Preparation method of arginine and ursolic acid modified hyaluronic acid polymer and micelle

Abstract

The invention relates to the field of pharmaceutical preparations, and discloses a preparation method of ursolic acid and arginine modified hyaluronic acid polymer and micelle.The preparation method comprises the steps that cystamine serves as a connecting bond, hydrophobic ursolic acid is modified to a hyaluronic acid framework, then arginine is modified to the hyaluronic acid framework through esterification reaction, and the polymer is synthesized. A drug-loaded micelle is prepared by taking an indissolvable drug as a model drug and adopting a dialysis method. The preparation method disclosed by the invention is simple in process, the prepared polymer micelle is high in drug loading capacity, the problem of low solubility of insoluble drugs is effectively improved, and the polymer micelle has a certain application value.

Description

Technical Field

[0001] The present invention relates to the field of pharmaceutical preparations, and particularly to a preparation method of a hyaluronic acid polymer modified with arginine and ursolic acid and micelles. Background Art

[0002] The solubility problem of poorly soluble drugs has always been a difficult point that people want to overcome. The nano-drug delivery system can increase the solubility of drugs, thereby increasing the drug utilization rate. The polymer micelles represented by the nano-drug delivery system have the advantages of good stability, strong drug-loading capacity, increased drug solubility, few adverse reactions, and improved bioavailability of drugs. Polymer micelles are mainly composed of a biocompatible hydrophilic chain segment and a hydrophobic chain segment of a multifunctional drug-loading core. When the hydrophobic chain segment and the hydrophilic chain segment reach or exceed the critical micelle concentration in an aqueous medium, a core-shell structure micelle solution with hydrophobic groups inside and hydrophilic groups outside is self-assembled due to intermolecular interactions. At present, polymer drug-loaded micelles have received more and more attention and have become a research hotspot in nano-pharmaceutical preparations. However, very few drug-loaded micelles have been successfully applied to the medical field. Therefore, polymer micelles have research significance as carriers in the administration of poorly soluble drug carriers.

[0003] In view of this, the present invention aims to prepare a novel polymer micelle to improve the solubility problem of poorly soluble drugs. Summary of the Invention

[0004] The object of the present invention is to provide a preparation method of a hyaluronic acid polymer modified with arginine and ursolic acid and micelles, using a poorly soluble drug as a model drug to improve its solubility.

[0005] Technical Solution: To achieve the above invention object, the technical solution adopted by the present invention is as follows:

[0006] A hyaluronic acid polymer modified with arginine and ursolic acid, the chemical structure of which is shown in general formula I:

[0007]

[0008] Preferably, in the polymer, the molecular weight of sodium hyaluronate is 1-570 kDa, the grafting rate of cystamine is 26-54%, the grafting rate of ursolic acid is 1-31%, and the grafting rate of arginine is 2-30%.

[0009] The synthesis method of the hyaluronic acid polymer modified with arginine and ursolic acid includes the following steps:

[0010]

[0011] (1) Prepare an aqueous solution of sodium hyaluronate, add EDC and NHS, and adjust the pH to the range of 4-5 to activate the carboxyl group;

[0012] (2) Prepare an aqueous solution of cystamine dihydrochloride, add sodium bicarbonate for neutralization reaction, dropwise add the cystamine aqueous solution into the activated sodium hyaluronate aqueous solution, adjust the pH to the range of 7-8, react under nitrogen protection in the dark, after the reaction, dialyze with deionized water and freeze-dry to obtain cystamine-modified hyaluronic acid (CYHA);

[0013] (3) Prepare an aqueous solution of ursolic acid, add EDC and NHS, after activating the carboxyl group, dropwise add the suspension aqueous solution of CYHA, react under nitrogen protection in the dark, after the reaction, dialyze with deionized water, wash thoroughly with dichloromethane and freeze-dry to obtain ursolic acid-modified hyaluronic acid (UAHA);

[0014] (4) Prepare an aqueous solution of arginine, add EDC and DMAP, add the UAHA solution, adjust the pH to 7, react under nitrogen protection in the dark, after the reaction, dialyze with deionized water and freeze-dry to obtain arginine and ursolic acid-modified hyaluronic acid

[0015] (AUHA).

[0016] Preferably, in step (1), the molar ratio of sodium hyaluronate, EDC, and NHS is 0.25-2:1-8:1-4; the concentration of sodium hyaluronate is 1-8 mg / mL; the time for activating the carboxyl group is 20-30 min.

