Preparation method and application of graphene oxide microcapsule carrying hydrophilic drug

Abstract

The invention discloses a preparation method and application of a hydrophilic drug-loaded graphene oxide microcapsule, and the graphene oxide microcapsule is a microstructure material which takes a cross-linked membrane constructed by graphene oxide as a shell and takes an organic solvent loaded with a hydrophilic drug as a core. The preparation method comprises the following steps: carrying out high-intensity ultrasonic radiation on an organic solvent containing the hydrophilic drug and a water phase containing sulfhydrylated graphene oxide, so as to prepare the graphene oxide microcapsule loaded with the hydrophilic drug in one step. The method is rapid, simple, convenient, efficient, environmentally friendly, wide in raw material source and high in packaging capacity, and can be widely applied to loading of various hydrophilic drugs. Besides, based on some characteristics of the graphene oxide microcapsule, such as pH responsiveness, reduction responsiveness, photothermal effect and the like of graphene oxide, photothermal therapy can be carried out while controllable release of hydrophilic drugs is realized. Therefore, the method disclosed by the invention is of great significance for simply and efficiently preparing the micro-structure carrier and synchronously realizing safe packaging of the hydrophilic medicine.

Description

Technical Field

[0001] The invention belongs to the cross-technical field of chemistry, material science and biomedicine, and relates to a preparation method and application of graphene oxide microcapsules carrying hydrophilic drugs. Background Art

[0002] Graphene oxide has the advantages of large specific surface area, low toxicity, good biocompatibility, good water solubility, etc., and has been widely used in the biomedical field. Wang et al. modified graphene oxide with polyethyleneimine and 6-arm polyethylene glycol as a drug delivery system, and both complexes were internalized by Raw 264.7 cells. Mona Alibolandi et al. covalently conjugated dextran to the surface of graphene oxide, and the drug delivery system was non-toxic to the 4T1 breast cancer cell line at a concentration of up to 300 μg / mL.

[0003] Microcapsules are solids, liquids, and gases encapsulated inside a wall-forming material as the capsule core. Since the internal substances can be effectively protected by the external wall-forming material, they are widely used in catalysis, energy storage, synthesis, drug delivery and other fields. The materials used to prepare the microcapsule wall are varied and can be organic polymer materials or inorganic materials. There are many methods for preparing microcapsules, such as layer-by-layer self-assembly, chemical cross-linking, in-situ polymerization, spray drying and aqueous phase separation. Although there are many types of microcapsule preparation technologies, the operation process is mostly cumbersome, the reaction time is long, and it is difficult to achieve mass production.

[0004] Ultrasonic radiation was invented by American scientists Suslick and others in the 1980s. Suslick and others used an ultrasonic machine gun to destroy the internal chemical environment of the solution, and coated the gas or water-insoluble liquid with protein as a shell to prepare protein microcapsules. The sonochemical method is simple to operate, low-cost, efficient and green. In recent decades, the sonochemical method has not only been explored in polymer chemistry, biomass proliferation, extraction and environmental remediation, but also widely used in sonocatalysis, organic sonochemistry and sonochemical preparation of materials. Although there are many papers on the preparation of microcapsules by sonochemical method at home and abroad, there are no domestic and foreign literature and patent reports on the preparation of hydrophilic drug-carrying graphene oxide microcapsules by sonochemical method. Summary of the invention

[0005] In view of the above-mentioned deficiencies of the prior art, the present invention provides a method for preparing graphene oxide microcapsules carrying hydrophilic drugs; the preparation method can achieve the encapsulation of the organic solvent carrying the hydrophilic drugs while completing the preparation of the graphene oxide microcapsules.

[0006] The specific technical solutions of the present invention are as follows:

[0007] A method for preparing hydrophilic drug-carrying graphene oxide microcapsules comprises the following steps: ultrasonically irradiating an organic solvent containing the hydrophilic drug and an aqueous solution containing thiolated graphene oxide, thereby finally forming microcapsules with graphene oxide as a shell and an organic solvent carrying the hydrophilic drug as a core; wherein the organic solvent containing the hydrophilic drug is prepared by the following method: dissolving the hydrophilic drug in a flowing hydrogel, and then dispersing the hydrogel containing the hydrophilic drug in an organic solvent containing an oil-in-water emulsifier by ultrasound, thereby forming an organic solvent containing the hydrophilic drug.

