阅读说明：本技术 一种近红外光控制按需供氧纳米诊疗剂及其制备方法和应用 (Near-infrared light controlled oxygen supply on-demand nano diagnosis and treatment agent and preparation method and application thereof ) 是由 刘湘梅 田康 吴昊明 周严莉 赵强 黄维 刘淑娟 于 2019-06-04 设计创作，主要内容包括：本发明公开了一种近红外光控制按需供氧纳米诊疗剂及其制备方法和应用。该纳米诊疗试剂采用近红外光敏剂IR780和罗丹明B同时负载得到二氧化硅纳米复合材料,然后再通过聚乙烯吡咯烷酮修饰负载双氧水,得到生物相容性好、粒径小且均匀、稳定性好、并能按需供氧的纳米诊疗剂,并测试其性能。该纳米诊疗剂有利于癌细胞以及肿瘤的自供氧光动力治疗,其具有制备工艺简单、成本低以及适用范围广等技术优势,因此在生物医药、化工工业、功能材料等领域具有重要的应用前景。(The invention discloses a near infrared light control oxygen supply on-demand nano diagnosis and treatment agent, and a preparation method and application thereof. The nano diagnosis and treatment reagent adopts near infrared photosensitizer IR780 and rhodamine B to load simultaneously to obtain a silicon dioxide nano composite material, then polyvinyl pyrrolidone is used for modifying and loading hydrogen peroxide to obtain the nano diagnosis and treatment reagent which has good biocompatibility, small and uniform particle size and good stability and can supply oxygen according to needs, and the performance of the nano diagnosis and treatment reagent is tested. The nano diagnosis and treatment agent is beneficial to the self-oxygenation photodynamic therapy of cancer cells and tumors, has the technical advantages of simple preparation process, low cost, wide application range and the like, and has important application prospect in the fields of biological medicine, chemical industry, functional materials and the like.)

1. The near infrared light controlled oxygen supplying nanometer diagnosis and treatment agent features that near infrared dye and fluorescent dye are mixed and loaded inside nanometer silica material, and nanometer silica material with polyvinyl pyrrolidone and hydrogen peroxide loaded successively is prepared.

2. The near-infrared controlled oxygen-on-demand nano diagnostic and therapeutic agent as claimed in claim 1, wherein the near-infrared dye is near-infrared photosensitizer IR 780.

3. The near-infrared controlled oxygen-supply-on-demand nano diagnostic and treatment agent as claimed in claim 1, wherein the fluorescent dye is rhodamine B.

4. The near-infrared controlled oxygen-supply-on-demand nano diagnostic and therapeutic agent as claimed in claim 1, wherein the particle size of the silica nano material is 60 to 80 nm.

5. The near-infrared light controlled oxygen supply on-demand nano diagnostic and therapeutic agent as claimed in claim 1, wherein the molar concentration of the hydrogen peroxide is 5.9mol/L to 10.58 mol/L.

6. The near-infrared controlled oxygen-supply-on-demand nano diagnostic and therapeutic agent as claimed in claim 1, wherein the polyvinylpyrrolidone has a relative number average molecular weight of 28K to 40K.

7. The preparation method of the near infrared light controlled oxygen supply on demand nano diagnosis and treatment agent as claimed in any one of claims 1 to 6, characterized by comprising the following steps:

s1: dissolving 20-30 mg of near-infrared photosensitizer IR780 and 8-12 mg of rhodamine B in absolute ethyl alcohol at room temperature, and stirring for 25-45 min;

s2: slowly dripping 70-100 uL of tetraethoxysilane into the solution prepared in the step S1 at room temperature, and stirring for 20-30 min;

s3: rapidly adding 3-5 mL of ammonia water into the solution prepared in the step S2 at room temperature, reacting for 20-24 h at a stirring speed of 600r/min, centrifugally washing the product for 3 times by using ultrapure water, and drying to obtain a nano particle solid product;

s4: dissolving 4mg of the nanoparticle solid product prepared in the step S3 in 2mL of ultrapure water at room temperature, dissolving 16-24 mg of polyvinylpyrrolidone in 2mL of ultrapure water, dropwise adding the nanoparticle solution into the ultrapure water solution of polyvinylpyrrolidone, and stirring for 8-12 h;

s5: and (5) dispersing the product obtained in the step S4 into a hydrogen peroxide solution at room temperature, and stirring for 3-6 hours to obtain a final product.

8. The method for preparing the near-infrared light controlled oxygen supply-on-demand nano diagnosis and treatment agent according to claim 7, wherein the mass ratio of the near-infrared photosensitizer IR780 to the rhodamine B in step S1 is 5: 2.

9. the method according to claim 7, wherein the mass ratio of the nanoparticle solid product to the polyvinylpyrrolidone in step S4 is 1: 4 to 6.