[0017] Preferably, in step (2), the molar ratio of sodium hyaluronate, cystamine dihydrochloride, and sodium bicarbonate is 0.25-2:0.5-20:1-40; the time for the reaction under nitrogen protection in the dark is 40-56 h, and the time for dialyzing with deionized water after the reaction is 48-72 h.

[0018] Preferably, in step (3), the ursolic acid solution is ursolic acid dissolved in DMSO with a concentration of 1-30 mg / mL, the molar ratio of ursolic acid, EDC, and NHS is 0.25-3:1-12:4-12, the molar ratio of ursolic acid to CYHA is 1-2:1, the time for activating the carboxyl group is 20-30 min, the time for the reaction under nitrogen protection in the dark is 40-56 h, and the time for dialyzing with deionized water after the reaction is 48-72 h.

[0019] Preferably, in step (4), the concentration of arginine is 1-300 mg / mL, the molar ratio of arginine, EDC, and DMAP is 0.25-2:1-5:0.05-0.3, the molar ratio of arginine to UAHA is 1-2:1, the UAHA solution is UAHA dissolved in DMSO with a concentration of 1-30 mg / mL, the time for the reaction under nitrogen protection in the dark is 40-56 h, and the time for dialyzing with deionized water after the reaction is 48-72 h.

[0020] Preferably, the preparation method of the blank nanomicelles comprises: dissolving AUHA in water with a solution concentration of 0.1 - 50 mg / mL, performing water bath sonication and then freeze-drying to obtain the target product, i.e., AUHA polymeric micelles.

[0021] The above preparation process has the advantages of simple operation, simple process and low manufacturing cost. The substitution degree of the amphiphilic copolymer can be adjusted by changing the feeding ratio and reaction time, thereby changing the particle size of the nanomicelles. The average particle size of the hyaluronic acid polymeric micelles modified with arginine and ursolic acid is 150 - 500 nm, the average potential is -27.3 ± 1.8 mV, and the critical micelle concentration is 21 μg / mL.

[0022] The present invention also provides a preparation method of the drug-loaded hyaluronic acid polymeric micelles modified with arginine and ursolic acid, comprising the following steps:

[0023] Dissolve AUHA in water, and while stirring at high speed, dropwise add the drug ethanol solution, stir vigorously, perform probe sonication for 30 min, dialyze, and filter through a membrane to remove the free drug to obtain the drug-loaded micelle solution.

[0024] Preferably, the concentration of AUHA is 3 - 8 mg / mL, the drug is a poorly soluble drug such as magnolol, paeonol, paclitaxel, docetaxel, carboplatin, cisplatin, oxaliplatin, gemcitabine, capecitabine, vincristine, hydroxycamptothecin, etc., the concentration of the drug ethanol solution is 10 - 25 mg / mL, the mass ratio of AUHA to the drug is 1 - 6:1, the time of vigorous stirring is 0.5 - 6 h, and the dialysis time is 12 - 48 h.

[0025] Compared with the prior art, the beneficial effects of the present invention are as follows:

[0026] The present invention modifies hyaluronic acid with ursolic acid and arginine. The prepared polymeric micelle carrier has the advantages of high drug loading capacity and high encapsulation efficiency. The drug-loaded micelles have good stability, effectively improving the solubility of poorly soluble drugs in water, and also solving the problems of toxicity, aging, high cost, etc. existing in current methods such as solid dispersion, liposome preparation, self-microemulsion drug delivery system, etc., providing a new idea for solving the solubility problem of poorly soluble drugs. BRIEF DESCRIPTION OF THE DRAWINGS

[0027] Figure 1 It is the infrared spectrum of hyaluronic acid modified with arginine and ursolic acid.

[0028] Figure 2 It is the NMR spectrum of hyaluronic acid modified with arginine and ursolic acid.

[0029] Figure 3 It is the particle size diagram of hyaluronic acid modified with arginine and ursolic acid.

[0030] Figure 4 Potential map of hyaluronic acid modified with arginine and ursolic acid.

[0031] Figure 5 Critical micelle concentration results of hyaluronic acid modified with arginine and ursolic acid.

[0032] Figure 6 In vitro release results of hyaluronic acid modified with arginine and ursolic acid. Detailed implementation manners

[0033] The present invention will be further described below in conjunction with embodiments for a comprehensive understanding of the present invention.

