CN108295255A - A kind of drug loaded magnetic graphene multifunctional composite and preparation method thereof - Google Patents

Abstract

The invention relates to a drug-loaded magnetic graphene multifunctional composite material and a preparation method thereof. Firstly, the graphene is modified by adding nitro groups and polyethylene glycol functionalization, and then the newly prepared magnetic nanoparticles and drug aqueous solution are uniformly added to the modified graphene aqueous solution, and the three are mixed with each other to form a homogeneous system, and then in the Formation of drug-loaded magnetic graphene multifunctional composites under the action of ultrasound. The preparation method is simple and the conditions are mild, and the obtained drug-loaded magnetic graphene composite material can concentrate the drug concentration in the tumor area under the action of a magnetic field to achieve magnetic targeting; the nitro group on the functionalized modified graphene can Improve the radiosensitivity in the hypoxic environment of the tumor area; the graphene-modified polyethylene glycol molecules can control the slow release of the drug carried, so that the drug is released in the tumor area and reduces the toxicity to normal tissues.

Description

A drug-loaded magnetic graphene multifunctional composite material and preparation method thereof

technical field

The invention belongs to the technical field of preparation of composite materials, in particular to a preparation technology of a drug-loaded magnetic graphene multifunctional composite material.

Background technique

Tumor has always been a worldwide problem that plagues public health. Radiotherapy, chemotherapy, and surgery are currently three important methods for treating tumors. For some tumors that have spread or grown around vital organs, surgery cannot resolve them, and radiation and chemotherapy are required. However, for radiotherapy, high-dose irradiation will cause great damage to normal tissues, and due to the hypoxia inside tumor cells, they generally have radiotherapy resistance, which makes the tumor after radiotherapy have the risk of recurrence. Therefore, looking for safe Effective radiosensitizers for hypoxic cells have become an urgent problem in cancer treatment. The nitro group of nitroimidazoles can be transformed into an amino group through a series of electron releases under low oxygen concentration, thus becoming a recognized radiosensitizer. Among these drugs, metronidazole has entered clinical trials. However, high-dose metronidazole has neurotoxicity, and low-dose metronidazole cannot play the role of sensitization, which limits its use. Therefore, reducing the toxicity of nitroimidazoles and effectively targeting them to the tumor area has become a research direction.

Pentafluorouracil is an anti-metabolite synthesized according to certain assumptions and widely used clinically. It interferes with DNA synthesis by inhibiting the action of thymidine nucleotide synthetase. However, since the drug is quickly distributed to various tissues of the body after being administered by injection, it is preferentially taken up by active division tissues, and can enter the brain tissue through the blood-brain barrier, thus causing mild or severe systemic side effects. Therefore, multifunctional targeting carrier drugs have become a hot spot in the research of radiotherapy and chemotherapy.

The development of nanotechnology has brought new opportunities for tumor treatment. Combining nanomaterials with radiotherapy and chemotherapy drugs can well solve the problem that radiotherapy sensitizers and chemotherapy drugs cannot be enriched in the tumor area. Graphene is a new type of single-layer two-dimensional carbon structure material. It has many excellent properties and has attracted the attention of many fields. It has become a research hotspot in the medical field to apply it as a drug carrier to organisms. The oxidized graphite has multiple groups such as hydroxyl group, carboxyl group, epoxy group, etc., which not only improves water solubility, but also provides a variety of sites for the attachment of other molecules, but graphene is very easy in high-salt solution The reunion has become an obstacle to the application to the biological field. While magnetic nanomaterials are widely used as contrast agents and radiotherapy sensitizers because of their superparamagnetism, high atomic number and low biological toxicity, they are also easy to agglomerate and have little effect and metabolic mechanisms in vivo. Uncertainty limits its clinical application.

If nanomaterials can be compounded with radiotherapy and chemotherapy drugs to maximize their respective advantages while avoiding their respective disadvantages, it will be a major advancement in the application of nanotechnology in the field of tumor treatment.