[0008] The specific steps are as follows:

[0009] (1) ultrasonically dispersing graphene oxide in deionized water, adding a compound containing both a thiol group and an amino group to the solution, magnetically stirring for 24 hours under N2 protection, centrifuging and washing, removing unreacted compounds containing both a thiol group and an amino group, and obtaining thiolated graphene oxide;

[0010] (2) dispersing the thiolated graphene oxide in deionized water; dissolving a hydrophilic drug in the flowing hydrogel, and then dispersing the hydrogel containing the hydrophilic drug in an organic solvent containing an oil-in-water emulsifier by ultrasound to form an organic solvent containing the hydrophilic drug;

[0011] (3) adding an aqueous solution of thiolated graphene oxide and an organic solvent containing a hydrophilic drug, placing the solution in an ice water bath environment, placing an ultrasonic probe at the interface of the organic solvent / water phases, and performing high-intensity radiation; the volume ratio of the aqueous phase containing thiolated graphene oxide to the organic solvent containing the hydrophilic drug is 3:1-10:1; the temperature of the ice water bath is -5-10°C; the power of the ultrasonic radiation is 200-900W, and the time of the ultrasonic radiation is 3-10min.

[0012] (4) After the ultrasound treatment, the reaction solution is dialyzed, and the dialyzed product is collected to finally obtain microcapsules with graphene oxide as the shell and an organic solvent carrying a hydrophilic drug as the core.

[0013] Wherein, the hydrogel is a hyaluronic acid hydrogel or a carboxymethyl cellulose hydrogel. The water-in-oil emulsifier is one of Span 20, Span 40, Span 60, and Span 80. The hydrophilic drug is at least one of doxorubicin hydrochloride, epirubicin hydrochloride, and aclarubicin hydrochloride. The organic solvent is one of ethyl acetate, ethyl benzoate, isopropyl acetate, and isopentyl acetate. The compound containing both sulfhydryl and amino groups is one of cysteamine and its hydrochloride, cysteine ​​and its hydrochloride, homocysteine, and glutathione.

[0014] Further features, in step (1), in order to successfully thiolate graphene oxide, the graphene oxide and the compound containing both thiol groups and amino groups should maintain an appropriate content ratio. The content ratio of each raw material is as follows: the content of the graphene oxide in the water phase is 0.5-2 mg / mL; the content of the compound containing both thiol groups and amino groups in the water phase is 2-8 mg / mL.

[0015] Further features, in step (2), the content ratio of each raw material has an important influence on the synthesized hydrophilic drug-carrying graphene oxide microcapsules. In order to successfully construct the microcapsule shell of the organic solvent carrying the hydrophilic drug, the concentration of thiolated graphene oxide should be appropriate; the hydrogel can not only dissolve the hydrophilic drug, but also play a supporting role inside the microcapsule, so the hydrogel containing the hydrophilic drug and the organic solvent containing the oil-in-water emulsifier should maintain an appropriate content ratio; the content of the oil-in-water emulsifier should be appropriate. When the content of the oil-in-water emulsifier is too high, the viscosity of the organic solvent will increase, the water phase dispersion effect will deteriorate, and the size of the formed microcapsules will increase. When the content of the oil-in-water emulsifier is too low, the hydrogel containing the hydrophilic drug cannot be completely dispersed in the organic solvent, which is not conducive to the formation of microcapsules. Therefore, preferably, the content ratio of each raw material is as follows: the content of the thiolated graphene oxide in the aqueous phase is 0.5-1 mg / mL; the content of the hydrophilic drug in the hydrogel is 1-5 mg / mL; the volume ratio of the hydrogel containing the hydrophilic drug to the organic solvent containing the oil-in-water emulsifier is 1:3-1:10; the content of the oil-in-water emulsifier in the organic solvent is 5-20 mg / mL.

[0016] According to a further feature, in step (4), dialysis can extract and purify graphene oxide microcapsules, and the dialysis time is 48-96 hours.