[0034] Example 1

[0035] Weigh 0.5 g of sodium hyaluronate (molecular weight 8000) and dissolve it in 60 mL of pure water. Add an appropriate amount of 0.904 g of NHS and 0.543 g of EDC, adjust the pH to 4 - 5, and carry out an activation reaction for 20 min. Then, dropwise add an aqueous solution of cystamine dihydrochloride and sodium bicarbonate at 6 mg / mL, adjust the pH to 7 - 8, and react under nitrogen protection in the dark for 48 h. After completion, transfer it to a dialysis bag with a molecular weight cut-off of 3500 and dialyze with pure water for 48 h, and then freeze-dry to obtain cystamine-modified hyaluronic acid (CYHA); weigh 0.5 g of ursolic acid and dissolve it in 50 mL of DMSO. Add an appropriate amount of 0.252 g of NHS and 0.420 g of EDC, carry out an activation reaction for 20 min, and dropwise add the CYHA suspension (0.293 g of CYHA powder dissolved in DMSO), and react under nitrogen protection in the dark for 48 h. After completion, transfer it to a dialysis bag with a molecular weight cut-off of 3500 and dialyze with pure water for 48 h, and then wash thoroughly with dichloromethane and freeze-dry to obtain ursolic acid-modified hyaluronic acid (UAHA); weigh 0.279 g of UAHA and dissolve it in DMSO. Add 0.33 g of EDC and 0.035 g of DMAP, stir evenly, add 0.5 g of arginine, adjust the pH to 7, and react under nitrogen protection in the dark for 48 h. After completion, transfer it to a dialysis bag with a molecular weight cut-off of 3500 and dialyze with pure water for 48 h, then carry out suction filtration, and freeze-dry the filtrate to obtain arginine and ursolic acid-modified hyaluronic acid (AUHA); dissolve 180 mg of AUHA in 30 ml of water and perform probe sonication to obtain the arginine and ursolic acid-modified hyaluronic acid polymer micelles.

[0036] Preparation of AUHA-loaded micelles: Weigh AUHA and dissolve it in deionized water. Sonicate it with a probe for 30 times at a power of 200 w, working for 2 s and pausing for 3 s to prepare a micelle solution with a concentration of 6 mg / mL. Take magnolol separately, add ethanol to it, and prepare a solution with a concentration of 25 mg / mL. Add the ethanol solution containing 25 mg / mL magnolol according to the dosage ratio of magnolol:AUHA of 25%. Stir it in the dark at room temperature for 2 h. After that, transfer it to a dialysis bag with a molecular weight cut-off of 3500 and dialyze it with pure water for 24 h. Collect the dialyzed product and centrifuge it at 8000 rpm for 10 min at low temperature to remove the magnolol not encapsulated by the micelles. Collect the supernatant to obtain hyaluronic acid-loaded micelles modified with arginine and ursolic acid (MAG-AUHA).

[0037] Determine the drug loading and encapsulation efficiency of magnolol in MAG-AUHA by HPLC. Take an appropriate amount of MAG-AUHA and dilute it with the mobile phase. The chromatographic column is Supersil ODS2 C18 (4.6×250 mm, 5 μm), the mobile phase is methanol:water (78:22, v / v), the flow rate is 0.8 mL / min, the detection wavelength is 292 nm, and the injection volume is 20 μL. Calculate the drug concentration in MAG-AUHA according to the standard curve.

[0038] Encapsulation efficiency = (mass of magnolol dosed - mass of free magnolol not encapsulated) / mass of magnolol dosed × 100%

[0039] Drug loading = mass of magnolol in the drug-loaded micelles / mass of micelles × 100%.

[0040] After calculation, the drug loading of MAG-AUHA is 20.0% and the encapsulation efficiency is 97.33%.

[0041] 1.1 Fourier transform infrared spectroscopy (FTIR)

[0042] The infrared spectrum was measured using a Nicolet 2000 Fourier transform infrared spectrometer (Nicolet, USA). The measurement method is as follows: Add about 1 - 2 mg of the freeze-dried sample of hyaluronic acid modified with arginine and ursolic acid prepared above to about 200 mg of ground and dried KBr powder. After thorough mixing, press it into a tablet and perform infrared spectrum scanning. Scanning range: 4000 cm -1 -400 cm -1 , record its infrared spectrum, as Figure 1 shown.