CN106177981A discloses a preparation method of a magnetic graphene drug-carrying system, using two surfactants to modify graphene oxide, and synthesizing magnetic graphene oxide by hydrothermal synthesis, and then modifying the amphiphilic polymer PF127 and hydrazine hydrate , the anticancer drug paclitaxel was loaded through π-π stacking interaction, and a magnetic graphene drug-loading system integrating anti-tumor, imaging and biocompatibility was obtained. However, this reaction needs to be carried out in a polytetrafluoroethylene-lined reactor at high temperature (190-210° C.) for 11-13 hours, which has certain requirements on equipment and environment. Moreover, the prepared magnetic graphene drug-loading system can produce a good inhibitory effect on tumors only at a concentration of 200ug/ml.

CN107050453A discloses a method for preparing a magnetic nano-targeted graphene oxide drug carrier, which uses graphene oxide, ferric chloride and aniline monomer as raw materials, and is prepared by chemical co-precipitation and in-situ synthesis. The magnetic nano-targeted graphene oxide drug carrier prepared by the method is based on the magnetic targeting and drug loading functions, and has the functions of combining magnetic hyperthermia, photothermotherapy and chemotherapy. However, similar to the above method, the reaction needs to be able to withstand high temperature and high pressure conditions, and there are certain requirements for the experimental environment. In addition, the magneto-induced heating and photo-induced heating are currently only at the experimental level. In the actual treatment In the process, the magnetothermal and photothermal methods have not been realized due to the practical problems of how difficult they are to overcome.

CN104815337A discloses a preparation method of PEG-modified magnetic graphene oxide. At normal temperature and pressure, carboxylated magnetic nanoparticles are mixed with aminated PEGylated graphene oxide, and prepared under EDC condensation conditions. The capacitive magnetic graphene oxide nanomaterial is loaded with the anticancer drug doxorubicin through the π-π stacking effect, and a composite material integrating anti-tumor, imaging and biocompatibility is prepared. However, it is very difficult to collect PEG-Go in the process, and it needs to be centrifuged at 13000rmp for 1h, then the supernatant is discarded, and it can be ultrafiltered with a 100kDa ultrafiltration tube.

Contents of the invention

The purpose of the present invention is to provide a method for preparing a drug-loaded magnetic graphene multifunctional composite material with simple preparation methods, mild preparation conditions, and no special requirements for instruments in view of the current problems in nanomaterials and tumor treatment. The material can integrate magnetic targeting, hypoxic cell sensitization and drug treatment.

The invention provides a method for preparing a drug-loaded magnetic graphene multifunctional composite material, which adopts the following technical scheme:

A preparation method of drug-loaded magnetic graphene multifunctional composite material, comprising the steps of:

1) preparing magnetic nanoparticles;

2) preparing a functionalized graphene material modified with nitroimidazole drugs and polyethylene glycol;

3) preparing an aqueous solution with a certain drug concentration;

4) adding the magnetic nanoparticles obtained in step 1) and the aqueous drug solution prepared in step 3) to the functionalized graphene material aqueous solution prepared in step 2), and mixing;

5) Ultrasonic the mixture obtained in step 4) at room temperature, and then centrifuge to collect the drug-loaded magnetic graphene composite.

Preferably, the magnetic nanoparticles in step 1) include any one of FePt, Fe ₂ O ₃ , Fe ₃ O ₄ , and MnFe ₂ O ₄ nanoparticles, which can be purchased.

Preferably, the magnetic nanomaterial is a FePt magnetic nanomaterial synthesized by a chemical co-reduction method, by combining a platinum source (chloroplatinic acid) and an iron source (iron acetylacetonate) under the action of sodium borohydride at 40° C. for 2 h FePt magnetic nanoparticles with single morphology were obtained.

Preferably, the particle size of the magnetic nanoparticles prepared in step 1) is 2-20 nm.

