A diatom-based nasal drug delivery sustained-release carrier, a preparation method and application thereof

Abstract

The application provides a diatom-based nasal drug delivery sustained-release carrier and a preparation method and application thereof, and belongs to the technical field of biological processing forming and micro-nano drug carrier manufacturing. The preparation method comprises the following steps: diatomite particle cleaning and purification, loading of anti-rhinitis drugs on the purified diatomite particles, coating of polydopamine on the surfaces of the drug-loaded diatomite particles, and grafting of carboxymethyl chitosan on the surfaces of the polydopamine-coated drug-loaded diatomite particles. The method provided by the application can realize controllable and batch preparation of the diatom-based nasal drug delivery sustained-release carrier, and can realize controllable release of drugs in a stimulus-responsive manner under simulated pathological conditions of the nasal cavity. The preparation method provided by the application is simple and convenient, and the conditions are mild, so that batch preparation of the diatom-based nasal drug delivery sustained-release carrier can be realized. The prepared nasal drug delivery sustained-release carrier has excellent bioadhesion and drug sustained and controlled release performance, and is suitable for intelligent drug delivery and other related fields.

Description

TECHNICAL FIELD

[0001] The present application relates to the technical field of biological processing forming and micro-nano drug carrier manufacturing, and particularly relates to a diatom-based nasal drug delivery sustained-release carrier and a preparation method and application thereof. BACKGROUND

[0002] With the development of micro-nano technology, new intelligent drug delivery carriers have been widely developed, which can improve the performance and efficacy of existing drug molecules and improve the quality of life of patients. Among various drug delivery routes, nasal drug delivery has the advantages of avoiding liver first-pass effect, rapid drug absorption, being suitable for patient first aid and self-help, etc., and is the best way to treat allergic rhinitis and related diseases. However, due to the inherent cilia clearance mechanism of the nasal cavity, the effective residence time of the drug in the nasal cavity is short (15-30 min), which reduces the therapeutic effect of the drug. Therefore, in order to improve the bioavailability of drug molecules through the nasal route and prolong the mucosal residence time, it is urgent to develop drug carriers with adhesion and controlled release properties to achieve safe and efficient adhesion of drugs in the nasal mucosa and achieve the therapeutic effect of sustained release of nasal drug delivery.

[0003] At present, many new drug carriers are used for nasal drug delivery, among which mesoporous SiO2 particles have attracted widespread attention due to their excellent physicochemical properties and have been proven to be useful for vaccine delivery through the nasal route, but their manufacturing cost is high and they are prone to cause pollution. Diatomite is a natural SiO2 porous material with good drug loading stability, environmental friendliness and renewability, and has been proven to be useful for drug loading and release. In addition, diatomite has a complex micro-nano multi-level structure and a large specific surface area, which can prolong drug release, especially for the delivery of hydrophobic drug molecules, but the sustained-release body prepared from diatomite has the problems of poor bioadhesion and controlled release performance. SUMMARY

[0004] Therefore, the present application aims to provide a diatom-based nasal drug delivery sustained-release carrier and a preparation method and application thereof. The nasal drug delivery sustained-release carrier provided by the present application can enhance bioadhesion and stimulate responsive drug release.

[0005] In order to achieve the above-mentioned application purposes, the present application provides the following technical solutions:

[0006] The present application provides a diatom-based nasal drug delivery sustained-release carrier, which takes drug-loaded porous diatomite microparticles as the core, and the surface of the porous diatomite microparticles is sequentially coated with a polydopamine coating and a carboxymethyl chitosan adhesion coating from the inside to the outside.

[0007] Preferably, the porous diatomite microparticles are petri dish-shaped, the diameter of the porous diatomite microparticles is 40-80 μm, and the thickness is 2-4 μm.

[0008] Preferably, the mass ratio of the porous diatomite microparticle, the polydopamine coating and the carboxymethyl chitosan adhesive coating is 1:0.5-2:1.

[0009] Preferably, the mass ratio of the porous diatomite microparticle, the polydopamine coating and the carboxymethyl chitosan adhesive coating is 2:1:2, 1:1:1, 2:3:2 or 1:2:1.

[0010] Preferably, the drug-loaded medicine comprises a rhinitis treatment medicine.