[0017] The hydrophilic drug-carrying graphene oxide microcapsules prepared by the above method are spherical or elliptical, with a particle size of 0.5-10 μm, and their structure includes a shell layer and a core; wherein the shell layer is a cross-linked membrane constructed by graphene oxide, and the core is an organic solvent carrying the hydrophilic drug. The graphene oxide microcapsules can be used as a material carrier. DETAILED DESCRIPTION

[0018] Example 1

[0019] 100 mg of graphene oxide was ultrasonically dispersed in 200 mL of deionized water, 400 mg of cysteine ​​hydrochloride was added to the above solution, and magnetic stirring was performed for 24 h under N2 protection. The unreacted cysteine ​​hydrochloride was removed by centrifugation and washed for 3 times to obtain thiolated graphene oxide; the thiolated graphene oxide was dispersed in 200 mL of deionized water; 20 mg of doxorubicin hydrochloride was dissolved in 20 mL of carboxymethyl cellulose hydrogel in a fluid state, and then 1 mL of carboxymethyl cellulose hydrogel was uniformly dispersed in 3 mL of ethyl acetate containing Span 20 (5 mg / mL) by ultrasound to form ethyl acetate containing doxorubicin hydrochloride; the thiolated graphene oxide aqueous solution and ethyl acetate containing doxorubicin hydrochloride were added to a beaker in a volume ratio of 3:1, and the beaker was placed in an ice-water bath at -5°C; the ultrasonic probe was placed at the interface of ethyl acetate / water phases, and high-intensity ultrasonic radiation (200 W, 3 min) was performed; after the ultrasonication, the reaction solution was dialyzed for 48 hours, and the dialyzed product was collected, and finally microcapsules with graphene oxide as the shell and ethyl acetate containing doxorubicin hydrochloride as the core were obtained. Under a polarizing microscope, the microcapsules appeared spherical or elliptical; under a laser confocal microscope, doxorubicin hydrochloride was found to be successfully encapsulated inside the microcapsules.

[0020] Example 2

[0021] 200 mg of graphene oxide was ultrasonically dispersed in 200 mL of deionized water, 800 mg of cysteine ​​hydrochloride was added to the above solution, and magnetic stirring was performed for 24 h under N2 protection. The unreacted cysteine ​​hydrochloride was removed by centrifugation and washed three times to obtain thiolated graphene oxide; the thiolated graphene oxide was dispersed in 200 mL of deionized water; 40 mg of doxorubicin hydrochloride was dissolved in 20 mL of carboxymethyl cellulose hydrogel in a flowing state, and then 1 mL of carboxymethyl cellulose hydrogel was uniformly dispersed in 4 mL of ethyl benzoate containing Span 20 (10 mg / mL) by ultrasound to form ethyl benzoate containing doxorubicin hydrochloride; according to Add thiolated graphene oxide aqueous solution and ethyl benzoate containing doxorubicin hydrochloride in a volume ratio of 4:1 to a beaker, and place it in an ice-water bath at 0°C; place the ultrasonic probe at the interface of ethyl benzoate / water, and perform high-intensity ultrasonic radiation (300W, 4min); after the end of the ultrasonic treatment, dialyze the reaction solution for 60h, collect the dialyzed product, and finally obtain microcapsules with graphene oxide as the shell and ethyl benzoate containing doxorubicin hydrochloride as the core. Under a polarizing microscope, the microcapsules are spherical or elliptical; under a laser confocal microscope, it is found that doxorubicin hydrochloride is successfully encapsulated inside the microcapsules.

