Drug-loaded microspheres of hydrophobic flavonoid active molecules as well as preparation method and application of drug-loaded microspheres

Abstract

The invention belongs to the technical field of pharmaceutical preparations, and particularly relates to drug-loaded microspheres of hydrophobic flavonoid active molecules as well as a preparation method and application of the drug-loaded microspheres. According to the invention, a preparation process of the drug-loaded microspheres, and components and contents of the drug-loaded microspheres are screened, so that optimized drug-loaded microspheres are obtained. Polyacryloyl chloride particles, hydrophobic flavonoid active molecules and polyethyleneimine are prepared into medicine carrying microspheres according to the weight part ratio of (5-10): (80-120): (15-25). The preparation method is simple and environment-friendly, and the prepared drug-loading microspheres are high in drug-loading capacity and encapsulation efficiency, have pH responsiveness during in-vitro release, and are excellent in stability, good in slow release performance and high in transdermal drug delivery efficiency. The solubility and bioavailability of hydrophobic flavonoid active molecules are improved, the development of flavonoid compounds in medicines is promoted, and the application prospect is good.

Description

Technical Field

[0001] The invention belongs to the technical field of pharmaceutical preparations, and in particular relates to drug-loaded microspheres of hydrophobic flavonoid active molecules and a preparation method and application thereof. Background Art

[0002] Flavonoids are a class of natural products widely found in plants and have a variety of biological activities, such as antioxidant, anti-inflammatory, and anti-tumor activities. However, many flavonoid active molecules are hydrophobic, which makes them less soluble in water and less bioavailable, limiting their application in the pharmaceutical field.

[0003] In order to improve the solubility and bioavailability of hydrophobic flavonoid active molecules, researchers have developed various drug delivery systems, such as nanoparticles, liposomes, microspheres, etc., and some studies on drug-loaded microspheres of flavonoid active molecules have been reported. For example, products prepared by the emulsification solvent volatilization method have considerable drug loading and encapsulation rates, but the existing preparation methods of drug-loaded microspheres are generally complex, with large amounts of organic solvents, and drug leakage is prone to occur. At the same time, in the field of transdermal drug delivery, the skin barrier hinders the absorption of peptides, and transdermal penetration affects the stability of active ingredients. The existing transdermal absorption technology has poor transdermal efficiency and post-transdermal stability, and is not sensitive to pH. Therefore, in terms of improving the solubility and bioavailability of flavonoid active molecules, there are problems such as complex preparation methods, large amounts of organic solvents, easy leakage, poor transdermal efficiency and post-transdermal stability.

[0004] Therefore, it is of great practical significance to develop a simple and environmentally friendly preparation method for synthesizing efficient and stable hydrophobic flavonoid active molecule drug-loaded microspheres. Summary of the invention

[0005] In view of the defects of the prior art, the present invention provides a hydrophobic flavonoid active molecule drug-loaded microsphere and a preparation method and use thereof.

[0006] The present invention provides a hydrophobic flavonoid active molecule drug-loaded microsphere, which is prepared from raw materials including the following components in parts by weight:

[0007] 5-10 parts of polyacryl chloride particles,

[0008] 80-120 hydrophobic flavonoid molecules,

[0009] 15-25 parts of polyethyleneimine;

[0010] The molecular weight of the polyacryl chloride in the polyacryl chloride particles is 1500 Da, and the molecular weight of the polyethyleneimine is 1800 Da.

[0011] Preferably, the hydrophobic flavonoid molecule is selected from at least one of dihydrobisflavonoids, chromones, and flavan-3-ols.

[0012] Preferably, the hydrophobic flavonoid molecule is stellerachalinensis chromone.

[0013] The present invention provides the drug-loaded microspheres described in any one of the above items, which are prepared according to the following steps:

[0014] Polyacryl chloride particles are mixed with hydrophobic flavonoid molecules and then reacted with polyethyleneimine to obtain the product.

[0015] Preferably, the polyacryl chloride particles are prepared by dissolving polyacryl chloride in acetone or ethanol.

