursor solution (0.1 mol/L);

synthesis of Na-TFA-OA (0.4M) precursor: adding 16.0mmol sodium trifluoroacetate and 40mL oleic acid into a 100mL three-necked flask, mixing, then placing under room temperature condition, vacuumizing and stirring uniformly until the solution is completely dissolved to obtain a pale yellow transparent precursor oleic acid solution;

core-shell structure beta-NaErF ₄ :2％Ho@NaYF ₄ Is prepared from the following steps: firstly, adding 4.0mL of oleic acid and 6mL of octadecene into a 50mL three-necked flask in sequence and uniformly mixing, and then adding 2.5mL of the synthesized beta-NaErF into the mixed solution ₄ A cyclohexane solution (0.25 mmol) of 2% Ho. Cyclohexane was removed from the system under vacuum, argon was introduced into the three-necked flask, and the temperature was raised to 280 ℃ (the rate of rise of temperature was 20 ℃/min). Then alternately adding yttrium-oleic acid (Y-OA) (0.10 mol/L,1.0 mL) and sodium trifluoroacetate-oleic acid (Na-TFA-OA) (0.40 mol/L,0.50 mL) precursor solutions, wherein the dripping time interval between the two precursors is 15min, and the nano particles coated with a shell layer are prepared after the dripping of the two precursors is completed; the total of three layers (i.e., repeatedly and alternately adding yttrium-oleic acid (Y-OA) (0.10 mol/L,1.0 mL) and sodium trifluoroacetate-oleic acid (Na-TFA-OA) (0.40M, 0.50 mL) precursor solutions) was coated three times, and the coating time interval between each layer was 15min, so the shell thickness could be controlled by the number of groups. After the reaction, the reaction mixture was cooled to room temperature to obtain beta-NaErF ₄ :2％Ho@NaYF ₄ The nanocrystals were separated by centrifugation at 3000rpm for 10min with absolute ethanol, and the solid product obtained by separation was dispersed in cyclohexane for storage by freezing.

FIG. 1 shows the core (. Beta. -NaErF) of near infrared two-b zone-excited rare earth nanocrystals prepared in this example ₄ 2% Ho nanocrystals) and core-shell structures (beta-NaErF ₄ :2％Ho@NaYF ₄ ) Is a transmission electron microscope image of (a). As can be seen from the figure, the nanocrystals of the core layer and the core shell layer each have a β -shaped structure with a uniform structure; and the particle size is uniform, wherein the particle size of the core layer is about 17.52+/-0.43 nm, and the particle size of the rare earth nanocrystalline with the core-shell structure is about 24.69 +/-0.52 nm.

Example 2

Preparation of the virus-like hollow mesoporous manganese oxide particles loaded with IR 1064:

(1) Synthesizing virus-like silicon mesoporous nano particles by a two-phase method;

60mL of ultrapure water and 1.5g of cetyltrimethylammonium bromide (CTAB) were placed in a 100mL flask, and after dissolution, the mixture was stirred at 60℃for 0.5 hour, and then 0.75mL of 25% triethanolamine was added. After 0.5h, 16mL of cyclohexane and 4mL of tetraethyl orthosilicate mixed solution are added, after 48h of reaction, precipitation is obtained through centrifugation at 10000rpm for 10min, and then the viroid silicon mesoporous nano particles are obtained through three times of washing with water and ethanol.

(2) Preparation of viroid hollow mesoporous manganese oxide

Dispersing 100mg of viroid silicon mesoporous nano particles prepared in the step (1) in 50mL of deionized water by ultrasonic, and then adding 0.09g of Mn (N)O ₃ ) ₃ ·6H ₂ O, and stirred in an oil bath at 90℃for 0.5h at 600 rpm. Then, 0.09g of urotropine was added thereto, and the reaction was continued with stirring for 2 hours. After the reaction is finished, the product is collected by centrifugal separation and washed three times with water and ethanol. Then etching in alkaline solution to remove the silicon dioxide template, wherein the specific operation is as follows: dispersing the reaction product into 2mol/L NaOH aqueous solution, standing for 24 hours in a 60 ℃ oven, then washing with water and ethanol for three times respectively to obtain the final product virus-like hollow mesoporous manganese oxide particles, and drying for later use.