Preferably, the graphene in step 2) can be ordinary graphene or graphene oxide. If it is ordinary graphene, it needs to be oxidized to graphene oxide by Hummer's method. The final concentration of graphene oxide is preferably 0.1-2mg /ml.

Preferably, the nitroimidazole drug in step 2) is metronidazole.

Preferably, the method for preparing functionalized graphene material in step 2) is: mixing nitroimidazole drugs and polyethylene glycol aqueous solution with graphene oxide aqueous solution, adding carbodiimide and dimethylaminopyridine, stirring at room temperature for 24 ~72h.

Preferably, the dosages of nitroimidazole drugs and polyethylene glycol are 10-50 times that of graphene oxide.

Preferably, the drug in step 3) is a chemotherapeutic drug, such as pentafluorouracil, which can be a powdered drug preparation solution, or a directly purchased injection.

Preferably, the final concentration of the magnetic nanoparticles in the mixed solution formed in step 4) is 0.1-3 mg/ml, and the final concentration of the drug is 0.1-10 mg/ml.

Preferably, step 5) ultrasonication time is 2-8 hours, and centrifugation is performed at 13000 rpm for 3 minutes.

A drug-loaded magnetic graphene multifunctional composite material is prepared by the above-mentioned method.

The application of the above drug-loaded magnetic graphene multifunctional composite material in the treatment of tumor drugs.

The drug-loaded magnetic graphene multifunctional composite material of the present invention uses a step-by-step synthesis method to separately synthesize magnetic nanomaterials and functionalized graphene. The preparation method is simple, the preparation conditions are mild, and there is no special requirement for the instrument. The prepared material Easy to collect. In addition, the drug-loaded magnetic graphene multifunctional composite material of the present invention, on the basis of the functions of magnetic targeting, anti-tumor, and imaging, also has radiosensitization for hypoxic cells due to the modification of nitroimidazole drugs. Therefore, radiotherapy and chemotherapy can be combined to treat tumors in multiple dimensions. Moreover, in the case of a very small dosage (20ug/ml), it exhibits an inhibitory effect on the proliferation of cancer cells.

The graphene multifunctional composite material obtained in the present invention combines the advantages of magnetic targeting, radiotherapy sensitization and chemotherapeutic drugs. Applying this material to tumor treatment can realize the enrichment of radiotherapy and chemotherapeutic drugs in the tumor area under an external magnetic field , to achieve magnetic targeting; the nitro group on the functionalized graphene can improve the radiosensitivity in the hypoxic environment of the tumor area; the graphene-modified polyethylene glycol molecule can control the slow release of the drug carried, The drug is concentrated and released in the tumor area, reducing the toxicity to normal tissues; in the case of low drug concentration, combined radiotherapy and chemotherapy for tumor cells can be achieved, which can greatly reduce the damage to normal tissues.

Description of drawings

The present invention will be further described by using the accompanying drawings, but the embodiments in the accompanying drawings do not constitute any limitation to the present invention.

Fig. 1 is the transmission electron microscope figure that the embodiment of the present invention 1 obtains the graphene multifunctional composite material, can obviously see that graphene wraps magnetic nanoparticles as sheet-like substrate from the figure, and the particle diameter of magnetic nanoparticles is at 3nm about. Metronidazole and pentafluorouracil cannot be displayed on the transmission electron microscope due to their small molecular weights.

Fig. 2 is the X-ray energy spectrogram that the embodiment of the present invention 2 obtains graphene multifunctional composite material, can see C from the figure, N, O, F, Pt, the peak of Fe, prove metronidazole and pentafluorouracil and FePt magnetic nanoparticles attached to graphene.

