Cyclodextrin medicine compound as well as preparation method and application thereof

Abstract

The invention provides a cyclodextrin medicine compound as well as a preparation method and application thereof, and belongs to the technical field of medicines. The preparation method comprises the following steps: performing hydroxylation modification on beta-cyclodextrin to prepare hydroxypropyl cyclodextrin, respectively embedding camptothecin, kushenin and bufalin, then performing spray drying with graphene oxide to prepare an embedded compound, depositing Ni-doped ferroferric oxide on the surface, and further performing folic acid modification to prepare the cyclodextrin medicine compound. The drug is conveyed to a tumor site through magnetic targeting, and the drug is combined with a receptor of tumor cells through active transportation of folic acid, so that the drug release precision is greatly improved, the drug has an extremely good treatment effect at a relatively low concentration, the adverse reaction of a patient is reduced, the compliance is improved, and the curative effect of the drug is improved. The medication cost is reduced, the bioavailability of the medicine is improved, and the application prospect is wide.

Description

Technical Field

[0001] The present invention relates to the field of medical technology, and in particular to a cyclodextrin drug complex and a preparation method and application thereof. Background Art

[0002] Many drugs are affected by heat, light, environment and air, and are easy to sublimate or volatilize, resulting in partial loss of efficacy. Some drugs also have very low solubility, which is not conducive to absorption by the human body and low bioavailability. Traditional drugs also have many problems in clinical applications, such as: large side effects, low elimination rate, and frequent medication to maintain efficacy. Therefore, developing new drugs to replace traditional drug systems has become an urgent problem to be solved. Since the cavity size of cyclodextrin is suitable for most drug molecules in clinical use or trials, the inclusion complex formed has certain stability and no toxic side effects, can increase the solubility of fat-soluble drugs, mask the bad smell of some drugs themselves, etc., resulting in more and more extensive applications of β-cyclodextrin and its derivatives.

[0003] Since Villiers discovered cyclodextrin in 1891, cyclodextrin chemistry has developed into an important part of supramolecular chemistry. In 1904, Schardinger first described the preparation of cyclodextrin in detail. The research work during this period progressed very slowly. From the mid-1930s to the late 1960s, Freudenberg obtained pure cyclodextrin, and he and his collaborators proposed that cyclodextrin is a cyclic molecule in which glucose units are combined in the form of maltose, and the molecule only contains α-1,4-glycosidic bonds. In 1953, Freudenberg and Cramer jointly summarized the important aspects of the application of cyclodextrin in pharmaceuticals. At the same time, the French and Cramer research groups studied the physical and chemical properties of cyclodextrin. During this period, people have realized that cyclodextrin is very likely to be used in the industrial field, but the understanding of its toxicity is still controversial, which seriously hinders its application. From the early 1970s to the present, people's research on cyclodextrin has entered a peak period. So far, cyclodextrin has been produced on a large scale in industry and applied to various fields.

[0004] With the changes in environment and lifestyle, the incidence of human malignant tumors has increased year by year. Common cancer treatments include surgical resection, chemotherapy, radiotherapy, etc., and chemotherapy, as one of the most commonly used treatments, often brings many serious side effects to patients due to the lack of selectivity of anticancer drugs. Therefore, establishing a targeted drug delivery system to kill cancer cells is a new direction for cancer detection and treatment. The construction of a targeted drug delivery system can be achieved by directly linking targeting groups to the drug, or by using targeted drug carriers to deliver drugs. Among them, since drug carriers can simultaneously have multiple functional effects such as drug protection and drug sustained release, in recent years, the research on the use of nanomaterials as drug carriers has gradually attracted attention. Summary of the invention

[0005] The purpose of the present invention is to propose a cyclodextrin drug complex and its preparation method and application, which can deliver drugs to tumor sites through magnetic targeting and active transport of folic acid to bind drugs to receptors of tumor cells, thereby greatly improving the accuracy of drug release, so that the drug has an excellent therapeutic effect at a lower concentration, thereby reducing adverse reactions of patients, improving compliance, reducing drug costs, and improving the bioavailability of drugs, and has broad application prospects.

[0006] The technical solution of the present invention is achieved in this way:

[0007] The invention provides a method for preparing a cyclodextrin drug complex. Beta-cyclodextrin is modified by hydroxylation to obtain hydroxypropyl cyclodextrin, camptothecin, matrine and bufotoxin are respectively embedded, and then they are spray-dried with graphene oxide to obtain an embedded complex, and Ni-doped ferrosoferric oxide is deposited on the surface, and further folic acid is modified to obtain a cyclodextrin drug complex.