[0022] Example 3

[0023] 300 mg of graphene oxide was ultrasonically dispersed in 200 mL of deionized water, and 1200 mg of cysteamine hydrochloride was added to the above solution. Under N2 protection, magnetic stirring was performed for 24 h. The unreacted cysteamine hydrochloride was removed by centrifugation and washed for 3 times to obtain thiolated graphene oxide; the thiolated graphene oxide was dispersed in 600 mL of deionized water; 60 mg of doxorubicin hydrochloride was dissolved in 20 mL of hyaluronic acid hydrogel in a flowing state, and then 1 mL of hyaluronic acid hydrogel was uniformly dispersed in 5 mL of ethyl acetate containing Span 40 (15 mg / mL) by ultrasound to form ethyl acetate containing doxorubicin hydrochloride; the thiolated graphene oxide aqueous solution and ethyl acetate containing doxorubicin hydrochloride were added to a beaker in a volume ratio of 5:1, and the beaker was placed in an ice-water bath at 5°C; the ultrasonic probe was placed at the interface of ethyl acetate / water phases, and high-intensity ultrasonic radiation (400 W, 5 min) was performed; after the ultrasonication, the reaction solution was dialyzed for 72 h, and the dialyzed product was collected, and finally microcapsules with graphene oxide as the shell and ethyl acetate containing doxorubicin hydrochloride as the core were obtained. Under a polarizing microscope, the microcapsules appeared spherical or elliptical; under a laser confocal microscope, doxorubicin hydrochloride was found to be successfully encapsulated inside the microcapsules.

[0024] Example 4

[0025] 400 mg of graphene oxide was ultrasonically dispersed in 200 mL of deionized water, 1600 mg of cysteamine hydrochloride was added to the above solution, and magnetic stirring was performed for 24 h under N2 protection. The solution was centrifuged and washed three times to remove the unreacted cysteamine hydrochloride to obtain thiolated graphene oxide; the thiolated graphene oxide was dispersed in 400 mL of deionized water; 100 mg of doxorubicin hydrochloride was dissolved in 20 mL of hyaluronic acid hydrogel in a flowing state, and then 1 mL of hyaluronic acid hydrogel was uniformly dispersed in 6 mL of ethyl benzoate containing Span 40 (20 mg / mL) by ultrasound to form ethyl benzoate containing doxorubicin hydrochloride; according to 6: Add thiolated graphene oxide aqueous solution and ethyl benzoate containing doxorubicin hydrochloride in a volume ratio of 1 to a beaker, and place it in an ice water bath at 10°C; place an ultrasonic probe at the interface of ethyl benzoate / water, and perform high-intensity ultrasonic radiation (500W, 6min); after the end of the ultrasonic wave, dialyze the reaction solution for 96h, collect the dialyzed product, and finally obtain microcapsules with graphene oxide as the shell and ethyl benzoate containing doxorubicin hydrochloride as the core. Under a polarizing microscope, the microcapsules are spherical or elliptical; under a laser confocal microscope, it is found that doxorubicin hydrochloride is successfully encapsulated inside the microcapsules.

[0026] Example 5

[0027] 100 mg of graphene oxide was ultrasonically dispersed in 200 mL of deionized water, 400 mg of homocysteine ​​was added to the above solution, and magnetic stirring was performed for 24 h under N2 protection. The thiolated graphene oxide was washed by centrifugation for 3 times to remove the unreacted homocysteine ​​and obtain the thiolated graphene oxide; the thiolated graphene oxide was dispersed in 200 mL of deionized water; 20 mg of epirubicin hydrochloride was dissolved in 20 mL of carboxymethyl cellulose hydrogel in a flowing state, and then 1 mL of carboxymethyl cellulose hydrogel was uniformly dispersed in 7 mL of isopropyl acetate containing Span 60 (5 mg / mL) by ultrasound to form isopropyl acetate containing epirubicin hydrochloride; the thiolated graphene oxide aqueous solution and isopropyl acetate containing epirubicin hydrochloride were added to a beaker in a volume ratio of 7:1, and the beaker was placed in an ice-water bath at -5°C; the ultrasonic probe was placed at the isopropyl acetate / water two-phase interface and high-intensity ultrasonic irradiation (600 W, 7 min) was performed; after the ultrasonication, the reaction solution was dialyzed for 48 hours, and the dialyzed product was collected, and finally microcapsules with graphene oxide as the shell and isopropyl acetate containing epirubicin hydrochloride as the core were obtained. Under a polarizing microscope, the microcapsules appeared spherical or elliptical; under a laser confocal microscope, it was found that epirubicin hydrochloride was successfully encapsulated inside the microcapsules.