[0043] Among them, 1640 and 1735 cm -1 are the symmetric contraction peaks of amide bonds, 2927 cm -1 is the methyl peak, and 3434 cm -1 is the stretching vibration peak of hydroxyl and amino groups.

[0044] 1.2 Nuclear magnetic resonance ( 1 H-NMR)

[0045] The freeze-dried sample of hyaluronic acid modified with arginine and ursolic acid prepared above was dissolved in D2O, irradiated at a frequency of 150 MHz, and recorded at 20 °C using a Bruker (AVACE) AV 500 nuclear magnetic resonance spectrometer (Bruker, Germany), as Figure 2 shown.

[0046] Among them, the characteristic peaks of hyaluronic acid are at δ 1.88 (N-acetylmethyl) and δ 3.2 - 3.7 (methylene and hydroxyl), the characteristic peak of cystamine methylene is at δ 2.63 - 2.8, the characteristic peak of ursolic acid methyl is at 61.12, and the characteristic peak of arginine α-amino is at δ 3.07.

[0047] 1.3 Particle size and potential

[0048] Blank micelles and drug-loaded micelle solutions were prepared, and the particle size and surface potential of the drug-loaded micelles were measured using a nanoparticle size and zeta potential analyzer. The measured particle size of the drug-loaded micelles was 110.01 ± 2.57 nm, and the Zeta potential was -20.6 ± 2.4 mV, as Figure 3 and Figure 4 shown.

[0049] 1.4 Critical micelle concentration

[0050] The critical micelle concentration (CMC) of the hyaluronic acid polymer modified with arginine and ursolic acid was measured using a fluorescence probe technique. First, a pyrene solution (0.1 mmol / L) prepared in acetone was added to a test tube. After the acetone evaporated naturally, different volumes of the AUHA stock solution were added, and the volume was made up to 10 mL with pure water. The mass concentrations of the prepared solutions were 0.00001, 0.00001, 0.0001, 0.0002, 0.001, 0.002, 0.01, 0.02, 0.1, 0.5 mg / mL, respectively. After ultrasonic treatment for 30 min at room temperature, the solutions were equilibrated at 37 °C in the dark for 24 h. The fluorescence intensity was measured using a fluorescence spectrophotometer. The excitation wavelength was 346 nm, and the emission wavelength range was 350 - 600 nm. Pyrene would be encapsulated by the micelles, and the polarity of the environment changed, and the ratio of the intensities of the first band (373 nm) and the third band (384 nm) also changed. The value of I 373 / I 384 was fitted, and the intersection of the two tangents determined the CMC value to be 21 μg / mL, as Figure 5 .

[0051] 1.5 Determination of solubility

[0052] Weigh 0.02 g of magnolol and 0.1 g of MAG-AUHA (calculated according to a drug loading of 20%) separately, dissolve them in 2 mL of pure water, stir and dissolve at room temperature for 24 h until saturation is reached, remove insoluble solids with a 0.22-μm filter membrane, and determine the solubility of magnolol and MAG-AUHA in water by high-performance liquid chromatography. After determination, the solubilities of magnolol and MAG-AUHA in pure aqueous solution are 0.04 mg / mL and 5.5 mg / mL respectively. The solubility of MAG-AUHA is increased by about 137 times compared with the original drug magnolol.

[0053] 1.6 In vitro release

[0054] The in vitro drug release was evaluated by the dialysis method. 1 mL of free magnolol solution and MAG-AUHA solution were separately placed into dialysis bags (with a molecular weight cut-off of 3500), and immersed in PBS solutions with different pH values (1.5, 6.8, 7.8) containing 1% (w / v) Tween-80 at time periods of 0 - 2 h, 4 - 6 h, and 7 - 48 h. 2 mL of release medium was collected at different time intervals (0.5, 1, 2, 4, 10, 12, 24 h), and the cumulative release amount was detected by high-performance liquid chromatography to plot the release curve. As Figure 6 shown, the in vitro release rate of MAG-AUHA is slower than that of free magnolol. The release rate is faster in the first 4 h when simulating artificial gastric juice with pH 1.5, and the release amount reaches 5.73%. The release rate starts to slow down at 4 - 6 h when simulating artificial intestinal juice with pH 6.8, and the release amount reaches 12.21%. The release rate is slow at 7 - 24 h when simulating artificial colon juice with pH 7.8, and the cumulative release rate reaches 20.29%, indicating that MAG-AUHA has the ability of slow and sustained drug release.