[0008] As a further improvement of the present invention, the following steps are included:

[0009] S1. Preparation of hydroxypropyl cyclodextrin: dissolving β-cyclodextrin in hot alkali solution, adding propylene oxide, adjusting the pH value of the solution, concentrating under reduced pressure, adding ethanol to dissolve, adding NaCl to the solution for precipitation, filtering, dialyzing the filtrate, and freeze-drying to obtain hydroxypropyl cyclodextrin;

[0010] S2. Encapsulation of oxymatrine and bufalin; dispersing oxymatrine and bufalin in water, adding hydroxypropyl cyclodextrin, heating and stirring to react, dialyzing, and freeze-drying to obtain an encapsulated oxymatrine and bufalin complex;

[0011] S3. Encapsulation of camptothecin: camptothecin was dispersed in water, hydroxypropyl cyclodextrin was added, heated and stirred for reaction, dialyzed, and freeze-dried to obtain encapsulated camptothecin;

[0012] S4. Preparation of the embedded complex: The embedded oxymatrine and bufalin complex and the embedded camptothecin were mixed uniformly, added to the graphene oxide aqueous dispersion, ultrasonically dispersed uniformly, and spray-dried to obtain an embedded complex;

[0013] S5. Deposition of Ni-doped ferroferric oxide: adding the embedding complex to water, adding ferric chloride, nickel chloride and ferrous chloride, dropping ammonia water, heating and stirring to react, centrifuging, washing, and drying to obtain a Ni-doped ferroferric oxide doped embedding complex;

[0014] S6. Preparation of cyclodextrin drug complex: dissolve folic acid in dimethyl sulfoxide and triethanolamine, add NHS and EDC, stir the reaction in the dark, evaporate, add the product and Ni-doped ferroferric oxide-doped embedding complex into water, adjust the pH value, incubate in the dark, and obtain the cyclodextrin drug complex.

[0015] As a further improvement of the present invention, the hot alkali solution in step S1 is a 10-20wt% NaOH or KOH solution at 55-65°C, the mass ratio of β-cyclodextrin and propylene oxide is 10:4-7, and the pH value of the adjusted solution is 6.8-7.2.

[0016] As a further improvement of the present invention, the mass ratio of matrine, bufalin and hydroxypropyl cyclodextrin in step S2 is 2-3:3-4:8-12, and the temperature of the heating and stirring reaction is 60-70°C and the time is 4-6h.

[0017] As a further improvement of the present invention, in step S3, the mass ratio of camptothecin to hydroxypropyl cyclodextrin is 2-3:4-7, the temperature of the heating and stirring reaction is 60-70° C., and the time is 3-5 h.

[0018] As a further improvement of the present invention, the mass ratio of the embedded matrine and bufalin complex, embedded camptothecin, and graphene oxide in step S4 is 3-5:7-10:18-25.

[0019] As a further improvement of the present invention, the mass ratio of the embedding complex, ferric chloride, nickel chloride, ferrous chloride and ammonia water in step S5 is 10:3.24:0.5-1:1.26:5-7, the concentration of ammonia water is 20-25wt%, the temperature of the heating and stirring reaction is 75-85°C, and the time is 3-5h.

[0020] As a further improvement of the present invention, the mass ratio of folic acid, dimethyl sulfoxide, triethanolamine, NHS and EDC in step S6 is 0.4-0.6:1.5-2.5:0.05-0.1:0.07-0.09:0.1-0.12, the time of the light-proof stirring reaction is 1-2 hours, the pH value is adjusted to 9-10, the time of the light-proof incubation is 2-4 hours, and the mass ratio of the product to the Ni-doped ferroferric oxide doped embedding complex is 0.5-1:10.

[0021] The present invention further protects a cyclodextrin drug complex prepared by the above preparation method.

[0022] The present invention further protects the use of the cyclodextrin drug complex in the preparation of anti-tumor drugs.

[0023] The present invention has the following beneficial effects:

[0024] Due to the interaction between the aggregation and surrounding water molecules in water, as well as the effect of lattice energy in the solid state, the solubility of cyclodextrin in water varies. Among them, β-cyclodextrin has the lowest solubility in water, which limits its application in formulations. Chemically modified β-cyclodextrin, such as hydroxypropyl-β-cyclodextrin, eliminates the above problems, greatly improving the water solubility of the prepared hydroxypropyl-β-cyclodextrin and significantly improving the drug loading capacity. After some insoluble drugs are encapsulated by cyclodextrin, their water solubility increases, they have membrane permeability, are easy to absorb, and can be made into injections. At the same time, cyclodextrin can improve the stability of drugs, so that some drugs that are sensitive to light, heat and oxygen remain in a stable state.

[0025] Camptothecin, as an anticancer drug, is a neutral alkaloid that exists in the form of lactone ring at pH < 4.5. It is easy to open the ring to form a water-soluble carboxylate in a dilute alkaline solution at room temperature. Camptothecin is poorly soluble in water and has strong toxic side effects. Its anti-tumor activity is significantly reduced after salt formation, and its biological toxicity is mainly related to its carboxylate form. Due to its serious biological toxic side effects, its direct use in clinical practice is restricted. The use of nano drug carriers can improve the solubility of camptothecin and reduce the cytotoxicity of camptothecin compounds, thereby enhancing its efficacy.