[0016] Preferably, the polyacryloyl chloride is prepared according to the following steps:

[0017] Acryloyl chloride is mixed with a photoinitiator and irradiated with light at 360-370nm to obtain the product.

[0018] Preferably, the irradiation dose is 1800-2200 mJ / cm 2 ; and / or, the volume ratio of acryloyl chloride to photoinitiator is 3-1:2-1, and the volume concentration of the photoinitiator is 1.5%-2.5%; and / or, the photoinitiator is Darocur 1173.

[0019] Preferably, the hydrophobic flavonoid molecules are dissolved in dimethyl sulfoxide, polyethylene glycol, or a mixed solution thereof; and / or, the mixing time is 20 to 40 minutes; and / or, after the mixing, the solution is added dropwise to water; and / or, the polyethyleneimine is dissolved in water and then added dropwise to the reaction solution; and / or, the reaction is stirred for 2-3 hours.

[0020] The present invention provides a method for preparing any of the above-mentioned drug-loaded microspheres, which comprises the following steps: mixing polyacryl chloride particles with hydrophobic flavonoid molecules, and then reacting with polyethyleneimine to obtain the drug-loaded microspheres.

[0021] The present invention provides use of any of the above-mentioned drug-loaded microspheres in the preparation of anti-inflammatory, antioxidant and anti-tumor drugs.

[0022] The present invention provides drug-loaded microspheres of hydrophobic flavonoid active molecules, and a preparation method and use thereof. The present invention screens the preparation process of drug-loaded microspheres, the components and content of drug-loaded microspheres, and obtains preferred drug-loaded microspheres. Polyacryl chloride particles, hydrophobic flavonoid active molecules and polyethyleneimine are prepared into drug-loaded microspheres in a weight ratio of 5-10:80-120:15-25. The preparation method is simple, the prepared drug-loaded microspheres have high drug loading and encapsulation rate, are pH responsive during in vitro release, have excellent stability, good sustained-release performance, and high transdermal drug delivery efficiency. The present invention improves the solubility and bioavailability of hydrophobic flavonoid active molecules, promotes the development of flavonoid compounds in medicines, and has good application prospects.

[0023] Obviously, according to the above contents of the present invention, in accordance with common technical knowledge and customary means in the art, without departing from the above basic technical ideas of the present invention, other various forms of modification, replacement or change may be made.

[0024] The above contents of the present invention are further described in detail below through specific implementation methods in the form of embodiments. However, this should not be understood as the scope of the above subject matter of the present invention being limited to the following examples. All technologies realized based on the above contents of the present invention belong to the scope of the present invention. BRIEF DESCRIPTION OF THE DRAWINGS

[0025] Figure 1 The scanning electron microscope images of polymer particles prepared under different polymer molecular weights and solvents;

[0026] Figure 2 This is the result of percutaneous penetration. DETAILED DESCRIPTION

[0027] In the following examples and experimental examples, reagents and materials not particularly described are all commercially available.

[0028] Example 1 Hydrophobic flavonoid active molecule drug-loaded microspheres and preparation method thereof

[0029] The hydrophobic flavonoid active molecule drug-loaded microspheres of this embodiment are prepared by the following method:

[0030] (1) Acryloyl chloride (3 mL) and photopolymerization initiator Darocur 1173 (2% v / v, 2 mL) were mixed in a glass bottle. The bottle was then placed horizontally under an ultraviolet light source (365 nm) at 2000 mJ / cm 2After irradiation, the polymer (200 mL) was mixed with excess toluene (HPLC grade, 400-500 mL) and purified by centrifugation (10000 rpm, 30 minutes), and then washed alternately with toluene and methanol three times to obtain purified polyacryl chloride (PAC) with a molecular weight of 1500 MW. Finally, the purified PAC was redispersed in acetone to prepare a 5 mg / mL PAC particle / acetone solution.

[0031] (2) Dissolve the stellerachaline chromone in DMSO solution to prepare a 5 mg / mL stellerachaline chromone solution.

[0032] (3) 20 mL of the stellera chamaejasme chromone solution (5 mg / mL) and 1.5 mL of the PAC particle / acetone solution (5 mg / mL) were mixed. After 30 minutes, the mixed solution was added dropwise to 20 mL of water to prepare an aqueous solution.

