A method for constructing a drug-loaded Pickering emulsion based on S N 2 Reaction of amphiphilic alginate derivatives / organic montmorillonite synergistically stabilized drug-loaded Pickering emulsion

Abstract

The application provides a kind of based on S N 2 reaction amphiphilic alginate derivative / organic montmorillonite synergistically stabilized drug-loaded Pickering emulsion construction method belongs to the technical field of hydrophobic anti-inflammatory drug preparation.The application uses chlorinated cholesterol as a hydrophobic modifier, prepares amphiphilic cholesteric alginate derivative through S N 2 reaction, alkyl glycoside is used as a modifier to prepare organic montmorillonite with certain interfacial activity, amphiphilic alginate derivative and OMMT are compounded to construct a new particle emulsifier, then the particle suspension of amphiphilic alginate derivative / OMMT compound is used as water phase, liquid paraffin dissolving anti-inflammatory drugs is used as oil phase, and the drug-loaded Pickerng emulsion stabilized by amphiphilic alginate derivative and OMMT is prepared.

Description

TECHNICAL FIELD

[0001] The present application belongs to the technical field of hydrophobic anti-inflammatory drug formulations, and particularly relates to a S N 2 Construction method of reaction amphiphilic alginate derivative / organic montmorillonite synergistically stabilized drug-loaded Pickering emulsion. BACKGROUND

[0002] Particle emulsifiers are one of the important components of Pickering emulsion, which has an important influence on the structure and stability of Pickering emulsion. With the in-depth study of Pickering emulsion, more and more particle emulsifiers have been studied to stabilize Pickering emulsion. According to the different types of particles, it can be divided into inorganic particles, organic particles, biological particles and Janus particles. According to the degree of deformation of the particles, it can be divided into rigid particles and soft particles. Rigid particles refer to a kind of particles with strong pressure resistance, and the elastic modulus is in the range of GPa, including inorganic particles, inorganic Janus particles and surface modified or hybrid solid particles. Soft particles refer to particles with easy deformation characteristics, and the elastic modulus is in the range of KPa, mainly including organic particles, biological particles, polymer Janus particles, polymer micelles and polymer microgels. In addition, according to the composition of the particles, it can be divided into single particles and composite particles. However, some single particles have the disadvantages of low solubility, easy aggregation and easy crosslinking, which limit their application in the functional application of Pickering emulsion. Therefore, the development of new composite particles as emulsifiers has gradually become the research hotspot of researchers.

[0003] Sodium alginate (SA) is a natural anionic polysaccharide, which has been widely used and explored in tissue engineering, drug delivery, wound healing, textiles, cosmetics and food science, etc. due to its excellent properties such as renewability, biocompatibility, biodegradability, antibacterial activity, non-immunogenicity, gelling ability and easy processing. However, due to its own swelling, strong hydrophilicity, low affinity for hydrophobic drugs and other shortcomings, its application in the pharmaceutical field is limited.

[0004] The amphiphilic alginate derivative as an emulsifier forms a thick hydrophilic coating on the surface of the oil droplets, making the oil droplets relatively stable to changes in ionic strength, pH value or temperature. At the same time, sodium alginate can be used as a thickening agent to further stabilize the emulsion. Inorganic minerals have unique nanostructure, hydrophilicity and biocompatibility. They can form a particle layer at the oil-water interface to prevent the aggregation of emulsion droplets. However, the existing composite particles prepared by using amphiphilic alginate derivatives and inorganic minerals for synergistically stabilizing Pickering emulsion cannot obtain Pickering emulsion with high stability.

[0005] Therefore, there is an urgent need for a kind of alginate-based polymer / solid particles of Pickering emulsion to solve the above problems. SUMMARY

[0006] To solve the above technical problems, the present application provides a kind of based on S N 2 reaction amphiphilic alginate derivative / organic montmorillonite synergistically stabilized drug-loaded Pickering emulsion construction method.

[0007] To achieve the above object, the present application provides the following technical solutions:

[0008] The present application provides a kind of based on S N 2 reaction amphiphilic alginate derivative / organic montmorillonite synergistically stabilized drug-loaded Pickering emulsion construction method, comprising the following steps:

[0009] (1) adding tetrabutylammonium hydroxide aqueous solution to alginate aqueous solution for neutralization reaction, after filtration, dialysis and freeze-drying, obtain alginate tetrabutylammonium salt TBA-Alg;

[0010] (2) the alginate tetrabutylammonium salt TBA-Alg obtained in step (1) is mixed with tetrabutylammonium fluoride solution, then cholesteryl chloride is added to carry out S N 2 reaction, then NaCl aqueous solution is added for displacement reaction, after precipitation, centrifugation, alcohol washing, centrifugation again, dialysis and freeze-drying, cholesteryl grafted alginate derivative DAD is obtained;

[0011] (3) sodium-based montmorillonite is placed in a ball mill tank, then alkyl glycoside aqueous solution is added for ball milling, after centrifugation, washing, freeze-drying, grinding and sieving, alkyl glycoside modified organic montmorillonite OMMT is obtained;

[0012] (4) the cholesteryl grafted alginate derivative DAD obtained in step (2) and the alkyl glycoside modified organic montmorillonite OMMT obtained in step (3) are mixed to obtain DAD / OMMT composite particle;The DAD / OMMT composite particle is added to water and dispersed by ultrasonic and shaken and placed to obtain DAD / OMMT composite particle suspension;

[0013] (5) the mixed solution of hydrophobic anti-inflammatory drug and liquid paraffin is used as oil phase, and the DAD / OMMT composite particle suspension obtained in step (4) is used as water phase, then the two are mixed and subjected to shear emulsification to obtain DAD / OMMT composite particle synergistically stabilized drug-loaded Pickering.

[0014] Preferably, in step (1), the amount of tetrabutylammonium hydroxide aqueous solution is used to adjust the pH value of alginate aqueous solution to 7-10.

[0015] Preferably, in step (2), the molar ratio of the chlorinated cholesterol to the uronic acid monomer of tetrabutylammonium alginate (TBA-Alg) is (1:1)-(3:10).

[0016] Preferably, in step (3), the CEC of the sodium-based montmorillonite is 1.20 mmol / g; the alkyl glycoside includes one or more of hexyl glycoside, octyl glycoside, decyl glycoside and dodecyl glycoside; and the amount of the alkyl glycoside used is 0.5-2.5 times the CEC of the sodium-based montmorillonite.

[0017] Preferably, in step (4), the mass ratio of DAD to OMMT in the DAD / OMMT composite particles is (1:2)-(1:10).

[0018] Preferably, in step (4), the mass fraction of the DAD / OMMT composite particles in the DAD / OMMT composite particle suspension is 0.1-1.5%.

[0019] Preferably, in step (5), the ratio of the hydrophobic anti-inflammatory drug to the liquid paraffin in the mixed solution of the hydrophobic anti-inflammatory drug and the liquid paraffin is 1.0-5.0 mg / mL.

[0020] Preferably, in step (5), the hydrophobic anti-inflammatory drug includes one or more of ibuprofen, ofloxacin and ciprofloxacin.

[0021] Preferably, in step (5), the volume ratio of the mixed solution of the hydrophobic anti-inflammatory drug and the liquid paraffin to the DAD / OMMT composite particle suspension is (1:9)-(4:6).

[0022] The application also provides a S N 2. A Pickering emulsion loaded with a reaction amphiphilic alginate derivative / organic montmorillonite synergistically stabilized drug.