[0055] Example 2

[0056] Weigh 0.25 g of sodium hyaluronate (molecular weight 8000) and dissolve it in 60 mL of pure water. Add an appropriate amount of 0.904 g of NHS and 0.543 g of EDC, adjust the pH to 4 - 5, and carry out an activation reaction for 20 min. Then, dropwise add a cystamine dihydrochloride and sodium bicarbonate aqueous solution with a concentration of 6 mg / mL, adjust the pH to 7 - 8, and carry out a reaction under nitrogen protection in the dark for 48 h. After completion, transfer it to a dialysis bag with a molecular weight cut-off of 3500, dialyze with pure water for 48 h, and freeze-dry to obtain cystamine-modified hyaluronic acid (CYHA); Weigh 0.25 g of ursolic acid and dissolve it in 50 mL of dimethyl sulfoxide. Add an appropriate amount of 0.252 g of NHS and 0.420 g of EDC, carry out an activation reaction for 20 min, and dropwise add the CYHA suspension (0.293 g of CYHA powder dissolved in DMSO). Carry out a reaction under nitrogen protection in the dark for 48 h. After completion, transfer it to a dialysis bag with a molecular weight cut-off of 3500, dialyze with pure water for 48 h, wash it thoroughly with dichloromethane, and freeze-dry to obtain ursolic acid-modified hyaluronic acid (UAHA); Weigh 0.279 g of UAHA and suspend it in pure water. Add 0.33 g of EDC and 0.035 g of DMAP, stir evenly, add 0.5 g of arginine, adjust the pH to 7, and carry out a reaction under nitrogen protection in the dark for 48 h. After completion, transfer it to a dialysis bag with a molecular weight cut-off of 3500, dialyze with pure water for 48 h, carry out suction filtration, and freeze-dry the filtrate to obtain an arginine and ursolic acid-modified hyaluronic acid polymer (AUHA); Dissolve 180 mg of AUHA in 30 mL of water and carry out probe sonication to obtain the AUHA polymer micelles.

[0057] Preparation of MAG-AUHA drug-loaded micelles: Weigh AUHA and dissolve it in deionized water. Carry out probe sonication 30 times with a power of 200 w, working for 2 s and stopping for 3 s to prepare a micelle solution with a concentration of 6 mg / mL. Separately, take honokiol, add ethanol, and prepare a solution with a concentration of 25 mg / mL. Add the ethanol solution containing 25 mg / mL of honokiol according to the dosing amount with a mass ratio of honokiol:AUHA of 25%, stir in the dark at room temperature for 2 h. After completion, transfer it to a dialysis bag with a molecular weight cut-off of 3500, dialyze with pure water for 24 h, collect the dialyzed product, and centrifuge it at 8000 rpm at low temperature for 10 min to remove the honokiol not encapsulated by the micelles. Collect the supernatant to obtain the arginine and ursolic acid-modified hyaluronic acid drug-loaded micelles (MAG-AUHA).

[0058] Determine the drug loading and encapsulation efficiency of honokiol in the drug-loaded micelles by HPLC. Take MAG-AUHA, dilute it with the mobile phase. The chromatographic column is Supersil ODS2 C18 (4.6×250 mm, 5 μm), the mobile phase is methanol:water (78:22, v / v), the flow rate is 0.8 mL / min, the detection wavelength is 292 nm, and the injection volume is 20 μL. Calculate the drug concentration in MAG-AUHA according to the standard curve.

[0059] Entrapment efficiency = (mass of magnolol administered - mass of free magnolol not encapsulated) / mass of magnolol administered × 100% Drug loading = mass of magnolol in micelles / mass of micelles × 100%.

[0060] The drug loading of MAG-AUHA was calculated to be 18.42% and the entrapment efficiency was 97.2%.

[0061] Blank micelles and drug-loaded micelle solutions were prepared, and the particle size and surface potential of the drug-loaded micelles were measured using a nanoparticle size potentiometer. The measured particle size of the drug-loaded micelles was 113.01 ± 2.17 nm, and the Zeta potential was -22.6 ± 1.4 mV.

Claims

1. A hyaluronic acid polymer modified with arginine and ursolic acid, the chemical structure of which is shown in general formula I:

2. The hyaluronic acid polymer modified with arginine and ursolic acid according to claim 1, characterized in that In the polymer, the molecular weight of sodium hyaluronate is 1-570 kDa, the grafting rate of cystamine is 26-54%, the grafting rate of ursolic acid is 1-31%, and the grafting rate of arginine is 2-30%.