[0028] Example 6

[0029] 200 mg of graphene oxide was ultrasonically dispersed in 200 mL of deionized water, and 800 mg of homocysteine ​​was added to the above solution. Under N2 protection, magnetic stirring was performed for 24 h. The thiolated graphene oxide was washed by centrifugation for 3 times to remove the unreacted homocysteine ​​and obtain the thiolated graphene oxide; the thiolated graphene oxide was dispersed in 200 mL of deionized water; 40 mg of epirubicin hydrochloride was dissolved in 20 mL of carboxymethyl cellulose hydrogel in a flowing state, and then 1 mL of carboxymethyl cellulose hydrogel was uniformly dispersed in 8 mL of isoamyl acetate containing Span 60 (10 mg / mL) by ultrasound to form isoamyl acetate containing epirubicin hydrochloride; the thiolated graphene oxide aqueous solution and isoamyl acetate containing epirubicin hydrochloride were added to a beaker in a volume ratio of 8:1, and the beaker was placed in an ice-water bath at 0°C; the ultrasonic probe was placed at the isoamyl acetate / water two-phase interface and high-intensity ultrasonic irradiation (700 W, 8 min) was performed; after the ultrasonication, the reaction solution was dialyzed for 60 hours, and the dialyzed product was collected, and finally microcapsules with graphene oxide as the shell and isoamyl acetate containing epirubicin hydrochloride as the core were obtained. Under a polarizing microscope, the microcapsules appeared spherical or elliptical; under a laser confocal microscope, it was found that epirubicin hydrochloride was successfully dispersed inside the microcapsules.

[0030] Example 7

[0031] 300 mg of graphene oxide was ultrasonically dispersed in 200 mL of deionized water, and 1200 mg of glutathione was added to the above solution, and magnetic stirring was performed for 24 h under N2 protection. The thiolated graphene oxide was washed by centrifugation for 3 times to remove the unreacted glutathione and obtain the thiolated graphene oxide; the thiolated graphene oxide was dispersed in 600 mL of deionized water; 60 mg of aclarubicin hydrochloride was dissolved in 20 mL of hyaluronic acid hydrogel in a flowing state, and then 1 mL of hyaluronic acid hydrogel was uniformly dispersed in 9 mL of isopropyl acetate containing Span 80 (15 mg / mL) by ultrasound to form isopropyl acetate containing aclarubicin hydrochloride; the thiolated graphene oxide aqueous solution and isopropyl acetate containing aclarubicin hydrochloride were added to a beaker in a volume ratio of 9:1, and the beaker was placed in an ice-water bath at 5°C; the ultrasonic probe was placed at the isopropyl acetate / water two-phase interface and high-intensity ultrasonic radiation (800 W, 9 min) was performed; after the ultrasonication, the reaction solution was dialyzed for 72 h, and the dialyzed product was collected, and finally microcapsules with graphene oxide as the shell and isopropyl acetate containing aclarubicin hydrochloride as the core were obtained. Under a polarizing microscope, the microcapsules appeared spherical or elliptical; under a laser confocal microscope, it was found that aclarubicin hydrochloride was successfully dispersed inside the microcapsules.

[0032] Example 8

[0033] 400 mg of graphene oxide was ultrasonically dispersed in 200 mL of deionized water, and 1600 mg of glutathione was added to the above solution. Under N2 protection, magnetic stirring was performed for 24 h. The unreacted glutathione was removed by centrifugation and washed three times to obtain thiolated graphene oxide; the thiolated graphene oxide was dispersed in 400 mL of deionized water; 100 mg of aclarubicin hydrochloride was dissolved in 20 mL of hyaluronic acid hydrogel in a flowing state, and then 1 mL of hyaluronic acid hydrogel was uniformly dispersed in 10 mL of isoamyl acetate containing Span 80 (20 mg / mL) by ultrasound to form isoamyl acetate containing aclarubicin hydrochloride; the mixture was stirred at a ratio of 10:1. Add thiolated graphene oxide aqueous solution and isoamyl acetate containing aclarubicin hydrochloride in a volume ratio of 1.5-2.5% to a beaker, and place it in an ice-water bath at 10°C; place the ultrasonic probe at the isoamyl acetate / water interface and perform high-intensity ultrasonic radiation (900W, 10min); after the ultrasonic treatment, dialyze the reaction solution for 96h, collect the dialyzed product, and finally obtain microcapsules with graphene oxide as the shell and isoamyl acetate containing aclarubicin hydrochloride as the core. Under a polarizing microscope, the microcapsules are spherical or elliptical; under a laser confocal microscope, it was found that aclarubicin hydrochloride was successfully dispersed inside the microcapsules.