[0023] Compared with the prior art, the application has the following advantages and technical effects:

[0024] In the application, chlorinated cholesterol is used as a hydrophobic modifier, and after the protonated alginate is converted into tetrabutylammonium alginate (TBA-Alg) by tetrabutylammonium hydroxide, a homogeneous solution is formed by adding a tetrabutylammonium fluoride solution, and then the S N2The amphiphilic cholesteric alginate derivative (DAD) is prepared by reaction. Meanwhile, the organic montmorillonite (OMMT) with certain interfacial activity is prepared by wet ball milling with the green non-ionic surfactant alkyl glycoside as the modifier. Further, the new type of particle emulsifier is constructed by compounding the amphiphilic alginate derivative and the OMMT. Then, the Pickering emulsion stabilized by the amphiphilic alginate derivative and the OMMT is prepared by taking the water dispersion system of the amphiphilic alginate derivative / OMMT complex particles as the water phase and the liquid paraffin dissolving the anti-inflammatory drug as the oil phase. The Pickering emulsion stabilized by the amphiphilic alginate derivative and the OMMT prepared in the application shows higher loading property and slow release behavior of the hydrophobic anti-inflammatory drug, has certain pH-responsive release behavior, and makes the cumulative release rate in the neutral environment lower. Meanwhile, the self-diffusion and swelling degradation of the drug in the Pickering emulsion stabilized by the amphiphilic alginate derivative and the OMMT are simultaneously carried out, and they jointly control the release rate of the hydrophobic drug. Based on the above results, the Pickering emulsion stabilized by the amphiphilic alginate derivative and the OMMT can be used as a hydrophobic drug delivery system with good slow-release property, and applied to the technical field of hydrophobic anti-inflammatory drug preparation. BRIEF DESCRIPTION OF DRAWINGS

[0025] The accompanying drawings, which form a part of this application, are included to provide a further understanding of the application and are incorporated in and constitute a part of this application. The embodiments illustrated in the drawings are provided to explain the present application and are not intended to limit the present application. In the drawings:

[0026] Figure 1 The synthetic route of DAD in the application is shown in the figure;

[0027] Figure 2 The FT-IR spectra (a) and H NMR spectra (b) of alginate SA and DAD in the embodiments 1-6 of the application are shown in the figures; 1 H NMR spectra (b);

[0028] Figure 3 The SEM images of sodium-based montmorillonite (a) and OMMT (b) in the embodiments 1-6 of the application are shown in the figures;

[0029] Figure 4 The TEM images of sodium-based montmorillonite (a) and OMMT (b) in the embodiments 1-6 of the application are shown in the figures;

[0030] Figure 5FT-IR spectra (a), XRD spectra (b), TGA graphs (c) of DAD, OMMT and DAD / OMMT composite particles in Examples 1-6, and particle size (d), Zeta potential (e) and surface tension (f) of DAD / OMMT composite particle suspensions in Comparative Example 1, Example 1 and Examples 5-6;

[0031] Figure 6 Emulsion destabilization index of stable emulsion of Pickering emulsion synergistically stabilized by ibuprofen-loaded DAD / OMMT composite particles in Comparative Example 1, Example 1 and Examples 5-6;

[0032] Figure 7 Actual photos of stable emulsion of Pickering emulsion synergistically stabilized by ibuprofen-loaded DAD / OMMT composite particles in Comparative Example 1, Example 1 and Examples 5-6 after standing for different time;

[0033] Figure 8 Plot of instability coefficient versus time of stable emulsion of Pickering emulsion synergistically stabilized by ibuprofen-loaded DAD / OMMT composite particles in Comparative Example 1, Example 1 and Examples 5-6;

[0034] Figure 9 Microscope photos, actual photos and size distribution plots of Pickering emulsion synergistically stabilized by ibuprofen-loaded DAD / OMMT composite particles in Comparative Example 1, Example 1 and Examples 5-6 after standing for 3h;

[0035] Figure 10 Actual photos and laser confocal electron microscope photos of Pickering emulsion synergistically stabilized by ibuprofen-loaded DAD / OMMT composite particles in Example 1 and Examples 5-6 after staining with Nile red and Rhodamine B, respectively;

[0036] Figure 11 In vitro cytotoxicity of Pickering emulsion synergistically stabilized by ibuprofen-loaded DAD / OMMT composite particles in Example 1 after incubation with RAW264.7 cells for 48h;

[0037] Figure 12 SEM photos of Pickering emulsion synergistically stabilized by ibuprofen-loaded DAD / OMMT composite particles in Comparative Example 1, Example 1 and Examples 5-6 (with 2.0 mol% AIBN of styrene as oil phase);

[0038] Figure 13Rheological property chart of the Pickering emulsion synergistically stabilized by the ibuprofen-loaded DAD / OMMT composite particles in Comparative Example 1, Example 1 and Examples 5-6;

[0039] Figure 14 Drug release curve of ibuprofen from the Pickering emulsion synergistically stabilized by the ibuprofen-loaded DAD / OMMT composite particles in Example 5 and Example 6. DETAILED DESCRIPTION

[0040] The technical solutions in the embodiments of the present application will be apparently and completely described below with the drawings in the embodiments of the present application. Obviously, the described embodiments are only part of the embodiments of the present application, rather than all the embodiments. Based on the embodiments in the present application, all the other embodiments obtained by a person of ordinary skill in the art without any creative work belong to the scope of protection of the present application.

[0041] In order to make the above objectives, characteristics and advantages of the present application more apparent and easy to understand, the present application will be further described in detail below with the drawings and specific embodiments.

[0042] The embodiments of the present application provide a kind of S N 2 reaction amphiphilic alginate derivative / organic montmorillonite synergistically stabilized drug-loaded Pickering emulsion construction method, comprising the following steps:

[0043] (1) preparation of tetrabutylammonium alginate TBA-Alg: adding tetrabutylammonium hydroxide aqueous solution to alginate aqueous solution for neutralization reaction, filtering, dialysis and freeze-drying to obtain tetrabutylammonium alginate TBA-Alg;

[0044] (2) preparation of cholesterol-based grafted alginate derivative DAD: mixing tetrabutylammonium alginate TBA-Alg obtained in step (1) with tetrabutylammonium fluoride solution, then adding cholesterol chloride for S N 2 reaction, and then adding NaCl aqueous solution for displacement reaction, precipitating, centrifuging, alcohol washing, centrifuging again, dialysis and freeze-drying to obtain cholesterol-based grafted alginate derivative DAD;

[0045] (3) preparation of alkyl glycoside modified organic montmorillonite OMMT: placing sodium-based montmorillonite in a ball mill jar, then adding alkyl glycoside aqueous solution for ball milling, centrifuging, washing, freeze-drying, grinding and sieving to obtain alkyl glycoside modified organic montmorillonite OMMT;

[0046] (4) Preparation of DAD / OMMT composite particle suspension: the DAD and OMMT obtained in steps (2) and (3) are mixed to obtain DAD / OMMT composite particles; the DAD / OMMT composite particles are added to water and dispersed by ultrasonic and left to stand by oscillation to obtain a DAD / OMMT composite particle suspension;

[0047] (5) Preparation of DAD / OMMT composite particle synergistically stabilized drug-loaded Pickering: a mixed solution of hydrophobic anti-inflammatory drug and liquid paraffin is used as the oil phase, and the DAD / OMMT composite particle suspension obtained in step (4) is used as the water phase; the two are mixed and then subjected to shear emulsification to obtain a DAD / OMMT composite particle synergistically stabilized drug-loaded Pickering.

[0048] In a preferred embodiment, in step (1), the amount of the aqueous solution of tetrabutylammonium hydroxide is used to adjust the pH of the aqueous alginate solution to 7-10.

[0049] In a preferred embodiment, in step (1), the mass average molecular weight Mw of the alginate in the aqueous alginate solution is ≤100000, and the molar ratio of monomer guluronic acid (G) to mannuronic acid (M) is ≤0.8.