[0011] The application also provides a preparation method of the diatom-based nasal administration sustained-release carrier.

[0012] The diatomite microparticles are cleaned and purified to obtain porous diatomite microparticles.

[0013] The porous diatomite microparticles are loaded with a drug to obtain drug-loaded diatomite microparticles.

[0014] The drug-loaded diatomite microparticles and a dopamine solution are mixed to obtain drug-loaded diatomite microparticles coated with polydopamine.

[0015] The drug-loaded diatomite microparticles coated with polydopamine and a Tris-HCl buffer solution are mixed to obtain a dispersion.

[0016] Carboxymethyl chitosan and a phosphate buffer solution are mixed to obtain a carboxymethyl chitosan solution.

[0017] The carboxymethyl chitosan solution, 1-ethyl-(3-dimethylaminopropyl) carbodiimide and N-hydroxysuccinimide are mixed to obtain an activation solution.

[0018] The dispersion is added to the activation solution to perform a cross-linking reaction to obtain the diatom-based nasal administration sustained-release carrier.

[0019] Preferably, the average diameter of the drug-loaded diatomite microparticles is 51.8±0.16 μm.

[0020] Preferably, the molecular weight of the carboxymethyl chitosan is 240 kDa.

[0021] Preferably, the mass ratio of the 1-ethyl-(3-dimethylaminopropyl) carbodiimide and N-hydroxysuccinimide is 2:1.

[0022] The application also provides application of the diatom-based nasal administration sustained-release carrier in preparation of a medicine for nasal administration.

[0023] The diatom-based nasal drug delivery sustained-release carrier has the porous diatomite microparticle as a core, and the surface of the porous diatomite microparticle is sequentially coated with a polydopamine coating and a carboxymethyl chitosan adhesion coating from inside to outside.

[0024] Compared with common purified diatomite particles, the diatom-based nasal drug delivery sustained-release carrier has significantly improved bioadhesion and controlled release performance. Under the corresponding nasal microenvironment conditions (35°C, pH = 5.0) of allergic rhinitis, the diatom-based nasal drug delivery sustained-release carrier has good bioadhesion and stimulus-responsive release characteristics, and can realize efficient combination of mucin, a main functional component of mucus, and controlled release of budesonide, a typical anti-rhinitis drug. The diatom-based nasal drug delivery sustained-release carrier has excellent bioadhesion and controlled drug release characteristics, and is suitable for use in the field of intelligent drug delivery.

[0025] The diatom-based nasal drug delivery sustained-release carrier has the porous diatomite microparticle as a core, and the surface of the porous diatomite microparticle is sequentially coated with a polydopamine coating and a carboxymethyl chitosan adhesion coating from inside to outside. The diatom-based nasal drug delivery sustained-release carrier has the porous diatomite microparticle as a core, and the surface of the porous diatomite microparticle is sequentially coated with a polydopamine coating and a carboxymethyl chitosan adhesion coating from inside to outside. BRIEF DESCRIPTION OF DRAWINGS

[0026] Figure 1 FIG. 1 is a flowchart of a preparation method of the diatom-based nasal drug delivery sustained-release carrier according to an embodiment of the present application;

[0027] Figure 2 FIG. 2 is a scanning electron microscope image of the diatom-based nasal drug delivery sustained-release carrier according to an embodiment of the present application;

[0028] Figure 3 FIG. 3 is a graph of drug encapsulation efficiency corresponding to different PDA coating concentrations and a thermogravimetric analysis graph of the diatom-based nasal drug delivery sustained-release carrier obtained under the optimal coating concentration according to an embodiment of the present application;

[0029] Figure 4 FIG. 4 is an N2 adsorption / desorption isotherm and pore size distribution graph of the diatom-based nasal drug delivery sustained-release carrier according to an embodiment of the present application;

[0030] Figure 5 FIG. 5 is a Fourier infrared spectrum of the diatom-based nasal drug delivery sustained-release carrier according to an embodiment of the present application;

[0031] Figure 6 FIG. 6 is a surface Zeta potential graph of the diatom-based nasal drug delivery sustained-release carrier under different physiological pH conditions according to an embodiment of the present application;

[0032] Figure 7Test results of mucin binding experiments of the diatom-based nasal drug delivery sustained-release carrier of the embodiment of the present application under different physiological pH conditions;

[0033] Figure 8 Test results of drug release experiments of the diatom-based nasal drug delivery sustained-release carrier of the embodiment of the present application under different physiological pH conditions. DETAILED DESCRIPTION

[0034] The present application provides a diatom-based nasal drug delivery sustained-release carrier, taking drug-loaded porous diatomite microparticles as the core, and successively coating the surface of the porous diatomite microparticles with a polydopamine coating and an adhesion coating of carboxymethyl chitosan from inside to outside.