10. The use of the near-infrared controlled on-demand oxygen supply nano diagnostic and therapeutic agent as claimed in any one of claims 1 to 6, characterized in that as a photodynamic material, a photothermal material or a fluorescent material, in the self-oxygen supply photodynamic therapy or photothermal therapy guided by fluorescence imaging of cancer cells or tumor cells.

Technical Field

The invention belongs to the technical field of preparation technology of nano materials and special materials, and particularly relates to a near-infrared light controlled oxygen supply on-demand nano diagnosis and treatment agent, and a preparation method and application thereof.

Background

Cancer was an unfamiliar vocabulary for Chinese people half a century ago. However, for as short as fifty years, cancer has become one of the most closely related diseases, and both morbidity and mortality are in the third half of the world. The prediction of traditional Chinese cancer statistics in 2015, which is completed by the national cancer center, namely Hijic, is that 429.2 ten thousand cancer cases, cancer death cases or more than 281.4 ten thousand are newly added in 2015 in China.

Hypoxia is a hallmark type characteristic of solid tumors, but this is contrary to the conditions of efficient PDT, so the hypoxic environment of tumors is a major factor in limiting solid tumors. In addition, oxygen is consumed during photodynamic therapy to further exacerbate the hypoxia condition of tumor tissues, and the hypoxia condition of the tumor persists. Blood vessels in tumor micro-regions can be damaged during the photodynamic therapy process, and the damage of the blood vessels causes oxygen transportation obstacle to further aggravate the hypoxia. To date, researchers have explored ways to improve oxygen concentration that affects photodynamic therapy, such as increasing blood vessel density, dilating blood vessels, and increasing oxygen transport. However, the experimental results show that the endogenous hydrogen peroxide content at the tumor tissue is not enough to provide the oxygen required for the treatment, and the exogenous oxygen is difficult to transport to the tumor tissue.

Patent CN109395081A discloses a nano-photodynamic reagent capable of near-infrared light excitation and self-oxygen supply, which adopts near-infrared photosensitizer IR780 with photodynamic effect and photothermal effect as photothermal and photodynamic treatment reagent, adopts small-sized sea urchin-shaped manganese dioxide nano-material as nano-platform, and adopts bovine serum albumin for coating; the composite nano material improves the oxygen concentration through the reaction of the manganese dioxide nano particles and the endogenous hydrogen peroxide of the tumor. Although the patent can relieve the hypoxic environment of the tumor tissue, the patent cannot provide oxygen for the tumor tissue in a fixed-point and timed mode.

Disclosure of Invention

The invention aims to solve the problems and provides a near-infrared light controlled oxygen supply on demand nano diagnosis and treatment agent, which can improve the biocompatibility of a near-infrared photosensitizer, can supply oxygen to the nano diagnosis and treatment agent on demand to improve the problem of tissue hypoxia, and can realize deep photodynamic therapy through effective delivery of the near-infrared photosensitizer.

In order to achieve the purpose, the invention adopts the technical scheme that:

a near-infrared light controlled oxygen supply-on-demand nano diagnosis and treatment agent is a nano material which is prepared by loading a near-infrared dye and a fluorescent dye in a silicon dioxide nano material through in-situ blending, and sequentially loading polyvinylpyrrolidone and hydrogen peroxide on the surface of the silicon dioxide nano material.

Preferably, the near-infrared dye is a near-infrared photosensitizer IR 780;

preferably, the fluorescent dye is rhodamine b (rb).

Preferably, the particle size of the silicon dioxide nano material is 60-80 nm.

Preferably, the polyvinylpyrrolidone has a relative number average molecular weight of 28K to 40K.

Preferably, the concentration of the hydrogen peroxide is 5.9-10.58 mol/L. With addition of sufficient amount of H₂O₂Can provide enough oxygen for the nano diagnosis and treatment agent to supplement endogenous H at tumor tissues₂O₂The content is insufficient; if H is present₂O₂Too high concentration of (A), H supported on the outer layer of PVP₂O₂Will reach saturation, resulting in H₂O₂Waste of (2);if H is present₂O₂Too low a concentration of (b) may result in insufficient oxygen supply at the tumor tissue and failure to achieve therapeutic effect.

The near-infrared photosensitizer IR780 and rhodamine B are loaded in the silica nanoparticle through in-situ doping, and the prepared nanoparticle has the capability of molecular release and structural collapse, which is beneficial to deep tumor penetration of the photosensitizer; and then the PVP and the hydrogen peroxide are sequentially loaded on the surfaces of the nanoparticles to obtain the nano diagnosis and treatment agent. The nano diagnosis and treatment agent not only improves the biocompatibility of the photosensitizer IR780, but also can supply oxygen as required to improve the problem of oxygen lack in tissues, and can realize deep photodynamic therapy through the effective delivery of the photosensitizer. Therefore, the nano diagnosis and treatment agent is beneficial to self-oxygen supply photodynamic therapy and photothermal therapy.