[0050] In a preferred embodiment, in step (1), the amount ratio of alginate to water in the aqueous alginate solution is 4.0 g:100 mL.

[0051] In a preferred embodiment, in step (1), the amount ratio of tetrabutylammonium hydroxide TBAOH to water in the aqueous solution of tetrabutylammonium hydroxide is 0.25 g:1.00 mL.

[0052] In a preferred embodiment, in step (1), the filter used for filtration is a Buchner funnel. The present application removes insoluble particulate impurities by filtration.

[0053] In a preferred embodiment, in step (1), the molecular weight cut-off of the dialysis bag used for dialysis is 3500, the dialysis fluid is double-distilled water, and the dialysis time is 3 days. The present application removes unreacted small molecules by dialysis.

[0054] In a preferred embodiment, in step (1), the temperature for freeze-drying is -60 to -50°C, and the time is 48 h.

[0055] In a preferred embodiment, in step (2), the solvent of the tetrabutylammonium fluoride solution is N,N-dimethylformamide; and the amount ratio of tetrabutylammonium fluoride TBAF to N,N-dimethylformamide DMF in the tetrabutylammonium fluoride solution is 0.8 g:80 mL.

[0056] In a preferred embodiment, in step (2), the mass ratio of the tetrabutylammonium alginate TBA-Alg to tetrabutylammonium fluoride in the tetrabutylammonium fluoride solution is 1.2 g:0.8 g.

[0057] In a preferred embodiment, in step (2), the S N 2The reaction temperature is room temperature, and the reaction time is 24 h.

[0058] In a preferred embodiment, in step (2), the molar ratio of the choline chloride to the uronic acid monomer in the tetrabutylammonium alginate TBA-Alg is (1:1)-(3:10), more preferably (1:2)-(3:10).

[0059] In a preferred embodiment, in step (2), the concentration of the NaCl aqueous solution is 2.5 mol / L.

[0060] In a preferred embodiment, in step (2), the time of the displacement reaction is 2 h. The present application displaces the TBA + .

[0061] In a preferred embodiment, in step (2), the reagent used for the precipitation is ethyl acetate, and the amount of the ethyl acetate is 5 times the volume of the reaction solution; the molecular weight cut-off of the dialysis bag used for the dialysis is 3500, the dialysis liquid used for the dialysis is double-distilled water, and the dialysis time is 3 d; the temperature of the freeze-drying is-60 to-50℃, and the time is 48 h.

[0062] In a preferred embodiment, in step (2), the degree of substitution of the DAD is 27.2-39.4%.

[0063] In a preferred embodiment, in step (2), the degree of substitution of the DAD is calculated by a saponification reaction; specifically, 50 mg of the DAD is weighed into a conical flask, 15 mL of a 0.03 mol / L NaOH standard solution is added, and the mixture is stirred overnight to allow a full reaction, then 0.02 mol / L HCl standard solution is used for back titration of the residual NaOH, the amount of substance of the NaOH consumed in the saponification reaction is calculated, and thus the DS of the DAD is obtained.

[0064] In a preferred embodiment, in step (3), the CEC of the sodium-based montmorillonite is 1.20 mmol / g; and the pretreatment method of the sodium-based montmorillonite comprises drying to a constant weight in an oven at 110℃.

[0065] In the preferred embodiments, in step (3), the alkyl glycoside includes one or more of hexyl glycoside, octyl glycoside, decyl glycoside and dodecyl glycoside, more preferably hexyl glycoside and / or dodecyl glycoside; the amount of alkyl glycoside used is 0.5-2.5 times the CEC of the sodium-based montmorillonite, more preferably 0.5-2.0 times the CEC of the sodium-based montmorillonite.

[0066] In the preferred embodiments, in step (3), the amount ratio of alkyl glycoside to water in the aqueous alkyl glycoside solution is (0.42-1.68) g:10 mL.

[0067] In the preferred embodiments, in step (3), when the amount of the sodium-based montmorillonite is 2.0 g, the grinding balls used for the ball milling include 2 maroon balls with a diameter of 20 mm, 8 maroon balls with a diameter of 10 mm and 12 maroon balls with a diameter of 6 mm.

[0068] In the preferred embodiments, in step (3), the rotation speed of the ball milling is 500 r / min, the ball milling time is 1 h, and the ball milling device is a PM planetary ball mill.

[0069] In the preferred embodiments, in step (3), the rotation speed of the centrifugation is 7000 r / min; the washing reagent is clean water; the temperature of the freeze-drying is -60 to -50℃, and the time is 48 h.

[0070] In the preferred embodiments, in step (4), the mass ratio of DAD to OMMT in the DAD / OMMT composite particles is (1:2)-(1:10), more preferably (1:2)-(1:5).

[0071] In the preferred embodiments, in step (4), the mass fraction of the DAD / OMMT composite particles in the DAD / OMMT composite particle suspension is 0.1-1.5%, more preferably 0.5-1.5%.

[0072] In the preferred embodiments, in step (4), the ultrasonic power of the ultrasonic dispersion is 100-200 W, and the ultrasonic mode is probe ultrasonic; the time of the oscillation and standing is more than 12 h.

[0073] In the preferred embodiments, in step (5), the amount ratio of the hydrophobic anti-inflammatory drug to the liquid paraffin in the mixed solution of the hydrophobic anti-inflammatory drug and the liquid paraffin is 1.0-5.0 mg / mL, more preferably 2.5-5.0 mg / mL.

[0074] In the preferred embodiments, in step (5), the hydrophobic anti-inflammatory drug includes one or more of ibuprofen, ofloxacin and ciprofloxacin, more preferably ibuprofen.

[0075] In a preferred embodiment, in step (5), the volume ratio of the mixed solution of the hydrophobic anti-inflammatory drug and liquid paraffin to the suspension of the DAD / OMMT composite particles is (1:9)-(4:6), more preferably (2:8)-(4:6).

[0076] In a preferred embodiment, in step (5), the shearing emulsification is performed at a speed of 12000r / min, at room temperature, for 2-3min, using an emulsifying machine.

[0077] The application also provides the S N 2 The Pickering emulsion stabilized by the amphiphilic alginate derivative / organic montmorillonite synergistically.

[0078] The application provides the S N 2 The Pickering emulsion stabilized by the amphiphilic alginate derivative / organic montmorillonite synergistically.

[0079] The application provides the S N 2 The Pickering emulsion stabilized by the amphiphilic alginate derivative / organic montmorillonite synergistically.

[0080] In the embodiment of the application, room temperature refers to 25±2℃.

[0081] Unless otherwise specified, the raw materials in the embodiments of the application are obtained by commercial purchase.

[0082] Embodiment 1

[0083] The application provides a method for constructing a S N 2 The Pickering emulsion stabilized by the amphiphilic alginate derivative / organic montmorillonite synergistically.

[0084] (1) Under the action of magnetic stirring, 4.0g of alginate is dispersed in 100mL of water to obtain an alginate aqueous solution, TBAOH aqueous solution (the amount ratio of TBAOH to water in the TBAOH aqueous solution is 0.25g:1.00mL) is added dropwise to the alginate aqueous solution for neutralization reaction, the pH value is adjusted to 8-10, then insoluble particle impurities are filtered by using a Buchner funnel, then the filtrate is loaded into a dialysis bag with a molecular weight cut-off of 3500, dialysis is performed in twice-distilled water for 3d, and then-60℃ freeze-drying is performed for 48h to obtain TBA-Alg.