Fig. 3 is the Fourier transform infrared spectrogram of the graphene multifunctional composite material that the embodiment of the present invention 3 obtains, among the figure 3414 represent the absorption peak of O-H, 2923 and 2852 represent CH respectively and CH2 absorption peak, 1623 represents C=C 1548 represents the absorption peak of NO2, 1261 represents the absorption peak of C-F, and 1204 represents the absorption peak of C-O, which proves that metronidazole and pentafluorouracil are indeed successfully attached to graphene.

Fig. 4 is the X-ray photoelectron spectrum figure of the graphene multifunctional composite material that the embodiment of the present invention 3 obtains, can see in the figure C, N, O, F, Pt, the characteristic peak of Fe, proves metronidazole and pentafluorouracil And FePt magnetic nanoparticles were successfully connected to graphene.

Fig. 5 is a diagram of the dynamic light scattering particle size distribution of the graphene multifunctional composite material obtained in Example 4 of the present invention. It can be seen from the figure that the particle size of the material is concentrated at about 240nm.

Figure 6 shows the results of the inhibition of tumor cell proliferation by the graphene multifunctional composite material obtained in Example 4 of the present invention, with a half-inhibitory concentration of 20ug/ml.

Detailed ways

In order to make the present invention easier to understand, specific embodiments of the present invention will be further described below.

Example 1

1) Synthesize FePt magnetic nanomaterials with a particle size of 3nm by co-reduction method;

2) preparing functionalized graphene modified by metronidazole and polyethylene glycol 4000, the final concentration ratio of metronidazole, polyethylene glycol 4000 and graphene is 20:20:1;

3) Prepare 25mg/ml pentafluorouracil aqueous solution;

4) Mix the magnetic nanoparticles obtained in step 1) and the pentafluorouracil aqueous solution prepared in step 3) with the functionalized graphene aqueous solution prepared in step 2), so that the final concentration of the magnetic nanoparticles is 0.25 mg/ml, and the final concentration of pentafluorouracil The concentration is 1.25mg/ml, and the final concentration of functionalized graphene is 0.15mg/ml;

5) Ultrasonic the mixed solution obtained in step 4) for 2 hours, and centrifuge at 13000 rpm for 3 minutes to collect the material to obtain a graphene multifunctional composite material.

Example 2

1) Synthesize FePt magnetic nanomaterials with a particle size of 3nm by co-reduction method;

2) preparing functionalized graphene modified by metronidazole and polyethylene glycol 4000, the amount ratio of metronidazole, polyethylene glycol 4000 and graphene is 30:15:1;

3) Prepare 30mg/ml pentafluorouracil aqueous solution;

4) Mix the magnetic nanoparticles obtained in step 1) and the pentafluorouracil aqueous solution prepared in step 3) with the functionalized graphene aqueous solution prepared in step 2), so that the final concentration of the magnetic nanoparticles is 2.5mg/ml, and the final concentration of pentafluorouracil The concentration is 8mg/ml, and the final concentration of functionalized graphene is 1mg/ml;

5) Ultrasonic the mixed solution obtained in step 4) for 8 hours, and centrifuge at 13000 rpm for 3 minutes to collect the material to obtain a graphene multifunctional composite material.

Example 3

1) Synthesize FePt magnetic nanomaterials with a particle size of 3nm by co-reduction method;

2) preparing functionalized graphene modified by metronidazole and polyethylene glycol 4000, the final concentration ratio of metronidazole, polyethylene glycol 4000 and graphene is 12:35:1;

3) Prepare 30mg/ml pentafluorouracil aqueous solution;

4) Mix the magnetic nanoparticles obtained in step 1) and the pentafluorouracil aqueous solution prepared in step 3) with the functionalized graphene aqueous solution prepared in step 2), so that the final concentration of the magnetic nanoparticles is 1 mg/ml, and the final concentration of pentafluorouracil 4mg/ml, the final concentration of functionalized graphene is 0.25mg/ml;

5) Ultrasonic the mixed solution obtained in step 4) for 6 hours, and centrifuge at 13000 rpm for 3 minutes to collect the material to obtain a graphene multifunctional composite material.