(3) Preparation of IR 1064-loaded viroid hollow manganese oxide

20mg of the viroid hollow mesoporous manganese oxide particles are dispersed in 20mL of water, then 5mg of IR1064 is added, stirring is carried out for 24 hours at room temperature, and the excess IR1064 is removed by centrifugation at 3000rpm for 5 minutes, so that the viroid hollow manganese oxide particles loaded with the IR1064 are successfully obtained.

FIG. 2 is a transmission electron microscope image of the viroid silicon mesoporous nano-particles prepared in the embodiment and the viroid hollow mesoporous manganese oxide synthesized based on the same. As can be seen from the figure, the invention successfully prepares the viroid mesoporous silica structure, the inside of the structure is a mesoporous structure, the surface of the structure grows cluster tubules, the whole structure is viroid structure, and meanwhile, the mesopores and the tubular body can be used for loading drug molecules; the viroid hollow mesoporous manganese oxide prepared by the subsequent hard template method successfully replicates the structure of viroid mesoporous silicon oxide, is similar to a virus shell, and has the functions of rapid cell infection and drug loading.

Example 3

Preparation of a composite probe of a rare earth nanocrystalline excited by a near infrared two b region modified on the surface of hollow virus manganese oxide loaded with IR 1064:

(1) Oleic acid coated beta-NaErF prepared in example 1 ₄ :2％Ho@NaYF ₄ Rare earth nanocrystals (0.1 mmol) were dispersed in 5mL chloroform, then 1mL containing 25mg DSEP-PEG was added ₂₀₀₀ -chloroform solution of COOH. After the mixed solution was stirred in a glass bottle for 24 hours, chloroform was spontaneously evaporated under an air atmosphere. After the chloroform is evaporated, the glass bottle is placed in a 50 ℃ oven0.5h to facilitate further evaporation of chloroform. Finally, 5mL of deionized water is added into the hydrophilic carboxyl phospholipid modified rare earth nanocrystalline particles, and the excessive carboxyl phospholipid is washed at least three times by an ultracentrifuge (17500 rpm,30 min) through ultrasonic treatment. Some large agglomerates present after the centrifugal washing can be removed by a 0.22 μm sieve. Finally, carboxyl functionalized rare earth nanocrystalline dispersed in water can be obtained;

(2) Dispersing 100mg of the virus-like hollow manganese oxide nanoparticle loaded with IR1064 prepared in example 2 in 50mL of ethanol, stirring in an oil bath at 80 ℃ for 0.5h, adding 100 mu L of aminosilane APTES (3-amino-propyl) -triethoxysilane), reacting for 12h, collecting samples, and washing with ethanol and water for three times at 3000rpm and 10min respectively to obtain amino-modified virus-like hollow manganese oxide particle loaded with IR 1064;

(3) Condensing carboxyl functionalized rare earth nanocrystalline obtained in step (1) and amino modified hollow virus-like manganese oxide particles loaded with IR1064 obtained in step (2) (mass ratio is 3:1) in a solution containing 8mg of 1- (3-dimethylaminopropyl) -3-ethylcarbodiimide hydrochloride (EDC) and 10mg of N-hydroxysuccinimide (NHS) and pH=8.5 for 12 hours to obtain the composite probe (MnO) loaded with the near infrared two b region excited rare earth nanocrystalline and modified on the surface of the hollow virus-like manganese oxide loaded with IR1064 ₂ -IR1064@β-NaErF ₄ :2％Ho@NaYF ₄ )。