[0085] (2) Put 1.2 g of dry TBA-Alg obtained in step (1) into 80 mL of DMF in which 0.8 g of TBAF is dissolved, and obtain a homogeneous solution by electric stirring; add 0.67 g of cholesteryl chloride (the molar ratio of cholesteryl chloride to uronic acid monomers in TBA-Alg is 1:2) into the homogeneous solution, continuously stir the reaction at room temperature for 24 h, then add 40 mL of 2.5 mol / L NaCl aqueous solution, continue to stir for 2 h, then add 5 times volume of ethyl acetate to precipitate the reaction product, centrifuge, alcohol wash, centrifuge again, and then transfer into a dialysis bag with a molecular weight cut-off of 3500, dialyze in double-distilled water for 3 d, and freeze dry at -60℃ for 48 h to obtain DAD with a degree of substitution DS of 39.4%.

[0086] (3) Dry Na-montmorillonite with a CEC of 1.20 mmol / g in an oven at 110℃ to constant weight, take 2 g of the Na-montmorillonite and put it into a 50 mL ball mill jar; mix 0.84 g of dodecyl glucoside (1.0 times the CEC of Na-montmorillonite) with 10 mL of deionized water to obtain a dodecyl glucoside aqueous solution, then add the dodecyl glucoside aqueous solution dropwise into the ball mill jar, use a PM planetary ball mill as the ball milling equipment, and use agate balls as the ball milling medium; add 2 agate balls with a diameter of 20 mm, 8 agate balls with a diameter of 10 mm, and 12 agate balls with a diameter of 6 mm into the ball mill jar, and set the rotation speed of the main disc to 500 r / min, and ball mill for 1 h; after the ball milling is completed, transfer the reaction solution into a beaker, wash the ball mill jar and the agate balls with deionized water, collect the washing liquid into the beaker, then centrifuge at 7000 r / min, repeat the washing and centrifuging with clean water for 3 times, freeze dry the precipitate obtained by centrifugation at -60℃ for 48 h, grind with an agate mortar, and sieve to obtain OMMT.

[0087] (4) Mix DAD obtained in step (2) and OMMT obtained in step (3) according to a mass ratio of 1:10 to obtain DAD / OMMT composite particle; take 80 mg of the DAD / OMMT composite particle and add it into 8 mL of deionized water, and perform ultrasonic dispersion at 100 W, and shake and stand for more than 12 h to obtain a DAD / OMMT composite particle suspension with a mass fraction of 1.0%.

[0088] (5) Take 10.0 mg of ibuprofen and add it into 2 mL of liquid paraffin to prepare a mixed solution of ibuprofen and liquid paraffin with a concentration of 5.0 mg / mL; mix the mixed solution of ibuprofen and liquid paraffin with the DAD / OMMT composite particle suspension obtained in step (4) according to a volume ratio of 2:8, then perform high-speed shearing at 12000 r / min at room temperature for 3 min by using an emulsifying machine to obtain a Pickering emulsion of DAD / OMMT composite particle loaded with ibuprofen.

[0089] Example 2

[0090] A method for constructing a drug-loaded Pickering emulsion based on S N 2 A method for constructing a drug-loaded Pickering emulsion based on a reaction amphiphilic alginate derivative / organic montmorillonite synergistic stabilization:

[0091] Step (1) is the same as in Example 1.

[0092] (2) 1.2 g of the dry TBA-Alg obtained in step (1) was added to 80 mL of DMF in which 0.8 g of TBAF was dissolved, and an electrically driven stirring was performed to obtain a homogeneous solution; 0.40 g of cholesteryl chloride (the molar ratio of cholesteryl chloride to the uronic acid monomer in TBA-Alg was 3:10) was added to the homogeneous solution, and the stirring was continuously performed at room temperature for 24 h, then 40 mL of a 2.5 mol / L NaCl aqueous solution was added, and the stirring was continuously performed for 2 h, then 5 times the volume of ethyl acetate was added to precipitate the reaction product, the product was centrifuged, washed with alcohol, and centrifuged again, and then transferred into a dialysis bag with a molecular weight cut-off of 3500, and dialyzed in double-distilled water for 3 d, and then freeze-dried at -60 °C for 48 h to obtain DAD with a degree of substitution DS of 27.2%.

[0093] (3) Sodium-based montmorillonite with a CEC of 1.20 mmol / g was dried in an oven at 110 °C to a constant weight, and 2 g of the sodium-based montmorillonite was weighed and placed in a 50 mL ball mill jar; 0.84 g of dodecyl glucoside (1.0 times the CEC of the sodium-based montmorillonite) was mixed with 10 mL of deionized water to obtain a dodecyl glucoside aqueous solution, and the dodecyl glucoside aqueous solution was added dropwise to the ball mill jar, then a PM type planetary ball mill was used as a ball milling device, and agate balls were used as ball milling media, 2 agate balls with a diameter of 20 mm, 8 agate balls with a diameter of 10 mm, and 12 agate balls with a diameter of 6 mm were added to the ball mill jar, the rotation speed of the main disc was 500 r / min, and the ball milling was performed for 1 h, after the ball milling was completed, the reaction solution was transferred to a beaker, and the ball mill jar and the agate balls were washed with deionized water, the washing liquid was collected into the beaker, and then centrifuged at 7000 r / min, the washing and centrifugation were repeated three times, the precipitate obtained by centrifugation was freeze-dried at -60 °C for 48 h, ground in an agate mortar, and sieved to obtain OMMT.

[0094] (4) The DAD obtained in step (2) and the OMMT obtained in step (3) were mixed according to a mass ratio of 1:5 to obtain DAD / OMMT composite particles; 80 mg of the DAD / OMMT composite particles was weighed and added to 8 mL of deionized water, and ultrasonic dispersion was performed at 100 W, and the system was shaken and allowed to stand for more than 12 h to obtain a DAD / OMMT composite particle suspension with a mass fraction of 1.0%.

[0095] (5) Weigh 10.0 mg of ibuprofen into 2 mL of liquid paraffin to prepare a mixed solution of ibuprofen and liquid paraffin with a concentration of 5.0 mg / mL; mix the above mixed solution of ibuprofen and liquid paraffin as the oil phase and the DAD / OMMT composite particle suspension obtained in step (4) as the water phase according to a volume ratio of 2:8, then perform high-speed shearing at 12000 r / min at room temperature for 3 min through an emulsifying machine to obtain a Pickering emulsion of DAD / OMMT composite particles loaded with ibuprofen and synergistically stabilized.

[0096] Example 3

[0097] A S N 2. A method for constructing a Pickering emulsion of a drug-loaded alginate derivative / organic montmorillonite synergistically stabilized:

[0098] Steps (1)-(2) are the same as in Example 1.

[0099] (3) Dry sodium-based montmorillonite with a CEC of 1.20 mmol / g in an oven at 110°C to a constant weight, weigh 2 g of the above sodium-based montmorillonite into a 50 mL ball mill jar; mix 1.68 g of dodecyl glucoside (2.0 times the CEC of sodium-based montmorillonite) with 10 mL of deionized water to obtain a dodecyl glucoside aqueous solution, then add the dodecyl glucoside aqueous solution dropwise into the ball mill jar, use a PM type planetary ball mill as the ball milling equipment and agate balls as the ball milling medium, add 2 agate balls with a diameter of 20 mm, 8 agate balls with a diameter of 10 mm and 12 agate balls with a diameter of 6 mm into the ball mill jar, the main disc rotates at a speed of 500 r / min, and ball milling is performed for 1 h; after ball milling, the reaction solution is transferred into a beaker, the ball mill jar and agate balls are rinsed with deionized water, the rinsing solution is collected into the beaker, then centrifugal separation is performed at 7000 r / min, the beaker is rinsed with clean water and centrifugal separation is repeated three times, the precipitate obtained by centrifugal separation is freeze-dried at -50°C for 48 h, ground in an agate mortar and sieved to obtain OMMT.