[0035] In the present application, the porous diatomite microparticles are preferably petri dish-shaped, the diameter of the porous diatomite microparticles is preferably 40-80 μm, and the thickness is preferably 2-4 μm.

[0036] In the present application, the mass ratio of the porous diatomite microparticles, the polydopamine coating and the adhesion coating of carboxymethyl chitosan is preferably 1:0.5-2:1, more preferably 2:1:2, 1:1:1, 2:3:2 or 1:2:1.

[0037] In the present application, the drug loaded is preferably a nasal inflammation treatment drug, and the nasal inflammation treatment drug is preferably budesonide.

[0038] The present application also provides a preparation method of the diatom-based nasal drug delivery sustained-release carrier described in the above technical solution, comprising the following steps:

[0039] The diatomite microparticles are washed and purified to obtain porous diatomite microparticles;

[0040] The porous diatomite microparticles are loaded with a drug to obtain drug-loaded diatomite microparticles;

[0041] The drug-loaded diatomite microparticles and a dopamine solution are mixed to obtain drug-loaded diatomite microparticles coated with polydopamine;

[0042] The drug-loaded diatomite microparticles coated with polydopamine and a Tris-HCl buffer solution are mixed to obtain a dispersion;

[0043] Carboxymethyl chitosan is mixed with a phosphate buffer solution to obtain a carboxymethyl chitosan solution;

[0044] The carboxymethyl chitosan solution, 1-ethyl-(3-dimethylaminopropyl) carbonyl diimide and N-hydroxysuccinimide are mixed to obtain an activation solution;

[0045] The dispersion is added to the activation solution to perform a cross-linking reaction to obtain the diatom-based nasal drug delivery sustained-release carrier.

[0046] In the present application, the raw materials used are all commercially available products in the art, unless otherwise specified.

[0047] The present application cleans and purifies diatomite microparticles to obtain porous diatomite microparticles.

[0048] In the present application, the cleaning agent used for cleaning is preferably anhydrous ethanol.

[0049] In the present application, the ratio of the amount of diatomite microparticles to anhydrous ethanol is preferably 8 g:100 mL.

[0050] In the present application, the cleaning is preferably ultrasonic cleaning, and the time of ultrasonic cleaning is preferably 30 min.

[0051] In the present application, the purification further removes impurities, and the purification preferably comprises the following steps: under the condition of deionized water flushing, 8 g of diatomite microparticles are filtered through a 200-mesh (Φ74 μm) stainless steel screen to remove larger impurities and collect the filtrate; the filtrate is then treated with a 400-mesh (Φ38 μm) stainless steel screen to remove smaller impurities and fragments, and deionized water is used to collect the microparticles left on the screen to obtain a crude treated diatomite solution; the crude treated diatomite solution is poured into a 50 mL graduated cylinder and diluted with deionized water to 50 mL, and after observing that particles have settled to the bottom (1-2 min), 45 mL of the upper mixture is quickly removed to another empty 50 mL graduated cylinder; after stirring with a glass rod and standing for 15 min, 20 mL of the solution above the scale is removed using a pipette, and then the graduated cylinder is filled with deionized water, and this process is repeated three times; the solution in the graduated cylinder is collected and completely dried in a vacuum drying oven at 60°C to obtain the porous diatomite microparticles.

[0052] After obtaining the porous diatomite microparticles, the present application loads the porous diatomite microparticles with drugs to obtain drug-loaded diatomite microparticles.

[0053] In the present application, the average diameter of the drug-loaded diatomite microparticles is preferably 51.8±0.16 μm.

[0054] In the present application, the drug preferably includes a nasal inflammation treatment drug, and the nasal inflammation treatment drug preferably includes Bud.