FIG. 3 shows A, B is a schematic diagram of IR1064 loaded hollow manganese oxide surface modified near infrared two b region excited rare earth nanocrystalline (MnO) prepared according to this example ₂ -IR1064@β-NaErF ₄ :2％Ho@NaYF ₄ ) Is excited by the near infrared two b regions (beta-NaErF) ₄ :2％Ho@NaYF ₄ Core-shell structure) and composite probes (MnO) ₂ -IR1064@β-NaErF ₄ :2％Ho@NaYF ₄ Core-shell structure). From the graph A, it can be observed that the rare earth nanocrystalline can be successfully attached to the surface of the viroid hollow manganese oxide, thereby proving the reliability of the amino carboxyl condensation reaction of the invention. From graph B, it can be seen that: the rare earth nanocrystalline prepared by the invention can excite light at 660nm and near infrared at 1532nmHas strong emission; in the composite probe, the viroid hollow manganese oxide is internally loaded with IR1064 and has a strong light absorption effect, so that fluorescence of the rare earth nanocrystalline is quenched.

FIG. 4 shows the virus-like manganese oxide particle-loaded rare earth nanocrystalline (MnO) ₂ -IR1064@β-NaErF ₄ :2％Ho@NaYF ₄ ) Incubation at ph=5.5 and 7.0 for 0 min, centrifugation after 30 min and 120 min gave a transmission electron micrograph of the precipitate. The results in the figures show that: the composite probe of the invention can degrade viroid hollow manganese oxide in the slightly acidic environment of tumor to release beta-NaErF ₄ :2％Ho@NaYF ₄ Rare earth nanocrystalline and IR1064, beta-NaErF ₄ :2％Ho@NaYF ₄ Fluorescence of the rare earth nanocrystals is not absorbed by IR1064, fluorescence recovery can be used for imaging, and released manganese ions can be used for chemo-kinetic treatment of tumor metastases.

Tissue penetration experiment: the 20% fat emulsion was diluted to 1% with water for use. Taking a hexagonal small dish, and respectively adding beta-NaErF with the same fluorescence intensity into the small dish ₄ :2％Ho@NaYF ₄ Rare earth nanocrystals and ICG solutions. Taking a culture dish with the diameter of 3.5cm, and placing the hexagonal small dishes with the samples at the bottoms of the culture dishes respectively, and fixing the hexagonal small dishes with the samples by using adhesive tapes. Will contain beta-NaErF ₄ :2％Ho@NaYF ₄ The culture dish of rare earth nanocrystalline is placed under an InGaAs imager, excited at 1532nm, filtered at 880nm, and photographed at 20mm exposure. Fat emulsions of 1% concentration were added dropwise to different thicknesses (0 mm, 2mm, 4mm, 6mm, 8mm, 10 mm) with a pipette, and photographed until no hexagonal dish was observed.

Setting ICG as a control group, exciting at 808nm, filtering at 880nm, and exposing and photographing at 20 mm; the above experiment was repeated. Analysis of signal to noise ratio by image J software gives a two b-region excited beta-NaErF ₄ :2％Ho@NaYF ₄ Rare earth nanocrystals have advantages in terms of penetrability and signal-to-noise ratio over clinically approved ICG.

FIG. 5 shows the virus-like manganese oxide particle-loaded rare earth nanocrystalline (MnO) ₂ -IR1064@β-NaErF ₄ :2％Ho@NaYF ₄ ) And clinically approved ICG penetration depth (steps such asBelow). From the figure, the virus-like manganese oxide particle loaded rare earth nano probe (MnO) excited by the two b regions prepared by the invention ₂ -IR1064@β-NaErF ₄ :2％Ho@NaYF ₄ ) The penetration depth of (2) can reach 10mm; compared with the traditional clinical comparative ICG with the penetrability of 6mm, the virus-like manganese oxide particle loaded rare earth nanocrystalline (MnO) excited by the two b regions prepared by the invention ₂ -IR1064@β-NaErF ₄ :2％Ho@NaYF ₄ ) Has remarkable advantages in tissue penetration.

The embodiments of the present invention have been described above. However, the present invention is not limited to the above embodiment. Any modification, equivalent replacement, improvement, etc. made within the spirit and principle of the present invention should be included in the protection scope of the present invention.