Example 4

1) Synthesize FePt magnetic nanomaterials with a particle size of 3nm by co-reduction method;

2) preparing functionalized graphene modified by metronidazole and polyethylene glycol 4000, the final concentration ratio of metronidazole, polyethylene glycol 4000 and graphene is 30:48:1;

3) Prepare 30mg/ml pentafluorouracil aqueous solution;

4) Mix the magnetic nanoparticles obtained in step 1) and the pentafluorouracil aqueous solution prepared in step 3) with the functionalized graphene aqueous solution prepared in step 2), so that the final concentration of the magnetic nanoparticles is 0.5 mg/ml, and the final concentration of pentafluorouracil The concentration is 3mg/ml, and the final concentration of functionalized graphene is 0.20mg/ml;

5) Ultrasonic the mixed solution obtained in step 4) for 4 hours, and centrifuge at 13000 rpm for 3 minutes to collect the material to obtain a graphene multifunctional composite material.

Figure 6 is the MTT method used to detect the inhibitory effect of the drug-loaded magnetic graphene multifunctional composite material obtained in this example on the proliferation of tumor cells, and the result of acting on the 1975 cell line for 24 hours, with a half-inhibitory concentration of 20ug/ml.

Finally, it should be noted that the above embodiments are only used to illustrate the technical solutions of the present invention rather than limit the protection scope of the present invention. Although the present invention has been described in detail with reference to the preferred embodiments, those of ordinary skill in the art should understand that The technical solution of the present invention can be modified or equivalently replaced without departing from the spirit and scope of the technical solution of the present invention.

Claims (11)

1. a preparation method of drug-loaded magnetic graphene multifunctional composite material, is characterized in that, comprises the steps: 1) preparing magnetic nanoparticles; 2) preparing a functionalized graphene material modified with nitroimidazole drugs and polyethylene glycol; 3) preparing an aqueous solution with a certain drug concentration; 4) adding the magnetic nanoparticles obtained in step 1) and the aqueous drug solution prepared in step 3) to the functionalized graphene material aqueous solution prepared in step 2), and mixing; 5) Ultrasonic the mixture obtained in step 4) at room temperature, and then centrifuge to collect the drug-loaded magnetic graphene composite. 2 . The preparation method according to claim 1 , wherein the magnetic nanoparticles in step 1) include any one of FePt, Fe ₂ O ₃ , Fe ₃ O ₄ , and MnFe ₂ O ₄ nanoparticles. 3. The preparation method according to claim 1, characterized in that the particle diameter of the magnetic nanoparticles prepared in step 1) is 2-20 nm. 4. The preparation method according to claim 1, characterized in that, the nitroimidazole drug in step 2) is metronidazole. 5. the preparation method as claimed in claim 1, is characterized in that, step 2) the method for preparing functionalized graphene material is: nitroimidazoles medicine and polyethylene glycol aqueous solution are mixed with graphene oxide aqueous solution, add carbonization Diimine and dimethylaminopyridine, stirred at room temperature for 24 to 72 hours. 6. The preparation method according to claim 5, characterized in that the amounts of nitroimidazoles and polyethylene glycol are 10 to 50 times that of graphene oxide. 7. The preparation method according to claim 1, characterized in that the drug in step 3) is a chemotherapeutic drug. 8. The preparation method according to claim 1, characterized in that the final concentration of the magnetic nanoparticles in the mixed solution formed in step 4) is 0.1-3 mg/ml, and the final concentration of the drug is 0.1-10 mg/ml. 9. The preparation method according to claim 1, characterized in that, step 5) ultrasonication time is 2-8 hours, and centrifugation is performed at 13000 rpm for 3 minutes. 10. A drug-loaded magnetic graphene multifunctional composite material, characterized in that it is prepared by the method according to any one of claims 1-9. 11. The application of the drug-loaded magnetic graphene multifunctional composite material according to claim 10 in the treatment of tumor drugs.