[0100] (4) Mix DAD obtained in step (2) and OMMT obtained in step (3) according to a mass ratio of 1:2 to obtain DAD / OMMT composite particles; weigh 80 mg of the above DAD / OMMT composite particles into 8 mL of deionized water and perform ultrasonic dispersion at 200 W, shake and stand for 12 h or more to obtain a DAD / OMMT composite particle suspension with a mass fraction of 1.0%.

[0101] (5) Take 5.0 mg ibuprofen and add to 2 mL liquid paraffin to prepare a mixed solution of ibuprofen and liquid paraffin with a concentration of 2.5 mg / mL; mix the above mixed solution of ibuprofen and liquid paraffin as the oil phase and the DAD / OMMT composite particle suspension obtained in step (4) as the water phase according to a volume ratio of 2:8, then pass through an emulsifying machine to perform high-speed shearing at 12000 r / min at room temperature for 3 min to obtain a Pickering emulsion of DAD / OMMT composite particles loaded with ibuprofen and synergistically stabilized.

[0102] Example 4

[0103] A S N 2. A method for constructing a drug-loaded Pickering emulsion of a reaction amphiphilic alginate derivative / organic montmorillonite synergistically stabilized:

[0104] Steps (1)-(2) are the same as in Example 2.

[0105] (3) Dry sodium-based montmorillonite with a CEC of 1.20 mmol / g in an oven at 110°C to a constant weight, take 2 g of the above sodium-based montmorillonite and place it in a 50 mL ball mill jar; mix 0.42 g of dodecyl glucoside (0.5 times the CEC of sodium-based montmorillonite) with 10 mL of deionized water to obtain a dodecyl glucoside aqueous solution, then add the dodecyl glucoside aqueous solution dropwise into the ball mill jar, use a PM-type planetary ball mill as the ball milling equipment and agate balls as the ball milling medium, add 2 agate balls with a diameter of 20 mm, 8 agate balls with a diameter of 10 mm and 12 agate balls with a diameter of 6 mm into the ball mill jar, the main disc rotates at a speed of 500 r / min, and ball milling is performed for 1 h; after ball milling, the reaction solution is transferred into a beaker, the ball mill jar and agate balls are rinsed with deionized water, the rinsing solution is collected into the beaker, then centrifugal separation is performed at 7000 r / min, the beaker is rinsed with clean water and the centrifugal separation is repeated three times, the precipitate obtained by centrifugal separation is freeze-dried at -50°C for 48 h, ground in an agate mortar and sieved to obtain OMMT.

[0106] (4) Mix DAD obtained in step (2) and OMMT obtained in step (3) according to a mass ratio of 1:10 to obtain DAD / OMMT composite particles; take 40 mg of the above DAD / OMMT composite particles, add to 8 mL of deionized water and perform ultrasonic dispersion at 200 W, shake and stand for 12 h or more to obtain a DAD / OMMT composite particle suspension with a mass fraction of 0.5%.

[0107] (5) 10.0 mg ibuprofen was weighed into 2 mL liquid paraffin to prepare a mixed solution of ibuprofen and liquid paraffin with a concentration of 5.0 mg / mL; the mixed solution of ibuprofen and liquid paraffin above was used as the oil phase, and the DAD / OMMT composite particle suspension obtained in step (4) was used as the water phase, and the two were mixed in a volume ratio of 2:8, and then high-speed shearing was performed at room temperature at 12000 r / min for 3 min by an emulsifying machine to obtain the ibuprofen-loaded DAD / OMMT composite particle synergistically stabilized Pickering emulsion.

[0108] The calculation process of the encapsulation efficiency of the DAD / OMMT composite particle synergistically stabilized drug-loaded Pickering emulsion against anti-inflammatory drugs:

[0109] Construction of the standard curve of the anti-inflammatory drug ibuprofen: 50.0 mg of ibuprofen powder was weighed into a beaker and heated to dissolve in 200 mL of PBS buffer, and then diluted to 250 mL in a volumetric flask. 0.25, 0.50, 1.00, 1.50 and 2.00 mL of ibuprofen standard solution were accurately pipetted into a colorimetric tube and supplemented with PBS buffer to make the solution volume 10 mL, respectively, to prepare ibuprofen solutions with concentrations of 5, 10, 20, 30 and 40 μg / mL, respectively. The absorbance value was measured at 221 nm by ultraviolet-visible spectrophotometer, thereby constructing the standard curve of the anti-inflammatory drug ibuprofen.

[0110] Determination of the encapsulation efficiency (EE): 2.0 mL of the ibuprofen-loaded DAD / OMMT composite synergistically stabilized Pickering emulsion in Examples 1-4 was weighed into 10.0 mL of methanol, and after ultrasonic treatment for 10 min, it was transferred to a 10 mL volumetric flask for constant volume. The supernatant was filtered through a 0.45 μm microporous filter membrane, and the absorbance at 221 nm was measured by ultraviolet-visible spectrophotometer. The concentration of ibuprofen in the DAD / OMMT composite particle synergistically stabilized Pickering emulsion was calculated according to the standard curve equation of ibuprofen above, and all experiments were repeated three times. The encapsulation efficiency (EE) of ibuprofen in the DAD / OMMT composite particle synergistically stabilized Pickering emulsion was calculated by the following formula:

[0111]

[0112] In the formula, m1 is the content of ibuprofen in the Pickering emulsion, and m2 is the content of ibuprofen added.

[0113] Test results: the encapsulation efficiency (EE) of the Pickering emulsion in Example 1 for ibuprofen was 82.43%, the encapsulation efficiency (EE) of the Pickering emulsion in Example 2 for ibuprofen was 80.60%, the encapsulation efficiency (EE) of the Pickering emulsion in Example 3 for ibuprofen was 95.34%, and the encapsulation efficiency (EE) of the Pickering emulsion in Example 4 for ibuprofen was 77.53%.

[0114] Example 5

[0115] The difference from Example 1 is only that in step (4), a DAD / OMMT composite particle suspension with a mass fraction of 0.5% is obtained.

[0116] Example 6

[0117] The difference from Example 1 is only that in step (4), a DAD / OMMT composite particle suspension with a mass fraction of 1.5% is obtained.

[0118] Comparative Example 1

[0119] The difference from Example 1 is only that in step (4), a DAD / OMMT composite particle suspension with a mass fraction of 0% is obtained.

[0120] Figure 1 is a synthetic route diagram of DAD in the present application. By Figure 1 it can be known that, in the present application, alginic acid is first neutralized with TBAOH, then cholesteryl chloride, DMF containing TBAF and an aqueous NaCl solution are added, and DAD is prepared through S N 2 reaction and displacement reaction.

[0121] Figure 2 is a FT-IR spectrum (a) and 1 H NMR spectrum (b) of sodium alginate (SA) and DAD in Examples 1-6 of the present application. By Figure 2 it can be seen that the characteristic absorption peaks of SA appear at 3437, 2927 and 1621, 1418 and 1031 cm -1 , which are respectively the stretching vibration absorption peaks of -OH, C-H, -COO- (asymmetric), -COO- (symmetric) and C-O. The absorption peak of DAD at 2927 cm -1 is obviously enhanced, which is attributed to the stretching vibration absorption peak of -CH3 on decyl group. And a new stretching vibration absorption peak appears at 1740 cm -1 , which is attributed to the stretching vibration peak of ester group C=O produced in the reaction process. The result shows that cholesteryl chloride is successfully grafted onto the main chain of alginic acid through S N 2 reaction, and an amphiphilic DAD is synthesized. The FT-IR spectrum and1 H NMR spectrum as shown Figure 2 As shown in b, the signal peaks with chemical shifts in the 3.5-5.0 ppm range are proton signal peaks of the methine group on the SA backbone. Compared with SA, DAD basically retains the proton peaks of the SA backbone and some new signal peaks appear. The new proton signal peak at δ 1.18 ppm is attributed to the proton peak of -CH2- on the cholesterol group of DAD, and the signal peak at δ 0.75 ppm is the proton signal peak of -CH3 on the cholesterol group. The above results are based on S N The reaction successfully modified SA and synthesized DAD.