[0033] (4) Polyethyleneimine (PEI, purity 99%, MW: 1800 Da, purchased from Polysciences, Inc.) was dissolved in water to prepare a 2 mg / mL polyethyleneimine solution.

[0034] (5) Add 10 mL of polyethyleneimine solution (2 mg / mL) to the aqueous solution prepared in step (3) and stir with a magnetic bar for 3 hours to ensure that the acetone is completely evaporated. Centrifuge the solution after the reaction at 3000 rpm for 10 minutes to obtain the drug-loaded microspheres. The eluate is used to determine the drug loading amount. Wash with distilled water three times, take the precipitate, and vacuum dry it.

[0035] The technical solution of the present invention is further described below through experiments. The drug-loaded microspheres of hydrophobic flavonoid active molecules tested in the following experimental examples were prepared by the method of Example 1.

[0036] Experimental Example 1 Characterization of drug-loaded microspheres

[0037] The particle size and distribution of drug-loaded microspheres are measured by scanning electron microscopy (SEM) and laser particle size analyzer. By adjusting the preparation process parameters, such as stirring speed, temperature, material concentration ratio, etc., high-quality microspheres with uniform particle size and narrow distribution are prepared to meet different administration routes and drug release requirements.

[0038] 1. Experimental Methods

[0039] 1. Determination of the effects of solvent and polymer molecular weight on the particle size of drug-loaded microspheres

[0040] The structural properties of the carrier material mainly depend on the polymer and solvent interaction parameters, molecular structure and polymer molecular weight, concentration, pH, etc.

[0041] Drug-loaded microspheres were prepared according to the preparation method of Example 1, except that the purified PAC in step (1) was redispersed in different solvents, including acetone, DMSO, ethanol, tetrahydrofuran, pure water, and PBS; the molecular weights of polyacryl chloride in step (1) were 1500MW and 300MW, respectively. The prepared drug-loaded microspheres were subjected to SEM examination.

[0042] 2. Experimental Results

[0043] 1. Effects of polymer molecular weight and solvent on particle size of drug-loaded microspheres

[0044] The results of the effect of polymer molecular weight on the particle size of drug-loaded microspheres are shown in Figure 1 As shown: when the molecular weight of PAC is 1500MW, after being dissolved in different solvents, the prepared microspheres are small and evenly distributed, indicating that when the molecular weight of PAC is 1500MW, it is better for the subsequent preparation of drug-loaded microspheres.

[0045] The results of the effect of solvent on the particle size of drug-loaded microspheres are shown in Figure 1 As shown in Table 1: When the solvent is acetone, the diameter of the prepared microspheres is about 100nm, the maximum size is 200nm, and the size is uniform; when the solvent is DMSO, the diameter of the prepared microspheres is 200-600nm, and the polymer film exists at the same time; when the solvent is ethanol, the particle size of the microspheres is about 60nm, the maximum size is 100nm, and it is a translucent solution; when the solvent is tetrahydrofuran, the particle size of the prepared microspheres is about 400nm, and the surface is covered with a polymer film; when the solvent is water, the polymer is insoluble, and small-sized polymer nanoparticles with uniform particle size cannot be prepared. Therefore, when PAC is dissolved in solvent acetone or ethanol, small-sized polymer nanoparticles with uniform particle size can be prepared.

[0046] Therefore, when the molecular weight of PAC is 1500 MW, when the solvent acetone or ethanol is used to dissolve PAC, small-sized polymer nanoparticles with uniform particle size can be prepared.

[0047] Table 1 Effect of solvent on particle size of drug-loaded microspheres

[0048]

[0049]

[0050] Note: δD represents dispersion force parameter, δP represents polarity parameter, δH represents hydrogen bonding parameter, Molar volume is molar volume, and X(AC) represents the solubility of acryloyl chloride in different solvents.