Claims

1. The rare earth nanocrystalline is characterized by comprising a carrier and rare earth core-shell nano particles loaded on the surface of the carrier;

the carrier is connected with the rare earth core-shell nano particles through valence bonds; the rare earth nanocrystalline can be selectively degraded to release Mn in the weak acidic microenvironment of tumor ²⁺ ；

The carrier is hollow manganese oxide modified by amination;

the cavity of the carrier is loaded with a fluorescence quenching molecule IR1064.

2. The rare earth nanocrystalline according to claim 1, wherein the mass ratio of the carrier to the rare earth core-shell nanoparticles in the rare earth nanocrystalline is 1 (1-2); and/or the carrier has an amino group that forms an amide bond with a carboxyl group in the rare earth core-shell nanoparticle.

3. The rare earth nanocrystal of claim 1, wherein the compound used for the amination modification is aminosilane APTES.

4. The rare earth nanocrystalline according to claim 1, wherein the loading of the fluorescence quenching molecules is 10-15% of the total weight of the carrier.

5. The rare earth nanocrystal of claim 1, wherein the rare earth core-shell nanoparticle comprises a core-layer nanoparticle, at least one shell layer coating the core-layer nanoparticle, and a polymer modified outside the shell layer and containing carboxyl functionality, wherein the core-layer nanoparticle and the shell layer are each independently selected from AREF ₄ Wherein: a is Na or K, RE is at least one of Er, ho, gd and Y.

6. The rare earth nanocrystal of claim 5, wherein a is Na.

7. The rare earth nanocrystal of claim 5, wherein the RE element in the core nanoparticle and the shell are different.

8. The rare earth nanocrystalline according to claim 7, wherein in the core layer nanoparticle, RE is two of Er, ho; in the shell nanoparticle, RE is Y.

9. The rare earth nanocrystal of claim 8, wherein the core nanoparticle has a Ho doping level of 1 to 5%.

10. The rare earth nanocrystalline according to claim 5, wherein the particle diameter of the rare earth core-shell nanoparticle is 24 to 25.5nm.

11. The rare earth nanocrystalline according to claim 5, wherein the ratio of the particle diameter of the core layer nanoparticle to the thickness of the shell layer is 1 (1-1.5).

12. The rare earth nanocrystal of claim 5, wherein the core nanoparticle has a particle size of 17 to 18nm.

13. The rare earth nanocrystalline according to claim 5, wherein the polymer containing carboxyl functional groups is a PEG polymer, a small molecule acid, or a high molecule polymer containing carboxyl groups.

14. The rare earth nanocrystalline according to claim 13, wherein the polymer containing carboxyl functional groups is phospholipid polyethylene glycol (DSEP-PEG) ₂₀₀₀ -COOH), maleamic acid, and polyacrylic acid.

15. The method for preparing rare earth nanocrystalline according to any one of claims 1 to 14, characterized in that the method comprises reacting a carrier with rare earth core-shell nanoparticles to prepare the rare earth nanocrystalline.

16. The method of claim 15, wherein the mass ratio of rare earth core-shell nanoparticles to carrier is 2-5:1.

17. The method of preparing the rare earth core-shell nanoparticle of claim 15, comprising the steps of:

(2) Adding RE-OA and A-TFA-OA precursor solution into the product of the step (1) to react, so as to prepare the rare earth core-shell nanoparticle;

wherein A is Na or K, RE is at least one of Er, ho, gd and Y; OA is oleic acid; TFA is trifluoroacetic acid.

18. The method of claim 17, wherein step (2) is performed at least once to obtain nanoparticles having at least one shell layer.

19. The method of claim 17, wherein RE is Y.

20. The method of claim 17, wherein the alkali metal fluoride solution is NaF, NH ₄ One of the solutions of F and KF.

21. The method according to claim 17, wherein in the step (1), the molar ratio of the rare earth ion and the alkali metal fluoride in the rare earth salt is 1 (2-5).