[0122] Figure 3 SEM images of sodium montmorillonite (a) and OMMT (b) from Examples 1-6. Figure 3 It can be seen that the raw material sodium montmorillonite is in the form of tiny particles, while after mechanical ball milling and physical modification, OMMT is in the form of fine and exfoliated blocks and agglomerated fine blocks, which increases the specific surface area and interlayer spacing of montmorillonite, which is conducive to the embedding of nonionic modifiers into the layers to achieve organic modification.

[0123] Figure 4 TEM images of sodium montmorillonite (a) and OMMT (b) from Examples 1-6. Figure 4 It can be seen that, due to the embedding of the modifier between the layers of OMMT, the interlayer spacing increased from 1.20 nm to 1.56 nm, and the width increased by 0.36 nm. This further verifies the successful modification of sodium montmorillonite by alkyl glycoside molecules through high-speed wet ball milling.

[0124] Figure 5 The images show the FT-IR (a), XRD (b), and TGA (c) spectra of the DAD, OMMT, and DAD / OMMT composite particles in Examples 1-6, as well as the particle size (d), Zeta potential (e), and surface tension (f) of the DAD / OMMT composite particle suspensions in Comparative Example 1, Example 1, and Examples 5-6. Figure 5 As can be seen from the infrared absorption spectrum of the DAD / OMMT composite particles, characteristic absorption peaks of DAD and OMMT can be found. Furthermore, a peak at 3437 cm⁻¹ for DAD was also observed. -1 The -OH absorption peak at [location] shows a slight red shift, indicating that the -COOH of DAD and the -OH of OMMT may interact to form hydrogen bonds. The XRD pattern of the DAD / OMMT composite particles is shown below. Figure 5b shows that similar characteristic diffraction peaks of DAD and OMMT can be found in DAD / OMMT composite particles, and the diffraction peaks at 5.32° in OMMT and 23° in DAD are shifted to 5.73° and 26° respectively, while the hydration crystallization diffraction peak at 13.2° disappears. It is indicated that there should be hydrogen bonding between DAD and OMMT, and DAD is adsorbed on the surface of the OMMT sheet. Figure 5 c is the thermogravimetric analysis diagram of DAD, OMMT and DAD / OMMT composite particles in examples 1-6. Since the DAD / OMMT composite particles contain two substances, its thermal stability is lower than that of OMMT and higher than that of DAD, which indicates the successful preparation of DAD / OMMT composite particles. Figure 5 d and 5e show that with the addition of negatively charged OMMT, the Zeta potential and particle size of DAD / OMMT composite particle suspension become smaller. And with the increase of OMMT concentration, the DAD / OMMT composite particle suspension shows a gradually decreasing trend. This shows that the formation of DAD / OMMT composite particles is mainly due to hydrogen bonding and electrostatic force. It is known that electrostatic repulsion can prevent the aggregation of particles and help the dispersion of particles in liquid. With the increase of the concentration of negatively charged OMMT, more DAD is adsorbed on the surface of OMMT, rather than self-assembly to form colloids, so that the electrostatic repulsion between composite particles will increase. The increase of electrostatic repulsion can better prevent the aggregation of composite particles. Therefore, the particle size of composite particles becomes smaller and smaller, the absolute value of Zeta potential increases more and more, and the stability of dispersion system is better. From Figure 5 f shows that when the suspension system does not add OMMT, the surface tension of 1.0 mg / mL DAD solution is 71.4 mN / m, which is slightly lower than that of water (72.75 mN / m), indicating that the surface of hydrophobically modified DAD still has strong hydrophilicity. With the increase of OMMT concentration, the surface tension of the suspension system decreases. It is indicated that with the increase of OMMT in the system, more and more DAD is adsorbed on OMMT through hydrogen bonding and electrostatic force, so that more and more composite particles are formed in the system, thereby reducing the surface tension of the suspension system and increasing the stability of DAD / OMMT composite particles.

[0125] The Pickering emulsion stabilized by the ibuprofen-loaded DAD / OMMT composite particles in comparative example 1, example 1 and examples 5-6 was subjected to high-speed shearing to make stable emulsion.

[0126] Figure 6 The creaming index of the stable emulsion of the Pickering emulsion stabilized by the ibuprofen-loaded DAD / OMMT composite particles in comparative example 1, example 1 and examples 5-6. From Figure 6It can be seen that the Pickering emulsion stabilized by DAD / OMMT composite particles in Comparative Example 1 was obviously destabilized within 24 hours after shearing and standing, and had no ability to stabilize the emulsion after 12 hours. When the concentration of OMMT in the Pickering emulsion stabilized by DAD / OMMT composite particles was 1.5 wt%, the emulsion had the smallest creaming index of 30.4%. With the increase of OMMT in the emulsion, the stability of the emulsion showed a better and better phenomenon. This can be explained as follows: when DAD alone forms a micellar region to stabilize the emulsion, it is easy to agglomerate, and the particle size in the aqueous solution is large, so the larger particles are not easy to stabilize the emulsion. When OMMT appears in the emulsion system, DAD and OMMT form composite particles with small particle size and good dispersion stability through hydrogen bonds and electrostatic repulsion. With the increase of the concentration of OMMT, the composite particles increase, and more DAD / OMMT composite particles are adsorbed on the oil-water interface to form an interfacial film, which prevents the aggregation and destabilization of the droplets, so the stability of the emulsion is improved.

[0127] Figure 7 The actual photos of the stable emulsion of the Pickering emulsion stabilized by DAD / OMMT composite particles loaded with ibuprofen in Comparative Example 1, Example 1 and Examples 5-6 standing for different times, wherein (A) is standing for 3 hours, (B) is standing for 1 day, (C) is standing for 30 days, and (D) is standing for 30 days in an inverted state. It can be seen from the photos that the Pickering emulsion stabilized by DAD / OMMT composite particles has a certain long-term storage stability. Figure 7 It can be seen that the Pickering emulsion stabilized by DAD / OMMT composite particles has a certain long-term storage stability. After standing for 1 month, the emulsion with 1.5 wt% of OMMT has the best stability.

[0128] Figure 8 The graph of the instability coefficient of the stable emulsion of the Pickering emulsion stabilized by DAD / OMMT composite particles loaded with ibuprofen in Comparative Example 1, Example 1 and Examples 5-6 versus time. It can be seen from the graph that the Pickering emulsion stabilized by DAD / OMMT composite particles has a certain long-term storage stability. Figure 8 It can be seen that the Pickering emulsion stabilized by DAD / OMMT-1.5 wt% has the smallest instability index of 0.43. With the increase of time, the instability coefficients of all samples increase, and the slope of the curve shows a gradually decreasing trend. It shows that the emulsion becomes stable slowly with the passage of time. It shows that the increase of OMMT can increase the stability of the emulsion. This analysis result is the same as the above analysis result of standing stability.