The invention also provides a preparation method of the near infrared light controlled oxygen supply on-demand nano diagnosis and treatment agent, which comprises the following steps:

s1: dissolving 20-30 mg of near-infrared photosensitizer IR780 and 8-12 mg of rhodamine B in absolute ethyl alcohol at room temperature, and stirring for 25-45 min;

s2: slowly dripping 70-100 uL of tetraethoxysilane into the solution prepared in the step S1 at room temperature, and stirring for 20-30 min;

s3: rapidly adding 3-5 mL of ammonia water into the solution prepared in the step S2 at room temperature, reacting for 20-24 h at a stirring speed of 600r/min, centrifugally washing the product for 3 times by using ultrapure water, and drying; obtaining a nano particle solid product;

s4: dissolving 4mg of the nanoparticle solid product prepared in the step S3 in 2mL of ultrapure water at room temperature, dissolving 16-20 mg of polyvinylpyrrolidone in 2mL of ultrapure water, dropwise adding the nanoparticle solution into the ultrapure water solution of polyvinylpyrrolidone, and stirring for 8-12 h;

s5: and (5) dispersing the product obtained in the step S4 in a hydrogen peroxide solution at room temperature, and stirring for 3-6 hours to obtain a final product.

Preferably, in step S1 of the preparation method, the mass ratio of the near-infrared photosensitizer IR780 to the rhodamine B is 5: 2. if the rhodamine B is too low, the rhodamine B is not enough to collapse the silica nanoparticles after being released from the interior of the silica nanoparticles, and the silica nanoparticles cannot be completely decomposed; when the quality of rhodamine B is too high, the amount of doped near-infrared photosensitizer IR780 becomes low, and efficient photodynamic therapy cannot be achieved.

Preferably, in step S4, the mass ratio of the nanoparticle solid product to PVP is 1: 4 to 6. When the content of PVP is too small, the compatibility of the near-infrared photosensitizer IR780 is poor; when the content of PVP is too high, the prepared nano diagnosis and treatment agent composite nano particles have strong cohesiveness.

The invention provides a near-infrared light control oxygen supply on-demand nano diagnosis and treatment agent which is used for photodynamic, photothermal materials and fluorescent materials in self-oxygen supply photodynamic therapy and photothermal therapy guided by fluorescence imaging of cancer cells and tumors.

Compared with the prior art, the invention has the beneficial effects that:

1. the nano diagnosis and treatment agent can improve the photodynamic therapy effect, more importantly, the nano diagnosis and treatment agent controls the fixed-point timed release of oxygen through a photo-thermal conversion material, can be effectively used for diagnosis and treatment of cancer cells and tumors, and has better in-vivo biomedical application prospect;

2. the nano diagnosis and treatment agent is used as a good thermal control oxygen supply intelligent material according to needs, can effectively improve the hypoxic condition of tumors and enhance photodynamic therapy; meanwhile, the problems of biocompatibility and delivery efficiency of the near-infrared photosensitizer IR780 are improved, and meanwhile, the composite nano material improves the problem of penetration depth of light in photodynamic therapy;

3. the composite nano particle has small hydrated particle size, good biocompatibility and high specific surface area, and has good application prospect in visual-guided photodynamic therapy;

4. the functionalized composite nano material has the advantages of rich raw materials, easy preparation and wide application range, and can be used for industrial production.

Drawings

Fig. 1 is a transmission electron microscope photograph of the nano-sized medical agent prepared in example 1;

FIG. 2 is a transmission electron microscope photograph of the nano diagnostic and therapeutic agent prepared in comparative example 1;

FIG. 3 is a transmission electron microscope photograph of the nano diagnostic and therapeutic agent prepared in comparative example 2;

fig. 4 is a hydrated particle size diagram of composite nanoparticles of the nano-diagnostic and therapeutic agent prepared in example 1;

fig. 5 is a uv-vis absorption spectrum of the composite nanoparticle of the nano therapeutic agent prepared in example 1;

FIG. 6 is a graph showing the photothermal power of the composite nanoparticles of the nano diagnostic agent prepared in example 1

Fig. 7 is a fluorescence spectrum of the nano diagnostic and therapeutic agent prepared in example 1 in laser-triggered oxygen release;

FIG. 8 is a fluorescence spectrum of the nano-diagnostic agent prepared in example 1 triggered by oxygen release when heated in a water bath;

FIG. 9 is a singlet oxygen generation diagram under different conditions for the nano-sized therapeutic agent prepared in example 1;

fig. 10 is a diagram showing the generation of singlet oxygen with different concentrations of hydrogen peroxide in application example 8.

Detailed Description

The technical solutions of the present invention will be described clearly and completely with reference to the accompanying drawings, and it is obvious that the described embodiments are only some embodiments of the present invention, not all 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.