[0026] Although oxymatrine and bufalotoxin can work synergistically with camptothecin to improve the anti-tumor efficacy by enhancing the body's immune function and inhibiting the growth and metastasis of tumor cells, and have a synergistic effect, both of them have low solubility in water and are prone to cause gastrointestinal reactions, such as nausea, vomiting, bitter taste in the mouth, loss of appetite, diarrhea, upper abdominal discomfort or pain, etc., which is mainly due to the irritation of the drug to the gastrointestinal mucosa.

[0027] The present invention adopts hydroxypropyl-β-cyclodextrin as an ultramicro carrier of anticancer drugs. The anticancer drug composition (camptothecin, matrine, bufalin) which is too irritating to be taken is embedded in the cavity of cyclodextrin to prepare an ultramicrocapsule inclusion compound, and then the drug is released by enzymatic hydrolysis in the body. Due to its ultramicrostructure and molecular dispersion, it is easy to absorb, has high bioavailability, slow drug release, and low side effects. After subsequent graphene oxide spray drying and embedding, the sustained release effect of the drug is further improved, the protective effect of the drug is enhanced, and the gastrointestinal reaction is further reduced.

[0028] The present invention further deposits ferroferric oxide on the surface of the prepared embedding complex and dopes it with nickel. On the one hand, the ferromagnetism of ferroferric oxide can be improved, so that it has a better magnetic targeted drug delivery effect. On the other hand, the doping of nickel metal can also inhibit the replication of viruses and has a significant therapeutic effect on the drug resistance of clinical tumor cells. The effect is good and the drug resistance is reversed.

[0029] The surface of the prepared Ni-doped ferroferric oxide-doped embedding complex is further modified with folic acid, which can bind to the receptors of tumor cells through active transport and enter the cells through endocytosis to enhance the therapeutic effect, greatly improving the efficacy of targeted therapy.

[0030] The present invention delivers drugs to the tumor site through magnetic targeting and active transport of folic acid to bind the drugs to the receptors of tumor cells, thereby greatly improving the accuracy of drug release, so that the drugs have excellent therapeutic effects at lower concentrations, thereby reducing patients' adverse reactions, improving compliance, reducing drug costs, and improving drug bioavailability, and has broad application prospects. DETAILED DESCRIPTION

[0031] The technical solutions in the embodiments of the present invention are described clearly and completely below. Obviously, the described embodiments are only part of the embodiments of the present invention, not all of them. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without creative work are within the scope of protection of the present invention.

[0032] NHS, N-hydroxysuccinimide; EDC, 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide.

[0033] Graphene oxide, particle size D 50 The particle size was 10-20 (μm), the pH value was 6-7, and it was purchased from Changzhou Sixth Element Materials Technology Co., Ltd.

[0034] Example 1

[0035] This embodiment provides a method for preparing a cyclodextrin drug complex, comprising the following steps:

[0036] S1. Preparation of hydroxypropyl cyclodextrin: 10 g of β-cyclodextrin was dissolved in 200 mL of a 10 wt% NaOH solution at 55°C, 4 g of propylene oxide was added, the pH value of the solution was adjusted to 6.8, the solution was concentrated under reduced pressure, ethanol was added to dissolve, NaCl was added to the solution to precipitate, the solution was filtered, the filtrate was dialyzed, and freeze-dried to obtain hydroxypropyl cyclodextrin;

[0037] S2. Encapsulation of oxymatrine and bufalin; 2 g oxymatrine and 3 g bufalin were added to 200 mL of water, ultrasonically dispersed at 1000 W for 15 min, 8 g hydroxypropyl cyclodextrin was added, heated to 60 ° C, stirred for 4 h, dialyzed, and freeze-dried to obtain an oxymatrine and bufalin encapsulated complex;

[0038] S3. Encapsulation of camptothecin: 2 g of camptothecin was added to 200 mL of water, ultrasonically dispersed at 1000 W for 15 min, 4 g of hydroxypropyl cyclodextrin was added, heated to 60 ° C, stirred for 3 h, dialyzed, and freeze-dried to obtain encapsulated camptothecin;

[0039] S4. Preparation of the embedded complex: 3 mg of the embedded oxymatrine and bufalin complex and 7 mg of the embedded camptothecin were mixed for 10 min, added to 50 mL of water, 18 mg of graphene oxide was added, ultrasonically dispersed at 1000 W for 15 min, and spray dried to obtain an embedded complex;

[0040] S5. Deposition of Ni-doped ferroferric oxide: 10 g of the embedding complex was added to 500 mL of water, 3.24 g of ferric chloride, 0.5 g of nickel chloride and 1.26 g of ferrous chloride were added, 5 g of 20 wt% ammonia water was added dropwise, the mixture was heated to 75°C, stirred for reaction for 3 h, centrifuged, washed and dried to obtain a Ni-doped ferroferric oxide doped embedding complex;

[0041] S6. Preparation of cyclodextrin drug complex: Dissolve 0.4g folic acid in a mixed liquid of 1.5g dimethyl sulfoxide and 0.05g triethanolamine, add 0.07g NHS and 0.1g EDC, stir and react for 1h in the dark, evaporate to dryness, add 0.5g product and 10g Ni-doped ferroferric oxide-doped embedding complex into 100mL water, adjust the pH value to 9, incubate in the dark for 2h to obtain cyclodextrin drug complex.