[0055] In the present application, the mass ratio of the porous diatomite microparticles to the drug is preferably 2:1.

[0056] The present application preferably configures a Bud-absolute ethanol solution with a concentration of 2.5 mg / mL; 20 mg of the porous diatomite microparticles is weighed, added into 4 mL of the Bud-absolute ethanol solution, and mixed by ultrasonic mixing for 2 min, and then mixed at room temperature for 24 h using a vortex mixer; the bottom precipitate is collected by centrifugation (12000 rpm, 5 min) and the supernatant is removed; the precipitate is repeatedly washed with deionized water, collected, and freeze-dried to obtain the drug-loaded diatomite microparticles (DB-Bud, DBB).

[0057] After obtaining the drug-loaded diatomite microparticles, the present application mixes the drug-loaded diatomite microparticles and a dopamine solution to obtain polydopamine-coated drug-loaded diatomite microparticles.

[0058] The present application preferably mixes polydopamine hydrochloride and Tris-HCl buffer (pH = 8.5, 10 mM) to obtain dopamine solutions with different concentrations, and the concentration of the dopamine solution is preferably 1.0, 2.0, 3.0, or 4.0 mg / mL.

[0059] The present application preferably adds the drug-loaded diatomite particles into the dopamine solution, applies mechanical stirring (200 rpm) at room temperature for 6 h until the solution is black, and then filters and separates using a stainless steel screen with a pore size of 38 μm, repeatedly washes with deionized water to remove unreacted dopamine, collects, and freeze-dries to obtain the polydopamine-coated drug-loaded diatomite microparticles (DBB@PDA, DBBP).

[0060] After obtaining the polydopamine-coated drug-loaded diatomite microparticles, the present application mixes the polydopamine-coated drug-loaded diatomite microparticles and Tris-HCl buffer to obtain a dispersion.

[0061] In the present application, the use amount ratio of the polydopamine-coated drug-loaded diatomite microparticles to Tris-HCl buffer is preferably 10 mg:1 mL.

[0062] The present application preferably adds the polydopamine-coated drug-loaded diatomite microparticles into Tris-HCl buffer (pH 8.5, 10 mM).

[0063] The present application mixes carboxymethyl chitosan (CMCS) and phosphate buffer to obtain a carboxymethyl chitosan solution.

[0064] In the present application, the molecular weight of the carboxymethyl chitosan is preferably 240 kDa.

[0065] In the present application, the use amount ratio of the carboxymethyl chitosan to phosphate buffer is preferably 10 mg:1 mL.

[0066] The carboxymethyl chitosan is preferably added into a phosphate buffer (pH = 7.4, 0.1M) and subjected to severe mechanical stirring (450 rpm, 30 min) at room temperature until the CMCS is completely dissolved to obtain the carboxymethyl chitosan solution.

[0067] After obtaining the carboxymethyl chitosan solution, the carboxymethyl chitosan solution, 1-ethyl-(3-dimethylaminopropyl) carbodiimide and N-hydroxysuccinimide are mixed to obtain an activated solution.

[0068] In the present application, the mass ratio of the 1-ethyl-(3-dimethylaminopropyl) carbodiimide and N-hydroxysuccinimide is preferably 2:1, and the 1-ethyl-(3-dimethylaminopropyl) carbodiimide (EDC) and N-hydroxysuccinimide (NHS) act as carboxyl activators.

[0069] In the present application, the mass ratio of the carboxymethyl chitosan and 1-ethyl-(3-dimethylaminopropyl) carbodiimide is preferably 1:1.

[0070] In the present application, the mixing is preferably stirring, and the stirring time is preferably 20 min.

[0071] After obtaining the dispersion and the activated solution, the dispersion is added to the activated solution for cross-linking reaction to obtain the diatom-based nasal drug delivery sustained-release carrier.

[0072] In the present application, the cross-linking reaction is preferably carried out at room temperature, and the time is preferably 6 h.

[0073] In the present application, the cross-linking reaction is preferably carried out under stirring, and during the cross-linking reaction, the carboxyl groups of the carboxymethyl chitosan and the amino groups of the polydopamine are fully cross-linked to form amide bonds, and the carboxymethyl chitosan is grafted to the surface of the diatomite particles.