[0129] Figure 9The microscope photos, real photos and size distribution diagrams of the Pickering emulsion stabilized by the ibuprofen-loaded DAD / OMMT composite particles in Comparative Example 1, Example 1 and Examples 5-6 after being placed for 3h, wherein (a) is Comparative Example 1 (DAD), (b) is Example 5 (DAD / OMMT-0.5wt%), (c) is Example 1 (DAD / OMMT-1.0wt%), and (d) is Example 6 (DAD / OMMT-1.5wt%). From Figure 9 It can be seen that when only DAD is used for stabilization, the emulsion droplet size is uneven and the distribution is uneven. With the increase of OMMT, the particle size distribution range of the DAD / OMMT-stabilized emulsion droplets gradually narrows, the average diameter of the droplets gradually decreases, and the droplet size is more uniform. The DAD / OMMT-1.5wt% emulsion prepared in Example 6 has the smallest and most uniform particle size, with an average particle size of 33μm. This is because the greater the concentration of OMMT in the system, the more and more DAD / OMMT composite particles formed will be adsorbed at the oil-water interface, reducing the interfacial energy, forming a tight interfacial film or three-dimensional network structure, hindering the movement of the droplets, and improving the stability of the emulsion.

[0130] Figure 10 The real photos and laser confocal electron microscope photos of the Pickering emulsion stabilized by the ibuprofen-loaded DAD / OMMT composite particles in Example 1 and Examples 5-6 after being dyed with Nile Red and Rhodamine B, respectively, wherein the amount of Nile Red is 5mg / mL and the amount of Rhodamine B is 1mg / mL. From Figure 10 It can be seen that the oil phase is dyed with Nile Red and the water phase is dyed with Rhodamine B. Under the laser confocal electron microscope, the morphology and structure of the micro-particles of the Pickering emulsion can be observed.

[0131] The biocompatibility of the Pickering emulsion stabilized by the DAD / OMMT composite particles was evaluated by in vitro cultured RAW 264.7 cells. The cell culture solution was composed of 90% DMEM, 10% fetal bovine serum, 100U / mL penicillin and 100μg / mL streptomycin. After the RAW 264.7 cells were recovered and adherently proliferated on the surface dish, the 3-4 generation RAW 264.7 cells were cultured at 1×10 5The cells were seeded in 96-well plates at a density of 100 μL per well. Then 100 μL of the DAD / OMMT composite particle-stabilized Pickering emulsion prepared in Example 1 was added to each well, and the final concentrations of the emulsion were 100, 200, 300, 400 and 500 μg / mL, respectively. The cells without sample were used as a control group, and each group was cultured in triplicate. The plates were incubated in a 37°C incubator with 5% CO2. After 2 days of incubation, the cell viability was determined by the CCK-8 method. 10 μL of CCK-8 reagent was added to each well, and the plate was incubated at 37°C for 4 h. Then 100 μL of the solution was taken from each well and added to a new 96-well plate. After 5 min of gentle shaking, the absorbance (OD) value at 450 nm was determined by an enzyme-labeled instrument (X-mark, Bio-rad, USA), and then the MC3T3-E1 cell survival rate was calculated as follows: cell survival rate = [(As-Ab) / (Ac-Ab)]x100%, where As is the absorbance of the experimental well, Ac is the absorbance of the control well, and Ab is the absorbance of the blank well. The results are shown in Table 1. Figure 11 .

[0132] Figure 11 The in vitro cytotoxicity of the DAD / OMMT composite particle-stabilized Pickering emulsion loaded with ibuprofen in Example 1 after 48 h of RAW 264.7 cell culture. The RAW 264.7 cells were seeded in 96-well plates at a density of 100 μL per well. Then 100 μL of the DAD / OMMT composite particle-stabilized Pickering emulsion loaded with ibuprofen in Example 1 was added to each well, and the final concentrations of the emulsion were 100, 200, 300, 400 and 500 μg / mL, respectively. The cells without sample were used as a control group, and each group was cultured in triplicate. The plates were incubated in a 37°C incubator with 5% CO2. After 48 h of incubation, the cell viability was determined by the CCK-8 method. 10 μL of CCK-8 reagent was added to each well, and the plate was incubated at 37°C for 4 h. Then 100 μL of the solution was taken from each well and added to a new 96-well plate. After 5 min of gentle shaking, the absorbance (OD) value at 450 nm was determined by an enzyme-labeled instrument (X-mark, Bio-rad, USA), and then the RAW 264.7 cell survival rate was calculated as follows: cell survival rate = [(As-Ab) / (Ac-Ab)]x100%, where As is the absorbance of the experimental well, Ac is the absorbance of the control well, and Ab is the absorbance of the blank well. The results are shown in Table 2. Figure 10 It can be seen that when the emulsion concentration is in the range of 0-500 μg / mL, the DAD / OMMT composite particle-stabilized Pickering emulsion loaded with ibuprofen still maintains a cell survival rate of more than 86% after 48 h of RAW 264.7 cell culture, indicating that the DAD / OMMT composite particle-stabilized Pickering emulsion loaded with ibuprofen does not have obvious cytotoxicity to RAW 264.7 cells, and exhibits good cell compatibility.

[0133] The emulsion droplets were solidified by using a solution of 2.0 mol% AIBN in styrene instead of the mixed solution of ibuprofen and liquid paraffin in Comparative Example 1, Example 1 and Examples 5-6 as the oil phase, and the microstructure of the DAD / OMMT composite particles stabilized at the oil-water interface was observed by SEM, Figure 12 SEM images of the DAD / OMMT composite particle-stabilized Pickering emulsion loaded with ibuprofen in Comparative Example 1, Example 1 and Examples 5-6 (using 2.0 mol% AIBN in styrene as the oil phase). From the SEM images, it can be seen that the DAD / OMMT composite particles are evenly distributed at the oil-water interface, and the emulsion droplets are spherical and uniform in size. Figure 12It can be seen that only by DAD to stabilize the emulsion, the naked, smooth surface of the droplets. With the increase of OMMT concentration, the droplet surface gradually covered by DAD / OMMT composite particles, forming different roughness. When the DAD / OMMT-0.5wt% composite particles to stabilize the emulsion, the composite particles will be densely covered on the droplet surface, forming a dense, thick particle film, preventing the mutual collision between the droplets, with the increase of OMMT concentration, even unabsorbed composite particles will be accumulated in the continuous phase, gradually forming a three-dimensional network structure, so that the emulsion has good stability.

[0134] Figure 13 The rheological properties of the Pickering emulsion stabilized by the ibuprofen-loaded DAD / OMMT composite particles in Comparative Example 1, Example 1 and Examples 5-6 are shown in the figures, wherein (a) is the viscosity, measured by DHR-2 rotary rheometer, (b) is the angular frequency (strain is 1%, oil-water ratio is 2:8). It can be seen from Figure 13 aIt can be seen that all the emulsions show non-Newtonian fluid behavior, showing the shear thinning behavior of the fluid, and the viscosity value is low. In addition, with the increase of the concentration of OMMT, the viscosity of the emulsion also increases. This is because the greater the concentration of DAD / OMMT, the more likely to form a stable three-dimensional network structure, which increases the viscosity of the emulsion. In addition, the relationship between the storage modulus (G') and the loss modulus (G") of the emulsion was also investigated. When G' < G", the emulsion shows viscous or liquid behavior; when G' > G", the emulsion shows elastic or gel behavior. As shown in Figure 13 b, in all emulsion tests, G' is almost higher than G", and they are almost parallel. This again proves that the more DAD / OMMT composite particles, the more likely to form a dense three-dimensional network structure between the oil droplets, so that the Pickering emulsion shows weak gel structure, which is consistent with the visual emulsion stability results.