[0042] Example 2

[0043] This embodiment provides a method for preparing a cyclodextrin drug complex, comprising the following steps:

[0044] S1. Preparation of hydroxypropyl cyclodextrin: 10 g of β-cyclodextrin was dissolved in 200 mL of a 20 wt % KOH solution at 65° C., 7 g of propylene oxide was added, the pH value of the solution was adjusted to 7.2, the solution was concentrated under reduced pressure, ethanol was added to dissolve, NaCl was added to the solution to precipitate, the solution was filtered, the filtrate was dialyzed, and freeze-dried to obtain hydroxypropyl cyclodextrin;

[0045] S2. Encapsulation of oxymatrine and bufalin; 3 g oxymatrine and 4 g bufalin were added to 200 mL of water, ultrasonically dispersed at 1000 W for 15 min, 12 g hydroxypropyl cyclodextrin was added, heated to 70 ° C, stirred for 6 h, dialyzed, and freeze-dried to obtain an oxymatrine and bufalin encapsulated complex;

[0046] S3. Encapsulation of camptothecin: 3 g of camptothecin was added to 200 mL of water, ultrasonically dispersed at 1000 W for 15 min, 7 g of hydroxypropyl cyclodextrin was added, heated to 70 ° C, stirred for 5 h, dialyzed, and freeze-dried to obtain encapsulated camptothecin;

[0047] S4. Preparation of the embedded complex: 5 mg of the embedded oxymatrine and bufalin complex and 10 mg of the embedded camptothecin were mixed for 10 min, added to 50 mL of water, 25 mg of graphene oxide was added, ultrasonically dispersed at 1000 W for 15 min, and spray dried to obtain an embedded complex;

[0048] S5. Deposition of Ni-doped ferroferric oxide: 10 g of the embedding complex was added to 500 mL of water, 3.24 g of ferric chloride, 1 g of nickel chloride and 1.26 g of ferrous chloride were added, 7 g of 5 wt% ammonia water was added dropwise, the mixture was heated to 85°C, stirred for reaction for 5 h, centrifuged, washed and dried to obtain a Ni-doped ferroferric oxide doped embedding complex;

[0049] S6. Preparation of cyclodextrin drug complex: Dissolve 0.6g folic acid in a mixed liquid of 2.5g dimethyl sulfoxide and 0.1g triethanolamine, add 0.09g NHS and 0.12g EDC, stir and react for 2h in the dark, evaporate to dryness, add 1g product and 10g Ni-doped ferroferric oxide-doped embedding complex into 100mL water, adjust the pH value to 10, incubate in the dark for 4h to obtain cyclodextrin drug complex.

[0050] Example 3

[0051] This embodiment provides a method for preparing a cyclodextrin drug complex, comprising the following steps:

[0052] S1. Preparation of hydroxypropyl cyclodextrin: 10 g of β-cyclodextrin was dissolved in 200 mL of a 15 wt% NaOH solution at 60°C, 5 g of propylene oxide was added, the pH value of the solution was adjusted to 7, the solution was concentrated under reduced pressure, ethanol was added to dissolve, NaCl was added to the solution to precipitate, the solution was filtered, the filtrate was dialyzed, and freeze-dried to obtain hydroxypropyl cyclodextrin;

[0053] S2. Encapsulation of oxymatrine and bufalin; 2.5 g oxymatrine and 3.5 g bufalin were added to 200 mL of water, ultrasonically dispersed at 1000 W for 15 min, 10 g hydroxypropyl cyclodextrin was added, heated to 65 ° C, stirred for 5 h, dialyzed, and freeze-dried to obtain an oxymatrine and bufalin encapsulated complex;

[0054] S3. Encapsulation of camptothecin: 2.5 g of camptothecin was added to 200 mL of water, ultrasonically dispersed at 1000 W for 15 min, 5.5 g of hydroxypropyl cyclodextrin was added, heated to 65 ° C, stirred for 4 h, dialyzed, and freeze-dried to obtain encapsulated camptothecin;

[0055] S4. Preparation of the embedded complex: 4 mg of the embedded oxymatrine and bufalin complex and 8 mg of the embedded camptothecin were mixed for 10 min, added to 50 mL of water, 22 mg of graphene oxide was added, ultrasonically dispersed at 1000 W for 15 min, and spray dried to obtain an embedded complex;