3. The method for synthesizing the hyaluronic acid polymer modified with arginine and ursolic acid according to claim 1 or 2, characterized in that: The following steps are involved: (1) preparing a sodium hyaluronate aqueous solution, adding EDC and NHS, adjusting the pH to a range of 4 to 5, and activating the carboxyl group; (2) preparing an aqueous solution of cystamine dihydrochloride, adding sodium bicarbonate to carry out a neutralization reaction, adding the aqueous solution of cystamine dropwise to the activated aqueous solution of sodium hyaluronate, adjusting the pH to a range of 7 to 8, passing nitrogen gas to avoid light, dialyzing with deionized water after the reaction, and freeze-drying to obtain cystamine-modified hyaluronic acid (CYHA); (3) preparing an ursolic acid solution, adding EDC and NHS, activating the carboxyl group, and then dropwise adding a suspension of CYHA in water, passing nitrogen gas to avoid light, and after the reaction, dialyzing with deionized water, washing with dichloromethane, and freeze-drying to obtain ursolic acid-modified hyaluronic acid (UAHA); (4) Prepare an arginine aqueous solution, add EDC and DMAP, add UAHA solution, adjust the pH to 7, pass nitrogen gas to avoid light, dialyze with deionized water after the reaction, and freeze-dry to obtain arginine and ursolic acid modified hyaluronic acid (AUHA).

4. The method for preparing the hyaluronic acid polymer modified by arginine and ursolic acid according to claim 3, characterized in that: In the step (1), the molar ratio of sodium hyaluronate, EDC, and NHS is 0.25-2:1-8:1-4; the concentration of sodium hyaluronate is 1-8 mg / mL; and the activation time of the carboxyl group is 20-30 min.

5. The method for preparing the hyaluronic acid polymer modified by arginine and ursolic acid according to claim 3, characterized in that: In the step (2), the molar ratio of sodium hyaluronate, cystamine dihydrochloride and sodium bicarbonate is 0.25-2:0.5-20:1-40; the light-proof reaction time is 40-56 hours, and the deionized water dialysis time after the reaction is 48-72 hours.

6. The method for preparing the hyaluronic acid polymer modified by arginine and ursolic acid according to claim 3, characterized in that: In the step (3), the ursolic acid solution is ursolic acid dissolved in DMSO, the concentration is 1-30 mg / mL, the molar ratio of ursolic acid, EDC, and NHS is 0.25-3:1-12:4-12, the molar ratio of ursolic acid to CYHA is 1-2:1, the activation time of the carboxyl group is 20-30 minutes, the nitrogen-proof light-proof reaction time is 40-56 hours, and the deionized water dialysis time after the reaction is completed is 48-72 hours.

7. The method for preparing the hyaluronic acid polymer modified by arginine and ursolic acid according to claim 3, characterized in that: In the step (4), the concentration of arginine is 1-300 mg / mL, the molar ratio of arginine, EDC and DMAP is 0.25-2:1-5:0.05-0.3, the molar ratio of arginine to UAHA is 1-2:1, the UAHA solution is UAHA dissolved in DMSO with a concentration of 1-30 mg / mL, the reaction time in nitrogen-proof and light-proof is 40-56 h, and the deionized water dialysis time after the reaction is 48-72 h.

8. The drug-loaded micelles prepared by the hyaluronic acid polymer modified with arginine and ursolic acid according to claim 1 or 2, characterized in that: The following steps are involved: Dissolve AUHA in water, add the drug ethanol solution dropwise under high-speed stirring, stir vigorously, perform probe ultrasound for 30 minutes, dialyze, and filter membrane to remove free drugs to obtain drug-loaded micelles (MAG-AUHA).

9. The method for preparing drug-loaded micelles according to claim 8, characterized in that: The concentration of the AUHA solution is 3-8 mg / mL, the drugs are poorly soluble drugs such as magnolol, paeonol, paclitaxel, docetaxel, carboplatin, cisplatin, oxaliplatin, gemcitabine, capecitabine, vincristine, hydroxycamptothecin, etc., the concentration of the drug ethanol solution is 10-25 mg / mL, the mass ratio of the AUHA and the drug is 1-6:1, the time of the vigorous stirring is 0.5-6 h, and the dialysis time is 12-48 h.

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