22. The method of claim 17, wherein the rare earth ions in the rare earth salt are at least one of Er, ho, and Y.

23. The method of claim 22, wherein the rare earth ions in the rare earth salt are two of Er and Ho.

24. The preparation method of claim 23, wherein the molar ratio of the amount of Er to the amount of Ho is (48-49.5): 0.5-2.

25. The method of claim 17, further comprising dispersing the core nanoparticles in a solvent to obtain a core nanoparticle dispersion in step (1).

26. The method of claim 25, wherein the solvent is cyclohexane.

27. The method of claim 25, wherein the core nanoparticle dispersion has a concentration of 0.05 to 0.2mol/L.

28. The method of claim 17, wherein in step (1), the rare earth salt is a rare earth chloride, a rare earth nitrate, or a rare earth acetate.

29. The method of claim 17, wherein in step (1), the alkaline solution is a sodium hydroxide solution or a potassium hydroxide solution.

30. The method of claim 17, wherein the solvent used in the alkali metal fluoride solution and the alkaline solution is methanol.

31. The method according to claim 17, wherein in the step (1), the total amount of the rare earth salt is 0.5 to 2 mmol.

32. The method of claim 17, wherein in step (1), the ratio of oleic acid to octadecene is 1 (2-3).

33. The method according to claim 17, further comprising heating and stirring the mixed solution of oleic acid and octadecene after adding rare earth salt to remove water and oxygen in the system in step (1).

34. The method according to claim 33, wherein the temperature of the heating and stirring is 130 to 150 ℃, and the time of the heating and stirring is 20 to 40 minutes.

35. The method of claim 17, further comprising, in step (1), stirring the mixed solution after the alkali metal fluoride solution and the alkaline solution are added to nucleate.

36. The method of claim 35, wherein the stirring nucleation is performed by a two-stage heating process.

37. The method of claim 36, wherein in the two-stage heating mode:

the temperature of the first-stage heating is 40-60 ℃, and the time of the first-stage heating is 0.5-2 h;

The temperature of the second-stage heating is 60-80 ℃, and the time of the second-stage heating is 0.5-2 h.

38. The method of claim 25, wherein in step (2), the core layer nanoparticle dispersion is added to a mixed solution of oleic acid and octadecene prior to adding RE-OA and a-TFA-OA precursor solutions to the product of step (1).

39. The method of claim 38, wherein the volume ratio of the core nanoparticle dispersion to the oleic acid and octadecene mixed solution is 1 (1-3).

40. The process according to claim 39, wherein the volume ratio of oleic acid to octadecene in the mixed solution of oleic acid and octadecene is 1 (1-2).

41. The method according to claim 17, wherein in the step (2), the concentration of the RE-OA precursor solution is 0.05 to 0.2mol/L.

42. The method according to claim 17, wherein in the step (2), the concentration of the a-TFA-OA precursor solution is 0.3 to 0.5 mol/L.

43. The method of preparing as claimed in claim 25, wherein in step (2), the RE-OA precursor solution and the a-TFA-OA precursor solution are added to the core layer nanoparticle dispersion liquid prepared in step (1) in an alternately spaced manner.

44. The method of claim 43, wherein the RE-OA precursor solution and a-TFA-OA precursor solution are added alternately at least once each.

45. The process according to claim 17, wherein in the steps (1) and (2), the temperatures of the reactions are the same or different and are 200 to 400 ℃ independently of each other, and the reaction time is 40 to 80 min.

46. The process according to claim 45, wherein in the steps (1) and (2), the temperature rise rate of the reaction is 5 to 20℃per minute.

47. The method of manufacturing of claim 17, further comprising step (3): uniformly mixing the nano particles coated with at least one layer of shell layer obtained in the step (2) with a polymer containing carboxyl functional groups in an organic solvent, standing until the organic solvent volatilizes, adding water for dispersion, and centrifugally separating the carboxyl modified rare earth core-shell nano particles.