[0056] S5. Deposition of Ni-doped ferroferric oxide: 10 g of the embedding complex was added to 500 mL of water, 3.24 g of ferric chloride, 0.7 g of nickel chloride and 1.26 g of ferrous chloride were added, 6 g of 22 wt% ammonia water was added dropwise, the mixture was heated to 80°C, stirred for 4 h, centrifuged, washed and dried to obtain a Ni-doped ferroferric oxide doped embedding complex;

[0057] S6. Preparation of cyclodextrin drug complex: Dissolve 0.5g folic acid in a mixed liquid of 2g dimethyl sulfoxide and 0.07g triethanolamine, add 0.08g NHS and 0.11g EDC, stir and react for 1.5h in the dark, evaporate to dryness, add 0.7g product and 10g Ni-doped ferroferric oxide-doped embedding complex into 100mL water, adjust the pH value to 9.5, incubate in the dark for 3h to obtain cyclodextrin drug complex.

[0058] Comparative Example 1

[0059] Compared with Example 3, the difference is that step S1 is not performed.

[0060] The details are as follows:

[0061] S1. Encapsulation of oxymatrine and bufalin; 2.5 g oxymatrine and 3.5 g bufalin were added to 200 mL of water, ultrasonically dispersed at 1000 W for 15 min, 10 g of β-cyclodextrin was added, heated to 65 ° C, stirred for 5 h, dialyzed, and freeze-dried to obtain an oxymatrine and bufalin encapsulated complex;

[0062] S2. Encapsulation of camptothecin: 2.5 g camptothecin was added to 200 mL water, ultrasonically dispersed at 1000 W for 15 min, 5.5 g β-cyclodextrin was added, heated to 65 ° C, stirred for 4 h, dialyzed, and freeze-dried to obtain encapsulated camptothecin;

[0063] S3. Preparation of the embedded complex: 4 mg of the embedded oxymatrine and bufalin complex and 8 mg of the embedded camptothecin were mixed for 10 min, added to 50 mL of water, 22 mg of graphene oxide was added, ultrasonically dispersed at 1000 W for 15 min, and spray dried to obtain an embedded complex;

[0064] S4. Deposition of Ni-doped ferroferric oxide: 10 g of the embedding complex was added to 500 mL of water, 3.24 g of ferric chloride, 0.7 g of nickel chloride and 1.26 g of ferrous chloride were added, 6 g of 22 wt% ammonia water was added dropwise, the mixture was heated to 80°C, stirred for 4 h, centrifuged, washed and dried to obtain a Ni-doped ferroferric oxide doped embedding complex;

[0065] S5. Preparation of cyclodextrin drug complex: Dissolve 0.5g folic acid in a mixed liquid of 2g dimethyl sulfoxide and 0.07g triethanolamine, add 0.08g NHS and 0.11g EDC, stir and react for 1.5h in a dark place, evaporate to dryness, add 0.7g product and 10g Ni-doped ferroferric oxide-doped embedding complex into 100mL water, adjust the pH value to 9.5, incubate in a dark place for 3h to obtain cyclodextrin drug complex.

[0066] Comparative Example 2

[0067] Compared with Example 3, the difference is that oxymatrine is not added in step S2.

[0068] The details are as follows:

[0069] S2. Encapsulation of bufalon venom; add 6 g of bufalon venom to 200 mL of water, disperse at 1000 W ultrasonically for 15 min, add 10 g of hydroxypropyl cyclodextrin, heat to 65°C, stir and react for 5 h, dialyze, and freeze-dry to obtain the encapsulated bufalon venom.

[0070] Comparative Example 3

[0071] Compared with Example 3, the difference is that bufalin is not added in step S2.

[0072] The details are as follows:

[0073] S2. Encapsulation of oxymatrine; add 6 g oxymatrine into 200 mL water, disperse it at 1000 W ultrasonically for 15 min, add 10 g hydroxypropyl cyclodextrin, heat to 65°C, stir and react for 5 h, dialyze, and freeze-dry to obtain the encapsulated bufotoxin.

[0074] Comparative Example 4

[0075] Compared with Example 3, the difference is that no composite of encapsulated oxymatrine and bufalotoxin is added in step S4.

[0076] The details are as follows:

[0077] S4. Preparation of the embedded material: 12 mg of embedded camptothecin was mixed for 10 min, added into 50 mL of water, 22 mg of graphene oxide was added, ultrasonically dispersed at 1000 W for 15 min, and spray dried to obtain the embedded material.

[0078] Comparative Example 5

[0079] Compared with Example 3, the difference is that step S4 is not performed.