Claims

1. A method for preparing graphene oxide microcapsules carrying hydrophilic drugs, characterized in that: Ultrasonic radiation is applied to an organic solvent containing a hydrophilic drug and an aqueous phase containing thiolated graphene oxide, thereby finally forming a microcapsule having a cross-linked membrane constructed by graphene oxide as a shell and an organic solvent carrying the hydrophilic drug as a core; The organic solvent containing the hydrophilic drug is prepared by the following method: dissolving the hydrophilic drug into a fluidized hydrogel, and then dispersing the hydrogel containing the hydrophilic drug into an organic solvent containing an oil-in-water emulsifier by ultrasound to form an organic solvent containing the hydrophilic drug.

2. The method for preparing a hydrophilic drug-carrying graphene oxide microcapsule according to claim 1, characterized in that: The steps are as follows: (1) ultrasonically dispersing graphene oxide in deionized water, adding a compound containing both a thiol group and an amino group to the solution, magnetically stirring for 24 hours under N2 protection, centrifuging and washing, removing unreacted compounds containing both a thiol group and an amino group, and obtaining thiolated graphene oxide; (2) dispersing the thiolated graphene oxide in deionized water; Dissolving a hydrophilic drug into a fluidized hydrogel, and then dispersing the hydrophilic drug-containing hydrogel into an organic solvent containing an oil-in-water emulsifier by ultrasound to form an organic solvent containing the hydrophilic drug; (3) adding an aqueous solution of thiolated graphene oxide and an organic solvent containing a hydrophilic drug, placing the solution in an ice water bath environment, placing an ultrasonic probe at the interface of the organic solvent / water phases, and performing high-intensity radiation; the volume ratio of the aqueous phase containing thiolated graphene oxide to the organic solvent containing the hydrophilic drug is 3:1-10:1; the temperature of the ice water bath is -5-10°C; the power of the ultrasonic radiation is 200-900W, and the time of the ultrasonic radiation is 3-10min. (4) After the ultrasound treatment is completed, the reaction solution is dialyzed, and the dialyzed product is collected to finally obtain graphene oxide microcapsules loaded with hydrophilic drugs.

3. The method for preparing a hydrophilic drug-carrying graphene oxide microcapsule according to claim 2, characterized in that: The hydrogel is hyaluronic acid hydrogel or carboxymethyl cellulose hydrogel; The water-in-oil emulsifier is one of Siban 20, Siban 40, Siban 60 and Siban 80; The hydrophilic drug is at least one of doxorubicin hydrochloride, epirubicin hydrochloride, and aclarubicin hydrochloride; The organic solvent is one of ethyl acetate, ethyl benzoate, isopropyl acetate and isoamyl acetate; The compound containing both sulfhydryl and amino groups is one of cysteamine and its hydrochloride, cysteine ​​and its hydrochloride, homocysteine ​​and glutathione.

4. The method for preparing a hydrophilic drug-carrying graphene oxide microcapsule according to claim 2 or 3, characterized in that: In step (1), the content of the graphene oxide in the aqueous phase is 0.5-2 mg / mL; the content of the compound containing both thiol and amino groups in the aqueous phase is 2-8 mg / mL.

5. The method for preparing a hydrophilic drug-carrying graphene oxide microcapsule according to claim 4, characterized in that: In step (2), the content of the thiolated graphene oxide in the aqueous phase is 0.5-1 mg / mL; the content of the hydrophilic drug in the hydrogel is 1-5 mg / mL; the volume ratio of the hydrogel containing the hydrophilic drug to the organic solvent containing the water-in-oil emulsifier is 1:3-1:10; the content of the water-in-oil emulsifier in the organic solvent is 5-20 mg / mL.

6. The method for preparing a hydrophilic drug-carrying graphene oxide microcapsule according to claim 2, 3 or 5, characterized in that: In step (4), the dialysis time is 48-96 hours.

7. Use of the hydrophilic drug-carrying graphene oxide microcapsules prepared by the method according to any one of claims 2 to 5 as material carriers.