[0135] The drug release experiment was carried out at 37℃ to study the drug release behavior of the DAD / OMMT complex particle-stabilized drug-loaded Pickering emulsion. The PBS (0.1M, pH 7.4) was selected as the release medium. The specific operation steps were as follows: 4 mL of the DAD / OMMT complex particle-stabilized drug-loaded Pickering emulsion prepared in Example 5 and Example 6 was placed in a dialysis bag with a molecular weight cut-off of 8000, and the dialysis bag was immersed in 50 mL of PBS solution. At 37℃, the dialysis bag was re-immersed in the same volume of fresh PBS solution at the set time to avoid the saturation effect caused by drug release. At each time point, 5 mL of the solution was taken out, and the absorbance was measured at 221 nm by ultraviolet spectrophotometry to calculate the ibuprofen drug content. The drug content in each time period was added to the total amount of drug released at the previous time to obtain the cumulative ibuprofen drug release amount. The ratio of the total release amount to the drug loading amount was the cumulative drug release rate. The drug release kinetics curve of the cumulative drug release rate versus time was plotted according to the experimental data, and the results are shown in Figure 14 .

[0136] Figure 14 The drug release curves of the DAD / OMMT complex particle-stabilized Pickering emulsion loaded with ibuprofen in Example 5 and Example 6 are shown in Figure 14 It can be seen that, under the condition of fixing the concentration of DAD at 1.0 mg / mL, the DAD / OMMT-0.5wt% (Example 5) stabilized Pickering emulsion has a faster release rate and a larger drug release amount, reaching 79.5% in 60 h. This is because when the OMMT is less, the DAD / OMMT stabilized Pickering emulsion has poor stability, and the same time, it is more prone to demulsification, thereby rapidly releasing the drug. Therefore, the DAD / OMMT stabilized Pickering emulsion has better encapsulation rate and sustained release performance when the OMMT is 1.5wt%. On the one hand, the OMMT has a larger spacing between the layers, which can well adsorb and accommodate the drug, and the encapsulation rates of DAD / OMMT-0.5wt% and DAD / OMMT-1.5wt% are 77.16% and 85.87%, respectively. On the other hand, the more the concentration of OMMT, the more the DAD / OMMT complex particles are formed, and the DAD / OMMT complex particles can effectively adsorb on the surface of the oil droplets to form an effective interfacial barrier, preventing the diffusion of the drug and the destabilization of the emulsion, thereby improving the stability and sustained release performance of the emulsion. In addition, when the pH of the release system is 2, the DAD in the DAD / OMMT complex particle is easily protonated by the carboxyl group, which makes the DAD molecular chain contract, thereby making the size of the DAD / OMMT complex particle smaller, forming a more stable interfacial film, and effectively delaying the release rate of the drug.

[0137] In addition, the DAD / OMMT composite particle prepared by the present application has good standing stability and centrifugal stability, and good cell compatibility. The DAD / OMMT composite particle is observed by SEM to be adsorbed on the oil-water interface to form a uniform interface film to prevent droplet migration and drug diffusion, thereby providing good stability and viscosity for the emulsion. Moreover, the Pickering emulsion has slow drug release behavior and certain pH-responsive drug release behavior for the hydrophobic anti-inflammatory drug ibuprofen, so that the cumulative release rate thereof is lower in a neutral environment. At the same time, the self-diffusion and swelling degradation of the drug in the DAD / OMMT composite particle-stabilized drug-loaded Pickering emulsion are simultaneously performed, which together control the release rate of the hydrophobic drug. In view of the outstanding physicochemical properties of the DAD / OMMT composite particle-stabilized drug-loaded Pickering emulsion, it is used as a hydrophobic anti-inflammatory drug preparation with good sustained-release properties.

[0138] The above merely describes the preferred specific embodiments of the present application, but the protection scope of the present application is not limited thereto, and any person skilled in the art can easily think of changes or replacements within the technical scope disclosed by the present application, which should be covered within the protection scope of the present application. Therefore, the protection scope of the present application should be subject to the protection scope of the claims.

Claims

1. A method of producing a S N 2. A method for constructing a drug-loaded Pickering emulsion stabilized by alginate derivative / organic montmorillonite synergistically, characterized in that, The method comprises the following steps: (1) adding an aqueous solution of tetrabutylammonium hydroxide to an aqueous solution of alginic acid for neutralization reaction, and then performing filtration, dialysis and freeze-drying to obtain tetrabutylammonium alginate (TBA-Alg); (2) mixing the TBA-Alg obtained in step (1) with a tetrabutylammonium fluoride solution, and adding cholesteryl chloride to perform S N 2 reaction, then adding an aqueous NaCl solution to perform a replacement reaction, and performing precipitation, centrifugation, alcohol washing, re-centrifugation, dialysis, and freeze-drying to obtain a cholesteryl group grafted alginate derivative DAD; the molar ratio of the cholesteryl chloride to the uronic acid monomers in the TBA-Alg is (1:1)-(3:10); (3) placing Na-MMT in a ball mill tank, and then adding an aqueous solution of alkyl polyglycoside for ball milling, and then performing centrifugation, washing, freeze-drying, grinding and sieving to obtain alkyl polyglycoside modified organic MMT (OMMT); the CEC of the Na-MMT is 1.20 mmol / g; the alkyl polyglycoside is one or more of hexyl polyglycoside, octyl polyglycoside, decyl polyglycoside and dodecyl polyglycoside; and the amount of the alkyl polyglycoside used is 0.5-2.5 times the CEC of the Na-MMT; (4) mixing the cholesterol-based grafted alginic acid derivative (DAD) obtained in step (2) and the alkyl polyglycoside modified organic MMT (OMMT) obtained in step (3) to obtain DAD / OMMT composite particles; and adding the DAD / OMMT composite particles into water for ultrasonic dispersion and shaking and standing to obtain a DAD / OMMT composite particle suspension; the mass fraction of the DAD / OMMT composite particles in the DAD / OMMT composite particle suspension is 0.5-1.5%; and the mass ratio of DAD to OMMT in the DAD / OMMT composite particles is (1:2)-(1:5); (5) using a mixed solution of a hydrophobic anti-inflammatory drug and liquid paraffin as an oil phase, using the DAD / OMMT composite particle suspension obtained in step (4) as an aqueous phase, mixing the two phases, and then performing shear emulsification to obtain a DAD / OMMT composite particle synergistically stabilized drug-loaded Pickering; the volume ratio of the mixed solution of the hydrophobic anti-inflammatory drug and liquid paraffin to the DAD / OMMT composite particle suspension is (1:9)-(4:6).

2. The S N 2. A method for constructing a drug-loaded Pickering emulsion synergistically stabilized by a reaction amphiphilic alginate derivative / organic montmorillonite, characterized in that, In step (1), the amount of the aqueous solution of tetrabutylammonium hydroxide used is adjusted to the pH value of the aqueous solution of alginic acid to 7-10.

3. The S N 2. A method for constructing a drug-loaded Pickering emulsion synergistically stabilized by a reaction amphiphilic alginate derivative / organic montmorillonite, characterized in that, In step (5), the amount ratio of the hydrophobic anti-inflammatory drug to liquid paraffin in the mixed solution of the hydrophobic anti-inflammatory drug and liquid paraffin is 1.0-5.0 mg / mL.

4. The S N 2. A method for constructing a drug-loaded Pickering emulsion synergistically stabilized by a reaction amphiphilic alginate derivative / organic montmorillonite, characterized in that, In step (5), the hydrophobic anti-inflammatory drug includes one or more of ibuprofen, ofloxacin and ciprofloxacin.

5. The construction method of any one of claims 1-4, wherein the construction method is used to construct an S N 2 Drug-loaded Pickering emulsion stabilized by the synergistic effect of the reaction amphiphilic alginate derivative and organic montmorillonite.

Images