[0080] The details are as follows:

[0081] S1. Preparation of hydroxypropyl cyclodextrin: 10 g of β-cyclodextrin was dissolved in 200 mL of a 15 wt% NaOH solution at 60°C, 5 g of propylene oxide was added, the pH value of the solution was adjusted to 7, the solution was concentrated under reduced pressure, ethanol was added to dissolve, NaCl was added to the solution to precipitate, the solution was filtered, the filtrate was dialyzed, and freeze-dried to obtain hydroxypropyl cyclodextrin;

[0082] S2. Encapsulation of oxymatrine and bufalin; 2.5 g oxymatrine and 3.5 g bufalin were added to 200 mL of water, ultrasonically dispersed at 1000 W for 15 min, 10 g hydroxypropyl cyclodextrin was added, heated to 65 ° C, stirred for 5 h, dialyzed, and freeze-dried to obtain an oxymatrine and bufalin encapsulated complex;

[0083] S3. Encapsulation of camptothecin: 2.5 g of camptothecin was added to 200 mL of water, ultrasonically dispersed at 1000 W for 15 min, 5.5 g of hydroxypropyl cyclodextrin was added, heated to 65 ° C, stirred for 4 h, dialyzed, and freeze-dried to obtain encapsulated camptothecin;

[0084] S4. Preparation of the mixture: 4 mg of the embedded oxymatrine and bufalin complex and 8 mg of the embedded camptothecin were mixed for 10 min to obtain a mixture;

[0085] S5. Deposition of Ni-doped ferroferric oxide: 10 g of the mixture was added to 500 mL of water, 3.24 g of ferric chloride, 0.7 g of nickel chloride and 1.26 g of ferrous chloride were added, 6 g of 22 wt% ammonia water was added dropwise, the mixture was heated to 80°C, stirred for 4 h, centrifuged, washed and dried to obtain a Ni-doped ferroferric oxide doped mixture;

[0086] S6. Preparation of cyclodextrin drug complex: Dissolve 0.5g folic acid in a mixed liquid of 2g dimethyl sulfoxide and 0.07g triethanolamine, add 0.08g NHS and 0.11g EDC, stir and react for 1.5h in the dark, evaporate to dryness, add 0.7g product and 10g Ni-doped ferroferric oxide-doped mixture into 100mL water, adjust the pH value to 9.5, incubate in the dark for 3h to obtain a cyclodextrin drug complex.

[0087] Comparative Example 6

[0088] Compared with Example 3, the difference is that nickel chloride is not added in step S5.

[0089] The details are as follows:

[0090] S5. Deposition of ferroferric oxide: add 10 g of the embedding complex into 500 mL of water, add 3.24 g of ferric chloride and 1.26 g of ferrous chloride, add 6 g of 22 wt% ammonia water, heat to 80°C, stir and react for 4 hours, centrifuge, wash, and dry to obtain the ferroferric oxide doped embedding complex.

[0091] Comparative Example 7

[0092] Compared with Embodiment 3, the difference is that step S5 is not performed.

[0093] The details are as follows:

[0094] S1. Preparation of hydroxypropyl cyclodextrin: 10 g of β-cyclodextrin was dissolved in 200 mL of a 15 wt% NaOH solution at 60°C, 5 g of propylene oxide was added, the pH value of the solution was adjusted to 7, the solution was concentrated under reduced pressure, ethanol was added to dissolve, NaCl was added to the solution to precipitate, the solution was filtered, the filtrate was dialyzed, and freeze-dried to obtain hydroxypropyl cyclodextrin;

[0095] S2. Encapsulation of oxymatrine and bufalin; 2.5 g oxymatrine and 3.5 g bufalin were added to 200 mL of water, ultrasonically dispersed at 1000 W for 15 min, 10 g hydroxypropyl cyclodextrin was added, heated to 65 ° C, stirred for 5 h, dialyzed, and freeze-dried to obtain an oxymatrine and bufalin encapsulated complex;

[0096] S3. Encapsulation of camptothecin: 2.5 g of camptothecin was added to 200 mL of water, ultrasonically dispersed at 1000 W for 15 min, 5.5 g of hydroxypropyl cyclodextrin was added, heated to 65 ° C, stirred for 4 h, dialyzed, and freeze-dried to obtain encapsulated camptothecin;

[0097] S4. Preparation of the embedded complex: 4 mg of the embedded oxymatrine and bufalin complex and 8 mg of the embedded camptothecin were mixed for 10 min, added to 50 mL of water, 22 mg of graphene oxide was added, ultrasonically dispersed at 1000 W for 15 min, and spray dried to obtain an embedded complex;

[0098] S5. Preparation of cyclodextrin drug complex: Dissolve 0.5g folic acid in a mixed liquid of 2g dimethyl sulfoxide and 0.07g triethanolamine, add 0.08g NHS and 0.11g EDC, stir and react for 1.5h in the dark, evaporate to dryness, add 0.7g product and 10g embedding complex into 100mL water, adjust pH value to 9.5, incubate in the dark for 3h to obtain cyclodextrin drug complex.

[0099] Comparative Example 8

[0100] Compared with the embodiment 3, the difference is that step S6 is not performed.