[0051] The above results show that the molecular weight of the polymer and the type of organic solvent have a great influence on the particle size of the drug-loaded microspheres. When the molecular weight of PAC is 1500MW, when the solvent acetone or ethanol is used to dissolve PAC, small-sized polymer nanoparticles with uniform particle size can be prepared, with particle diameters of 60nm and 100nm, which is conducive to transdermal penetration.

[0052] Experimental Example 2 Determination of drug loading and encapsulation efficiency

[0053] 1. Experimental Methods

[0054] 1. Determination of drug loading and drug embedding rate

[0055] The aqueous solution obtained in step 4 of Example 1 was centrifuged, and the solution was taken to measure the drug loading efficiency and drug encapsulation efficiency of the PAC particles.

[0056] The eluate from step 5 of Example 1 was collected and used to determine the drug loading efficiency and drug encapsulation efficiency of the PAC / PEI particles. The calculation method is as follows:

[0057] Drug loading = (loaded flavonoid mass / total sample mass) x 100%

[0058] Drug embedding rate = (loaded flavonoid mass / initial total flavonoid mass added) x 100%

[0059] The mass of flavonoids loaded = the total mass of flavonoids initially added - the mass of free flavonoids in the eluate

[0060] The mass of free flavonoids in the eluate was detected by UV-vis (Varian Cary5000 spectrophotometer, with distilled water as control)

[0061] 2. Determination of zeta potential

[0062] The aqueous solution obtained in step 4 of Example 1 was centrifuged, and the precipitate was taken and dissolved in water. The zeta potential was measured by a laser particle size analyzer, and the zeta potential of the drug+PAC particles was measured.

[0063] The material prepared in Example 1 is drug + PAC / PEI particles, which are dissolved in water and the zeta potential is measured by a laser particle size analyzer. The zeta potential of the drug + PAC / PEI particles is measured.

[0064] 2. Experimental Results

[0065] The results are shown in Table 2: Compared with PAC particles, the drug loading and drug embedding rate of PAC / PEI particles are significantly increased, the zeta potential is higher, and the system is stable. The results show that the drug-loaded microspheres prepared by the method of Example 1 of the present invention have high drug loading and drug embedding rate, and the system is stable.

[0066] Table 2

[0067]

[0068] Experimental Example 3 Performance Determination of In Vitro Drug Release

[0069] 1. Experimental Methods

[0070] 1. In vitro drug release assay

[0071] The drug release was measured for 5 hours by UV spectrophotometer (Varian Cary 5000 spectrophotometer). The specific steps of the measurement are as follows:

[0072] 2 mL of sample (10 mM NaCl) was placed in a dialysis membrane bag (3000 molecular weight), which was then placed in 10 mL buffer solution (10 mM NaCl) at pH 7 for 2 hours, followed by 10 mL buffer solution (10 mM NaCl) at pH 4 for 3 hours. The released drug was quantified by absorbance at 210 nm.

[0073] 2. Determination of the effect of percutaneous penetration

[0074] The transdermal permeation effect was compared with stellerachaline chromone as the control sample. The in vitro transdermal absorption test was conducted using the Franz diffusion cell method. The specific steps are as follows:

[0075] (1) Take frozen skin with subcutaneous tissue removed, thaw it in physiological saline, and use a diffusion cell with a diameter of 1.5 cm to fix the skin tissue between the supply cell and the receiving cell of the diffusion cell, with the stratum corneum facing the supply cell.

[0076] (2) The supply tank and receiving tank are filled with saline solution, and the circulating water is 37-38 degrees Celsius.

[0077] (3) Use a pipette to add 1 ml of sample to the supply pool, so that the sample evenly covers the skin area. Turn on the magnetic stirrer and stir at a speed of 600 r / min.

[0078] (4) At intervals of 3, 6, 9, 12, 15, 18, and 21 hours, remove 1 ml of receiving solution and add an equal amount of constant temperature receiving solution.

[0079] (5) The receiving solution was compared with distilled water, and the absorbance was measured at 210 nm to calculate the transdermal penetration rate.

[0080] Transdermal rate = (content of receiving pool sample mg / ml) * (volume of receiving pool solution ml) / (initial mass of supply pool sample mg)

[0081] 2. Experimental Results

[0082] 1. pH-responsive drug release

[0083] The experimental results show that the drug-loaded microspheres of the present invention have pH-responsive drug release.