48. The process according to claim 47, wherein in the step (3), the organic solvent is chloroform or cyclohexane or n-hexane or tetrahydrofuran.

49. The process according to claim 48, wherein in step (3), the organic solvent is chloroform.

50. The method of claim 47, wherein in step (3), the ratio of the nanoparticles coating at least one shell layer to the polymer having carboxyl functional groups is 0.1 mmol (20-30) mg.

51. The method of claim 15, wherein the method of preparing the carrier comprises reacting a viroid silicon mesoporous nanoparticle with a manganese salt and urotropine to remove the viroid silicon mesoporous nanoparticle template by alkali treatment, thereby obtaining the viroid hollow manganese oxide.

52. The method of claim 51, wherein the virus-like silicon mesoporous nanoparticle is prepared by reacting a raw material comprising tetraethyl orthosilicate with cetyltrimethylammonium bromide (CTAB), triethylamine.

53. The method according to claim 52, wherein the ratio of tetraethyl orthosilicate to cetyltrimethylammonium bromide (CTAB) to triethylamine is 5.3 mL/2 g/1 mL.

54. The method according to claim 51, wherein the mass ratio of the viroid silicon mesoporous nano-particles to manganese salt and urotropine is 10:9:9.

55. The method according to claim 51, wherein the reaction temperature of the viroid silicon mesoporous nano-particles with manganese salt and urotropine is 80-100 ℃ and the reaction time is 3-5 h.

56. The method of claim 51, further comprising reacting the washed reaction product with a compound having an amino function to produce the virus-like hollow manganese oxide having an amino function modified on the surface.

57. The method of claim 56, further comprising reacting the above-mentioned hollow manganese oxide with a fluorescent quenching molecule to obtain hollow manganese oxide with amino groups.

58. The method according to claim 57, wherein the mass ratio of the virus-like hollow manganese oxide modified with amino functional groups to the fluorescence quenching molecule is (3-5): 1.

59. The method of claim 51, wherein the method of preparing the carrier comprises the steps of:

cetyl Trimethyl Ammonium Bromide (CTAB) is dissolved in water, and then mixed solution of triethylamine, cyclohexane and tetraethyl orthosilicate is added to prepare the viroid silicon mesoporous nano-particles through reaction;

(S2) preparation of viroid hollow manganese oxide

Adding Mn (NO) into the viroid silicon mesoporous nano particle dispersion liquid under the condition of heating and stirring ₃ ) ₂ ·6H ₂ O and urotropine, after reaction, centrifugally washing, and finally etching off the viroid mesoporous silicon nanoparticle template in NaOH aqueous solution to prepare viroid hollow manganese oxide; then, the viroid hollow manganese oxide reacts with a compound containing amino functional groups to prepare the viroid hollow manganese oxide with the amino functional groups modified on the surface;

And (3) dissolving the viroid hollow manganese oxide particles prepared in the step (S2) in water, then adding IR1064, stirring at room temperature for reaction, and centrifuging to remove the redundant IR1064, thereby preparing the viroid hollow manganese oxide particles loaded with the IR 1064.

60. The method of any one of claims 15-59, wherein the reacting the support with the rare earth core shell nanoparticle further comprises adding an activator.

61. The method of claim 60, wherein the activator is at least one of carbodiimide (EDC), N-hydroxysuccinimide (NHS), and Dimethylacetamide (DMAC).

62. The method of claim 60, wherein the activating with the activator is performed in a solution having a pH of 8.0 to 9.0.

63. The method of claim 60, wherein the mass ratio of rare earth core-shell nanoparticles to activator is 1:3-5.

64. A composite probe comprising the rare earth nanocrystal of any one of claims 1 to 14.

65. Use of the rare earth nanocrystalline according to any one of claims 1 to 14 or the composite probe according to claim 64 for the preparation or as a contrast agent in the field of surgical navigation, postoperative photodynamic therapy, fluorescence guidance of postoperative metastases, nuclear magnetic resonance imaging of tumors.