[0101] The details are as follows:

[0102] S1. Preparation of hydroxypropyl cyclodextrin: 10 g of β-cyclodextrin was dissolved in 200 mL of a 15 wt% NaOH solution at 60°C, 5 g of propylene oxide was added, the pH value of the solution was adjusted to 7, the solution was concentrated under reduced pressure, ethanol was added to dissolve, NaCl was added to the solution to precipitate, the solution was filtered, the filtrate was dialyzed, and freeze-dried to obtain hydroxypropyl cyclodextrin;

[0103] S2. Encapsulation of oxymatrine and bufalin; 2.5 g oxymatrine and 3.5 g bufalin were added to 200 mL of water, ultrasonically dispersed at 1000 W for 15 min, 10 g hydroxypropyl cyclodextrin was added, heated to 65 ° C, stirred for 5 h, dialyzed, and freeze-dried to obtain an oxymatrine and bufalin encapsulated complex;

[0104] S3. Encapsulation of camptothecin: 2.5 g of camptothecin was added to 200 mL of water, ultrasonically dispersed at 1000 W for 15 min, 5.5 g of hydroxypropyl cyclodextrin was added, heated to 65 ° C, stirred for 4 h, dialyzed, and freeze-dried to obtain encapsulated camptothecin;

[0105] S4. Preparation of the embedded complex: 4 mg of the embedded oxymatrine and bufalin complex and 8 mg of the embedded camptothecin were mixed for 10 min, added to 50 mL of water, 22 mg of graphene oxide was added, ultrasonically dispersed at 1000 W for 15 min, and spray dried to obtain an embedded complex;

[0106] S5. Deposition of Ni-doped ferroferric oxide: Add 10 g of the embedding complex into 500 mL of water, add 3.24 g of ferric chloride, 0.7 g of nickel chloride and 1.26 g of ferrous chloride, add 6 g of 22 wt% ammonia water, heat to 80°C, stir the reaction for 4 hours, centrifuge, wash, and dry to obtain a Ni-doped ferroferric oxide doped embedding complex, which is a cyclodextrin drug complex.

[0107] 10 5 MiaPaCa-2 cells were seeded at 100 / mL in 12-well plates and cultured overnight. 50 The cells were subjected to shock treatment for 24 hours with 1% paclitaxel (20% concentration) to induce pancreatic cancer cells to develop resistance to napaclitaxel. The culture medium was then replaced twice a week, and the drug concentration was gradually increased when the cells grew well. The culture was continued for 2 weeks to obtain a drug-resistant cell line. The drug-resistant cell line was transferred to a new tissue culture flask for further culture, passaged at 75-80% confluence, and directly seeded into a drug-containing culture medium, and divided into a control group, a positive drug group, Examples 1-3, and Comparative Example 6. Among them, the control group did not add drugs, and the positive drug group (camptothecin), Examples 1-3, and Comparative Example 6 (the corresponding cyclodextrin drug complexes) were added with different concentrations of the correspondingly prepared product solutions. Each group had 5 replicate wells, and the experiment was repeated 3 times. The blank culture medium without cells was used as the blank group for control, and the absorbance (A) value was detected at a wavelength of 450nm using an enzyme marker. Calculate the IC of the drug 50 The results are shown in Table 1.

[0108] Cell viability (%) = (A value of experimental group - A value of blank group) / (A value of control group - A value of blank group) × 100%

[0109] Inhibition rate (%) = (1-cell viability) × 100%.

[0110] Table 1

[0111] Group <![CDATA[IC 50 Value (mmol / mL)]]> Example 1 1.4 Example 2 1.2 Example 3 1.1 Comparative Example 6 57.8 Positive drug group 224.7

[0112] It can be seen from the above table that the cyclodextrin drug complexes prepared in Examples 1-3 of the present invention have a good in vitro inhibitory effect on drug-resistant cell lines and reverse the drug resistance.

[0113] Test Example 2

[0114] H-22 female tumor-bearing mice were selected and randomly divided into 12 groups, with 6 mice in each group. Physiological saline, cyclodextrin drug complex prepared in Example 1-3 or Comparative Example 1-8 were respectively injected into the tail vein (dosage was 10 mg / kg). The first administration time was set at day 0, and the drugs were administered on days 0, 3, and 6, respectively. At the same time as the administration, a magnetic field was applied to the tumor site with a magnetic field intensity of 0.2T. It was applied once on each administration day, each time for 30 minutes. The long diameter (L, mm) and short diameter (W, mm) of the tumor mass were measured with a vernier caliper every other day, and the tumor volume (V t , mm 3 ): V t =L×W 2 / 2, until the largest tumor grew to 3000mm, all mice were killed, the tumor was dissected out, photographed and weighed. The results are shown in Table 2.