[0084] 2. Effect of transdermal penetration

[0085] The results of percutaneous penetration are as follows Figure 2 As shown: the skin permeation rate of the drug-loaded microspheres of the present invention is significantly better than that of the control sample. As time increases, the permeation amount of the drug-loaded microspheres prepared by the present invention increases, reaching 35% permeation at 15 hours and 46% permeation at 21 hours. The results show that the drug-loaded microspheres prepared by the present invention can significantly improve the permeation effect of the skin.

[0086] Through the above embodiments and experimental examples, it can be seen that the present invention provides a drug-loaded microsphere of hydrophobic flavonoid active molecules and its preparation method and use. The present invention screened the preparation process of drug-loaded microspheres, the components and content of drug-loaded microspheres, and obtained preferred drug-loaded microspheres. Polyacryl chloride particles, hydrophobic flavonoid active molecules and polyethyleneimine are prepared into drug-loaded microspheres in a weight ratio of 5-10:80-120:15-25. The preparation method is simple and environmentally friendly, the prepared drug-loaded microspheres have high drug loading and encapsulation rate, are pH responsive during in vitro release, have excellent stability, good sustained-release performance, and high transdermal drug delivery efficiency. The present invention improves the solubility and bioavailability of hydrophobic flavonoid active molecules, promotes the development of flavonoid compounds in medicines, and has good application prospects.

Claims

1. A drug-loaded microsphere containing hydrophobic flavonoid active molecules, characterized in that: It is prepared from raw materials including the following components in parts by weight: 5-10 parts of polyacryl chloride particles, 80-120 hydrophobic flavonoid molecules, 15-25 parts of polyethyleneimine; The molecular weight of the polyacryl chloride in the polyacryl chloride particles is 1500 Da, and the molecular weight of the polyethyleneimine is 1800 Da.

2. The drug-loaded microspheres according to claim 1, characterized in that: The hydrophobic flavonoid molecules are selected from at least one of dihydrobisflavonoids, chromones, and flavan-3-ols.

3. The drug-loaded microspheres according to claim 2, characterized in that: The hydrophobic flavonoid molecule is stellera chamaejasme chromone.

4. The drug-loaded microspheres according to any one of claims 1 to 3, characterized in that: Prepared according to the following steps: The polyacryl chloride particles are mixed with hydrophobic flavonoid molecules and then reacted with polyethyleneimine to obtain the product.

5. The drug-loaded microspheres according to claim 4, characterized in that: The polyacryl chloride particles are prepared by dissolving polyacryl chloride in acetone or ethanol.

6. The drug-loaded microspheres according to claim 5, characterized in that: The polyacryloyl chloride is prepared according to the following steps: Acryloyl chloride is mixed with a photoinitiator and irradiated with light at 360-370nm to obtain the product.

7. The drug-loaded microspheres according to claim 6, characterized in that: The irradiation dose is 1800-2200 mJ / cm 2 ; and / or, the volume ratio of acryloyl chloride to photoinitiator is 3-1:2-1, and the volume concentration of the photoinitiator is 1.5%-2.5%; and / or, the photoinitiator is Darocur 1173.

8. The drug-loaded microspheres according to claim 4, characterized in that: The hydrophobic flavonoid molecules are dissolved in dimethyl sulfoxide, polyethylene glycol, or a mixed solution thereof; and / or, the mixing time is 20 to 40 minutes; and / or, after the mixing, the solution is dripped into water; and / or, the polyethyleneimine is dissolved in water and then dripped into the reaction solution; and / or, the reaction is stirred for 2-3 hours.

9. The method for preparing the drug-loaded microspheres according to any one of claims 1 to 8, characterized in that: The method comprises the following steps: polyacryl chloride particles are mixed with hydrophobic flavonoid molecules, and then reacted with polyethyleneimine to obtain the obtained product.

10. Use of the drug-loaded microspheres according to any one of claims 1 to 8 in the preparation of anti-inflammatory, antioxidant and anti-tumor drugs.

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