[0115] Tumor inhibition rate (%) = (1-W t / W c )×100%

[0116] Among them, W t W is the tumor weight of the drug-treated group (g); c is the tumor weight (g) of the normal saline control group.

[0117] Table 2

[0118]

[0119]

[0120] It can be seen from the above table that the cyclodextrin drug complexes prepared in Examples 1-3 of the present invention have a good effect of suppressing weight in vivo.

[0121] The above description is only a preferred embodiment of the present invention and is not intended to limit the present invention. Any modifications, equivalent substitutions, improvements, etc. made within the spirit and principle of the present invention should be included in the protection scope of the present invention.

Claims

1. A method for preparing a cyclodextrin drug complex, characterized in that: β-cyclodextrin was modified by hydroxylation to obtain hydroxypropyl cyclodextrin, which was used to encapsulate camptothecin, matrine and bufotoxin, respectively. The composite was then spray-dried with graphene oxide to obtain an encapsulation complex. Ni-doped ferrous oxide was deposited on the surface and further modified with folic acid to obtain a cyclodextrin drug complex.

2. The preparation method according to claim 1, characterized in that: The following steps are involved: S1. Preparation of hydroxypropyl cyclodextrin: dissolving β-cyclodextrin in hot alkali solution, adding propylene oxide, adjusting the pH value of the solution, concentrating under reduced pressure, adding ethanol to dissolve, adding NaCl to the solution for precipitation, filtering, dialyzing the filtrate, and freeze-drying to obtain hydroxypropyl cyclodextrin; S2. Encapsulation of oxymatrine and bufalin; dispersing oxymatrine and bufalin in water, adding hydroxypropyl cyclodextrin, heating and stirring to react, dialyzing, and freeze-drying to obtain an encapsulated oxymatrine and bufalin complex; S3. Encapsulation of camptothecin: camptothecin was dispersed in water, hydroxypropyl cyclodextrin was added, heated and stirred for reaction, dialyzed, and freeze-dried to obtain encapsulated camptothecin; S4. Preparation of the embedded complex: The embedded oxymatrine and bufalin complex and the embedded camptothecin were mixed uniformly, added to the graphene oxide aqueous dispersion, ultrasonically dispersed uniformly, and spray-dried to obtain an embedded complex; S5. Deposition of Ni-doped ferroferric oxide: adding the embedding complex to water, adding ferric chloride, nickel chloride and ferrous chloride, dropping ammonia water, heating and stirring to react, centrifuging, washing, and drying to obtain a Ni-doped ferroferric oxide doped embedding complex; S6. Preparation of cyclodextrin drug complex: dissolve folic acid in dimethyl sulfoxide and triethanolamine, add NHS and EDC, stir the reaction in the dark, evaporate, add the product and Ni-doped ferroferric oxide-doped embedding complex into water, adjust the pH value, incubate in the dark, and obtain the cyclodextrin drug complex.

3. The preparation method according to claim 2, characterized in that: The hot alkali solution in step S1 is a 10-20wt% NaOH or KOH solution at 55-65°C, the mass ratio of β-cyclodextrin to propylene oxide is 10:4-7, and the pH value of the adjusted solution is 6.8-7.

2.

4. The preparation method according to claim 2, characterized in that: In step S2, the mass ratio of matrine, bufalin and hydroxypropyl cyclodextrin is 2-3:3-4:8-12, and the temperature of the heating and stirring reaction is 60-70° C. and the time is 4-6 hours.

5. The preparation method according to claim 2, characterized in that: In step S3, the mass ratio of camptothecin to hydroxypropyl cyclodextrin is 2-3:4-7, the temperature of the heating and stirring reaction is 60-70° C., and the time is 3-5 hours.

6. The preparation method according to claim 2, characterized in that: The mass ratio of the embedded matrine and bufalin complex, embedded camptothecin, and graphene oxide in step S4 is 3-5:7-10:18-25.

7. The preparation method according to claim 2, characterized in that: In step S5, the mass ratio of the embedding complex, ferric chloride, nickel chloride, ferrous chloride and ammonia water is 10:3.24:0.5-1:1.26:5-7, the concentration of ammonia water is 20-25wt%, the temperature of the heating and stirring reaction is 75-85°C, and the time is 3-5h.

8. The preparation method according to claim 2, characterized in that: The mass ratio of folic acid, dimethyl sulfoxide, triethanolamine, NHS and EDC in step S6 is 0.4-0.6:1.5-2.5:0.05-0.1:0.07-0.09:0.1-0.12, the time of the light-proof stirring reaction is 1-2 hours, the pH value is adjusted to 9-10, the time of the light-proof incubation is 2-4 hours, and the mass ratio of the product to the Ni-doped ferroferric oxide doped embedding complex is 0.5-1:

10.

9. A cyclodextrin drug complex prepared by the preparation method according to any one of claims 1 to 8.

10. Use of the cyclodextrin drug complex according to claim 9 in the preparation of anti-tumor drugs.

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