[0074] After the cross-linking reaction is completed, the present application preferably uses a stainless steel screen with a pore size of 38 μm for filtration separation, and deionized water is used for washing multiple times to remove excess EDC, NHS and unreacted substances, and the diatom-based nasal drug delivery sustained-release carrier is collected and freeze-dried.

[0075] The present application also provides the use of the diatom-based nasal drug delivery sustained-release carrier in the preparation of a nasal drug delivery drug.

[0076] The present application does not have special limitations on the specific method of the use, and the method well known to those skilled in the art can be used.

[0077] The technical solutions in the present application will be clearly and completely described below with reference to the embodiments in the present application. Apparently, 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 creative effort belong to the protection scope of the present application.

[0078] The units in the present application are explained as follows:

[0079] min min ℃ Celsius g gram μm micrometer mL milliliter mg milligram

[0080] mg / L milligram per liter M mole per liter mM millimole per liter rpm revolutions per minute

[0081] The English abbreviations in the present application are explained as follows:

[0082] purified diatomite DB drug-loaded diatomite DB DBB PDA-coated drug-loaded diatomite microparticle DBB PCM CS / PDA-coated drug-loaded diatomite microparticle DBB PC

[0083] Embodiment 1

[0084] Figure 1 The flowchart of the preparation method of the diatom-based nasal drug delivery sustained-release carrier of the embodiments of the present application is shown. The preparation method of the diatom-based nasal drug delivery sustained-release carrier comprises the following steps:

[0085] Step 101: cleaning and purification of diatomite microparticles.

[0086] Cleaning: 8 g of diatomite sample is weighed and dispersed into 100 mL of anhydrous ethanol to obtain a mixed solution A. After stirring with a glass rod, the solution is cleaned by ultrasonic cleaning for 30 min. Purification treatment: to further remove impurities, the solution is filtered through a 200-mesh (Φ74 μm) stainless steel screen under the condition of deionized water washing to remove impurities with larger particle size and collect the filtrate B. Then, the filtrate B is treated with a 400-mesh (Φ38 μm) stainless steel screen to remove impurities and fragments with smaller particle size, and the microparticles left on the screen are collected using deionized water to obtain a coarsely treated diatomite solution C. The solution C is poured into a 50-mL measuring cylinder and diluted to 50 mL with deionized water. After observing that the particles have settled to the bottom (the time is 2 min), the upper mixed solution 45 mL is quickly removed to another empty 50-mL measuring cylinder. Then, the solution is stirred uniformly with a glass rod and left to stand for 15 min. The solution above the 20-mL scale is removed using a pipette, and then the measuring cylinder is filled with deionized water. This process is repeated three times. The solution in the measuring cylinder is collected and completely dried in a vacuum drying oven at 60 ℃ to obtain clean and petri dish-shaped diatomite particles with a particle size of 40-80 μm.

[0087] Step 102: loading of purified diatomite microparticles with nasal drugs.

[0088] Bud-anhydrous ethanol solution with concentration of 2.5mg / mL was prepared using hydrophobic glucocorticoid budesonide (Bud) as model drug for the treatment of rhinitis; 20mg of purified dish-shaped diatomite microparticles were weighed and added into 4mL of prepared drug solution and mixed by ultrasonic for 2min, then mixed by vortex mixer for 24h at room temperature; the bottom precipitate was collected by centrifugation (12000rpm, 5min) and the supernatant was removed; the precipitate was repeatedly washed with deionized water, collected and freeze-dried to obtain drug-loaded dish-shaped diatomite microparticles.

[0089] Step 103: Surface of drug-loaded diatomite microparticles was coated with polydopamine.

[0090] Polydopamine hydrochloride was weighed and added into 50mL of Tris-HCl buffer (pH=8.5, 10mM) to obtain dopamine solutions with different concentrations (1.0, 2.0, 3 and 4.0mg / mL); 100mg of drug-loaded diatomite particles were weighed and added into the above series of solutions, and mechanical stirring (200rpm) was applied at room temperature for 6h until the solution turned black; filtration separation was performed using a stainless steel screen with a pore size of 38μm, and the unreacted dopamine was removed by washing with deionized water for several times, and the polydopamine (PDA)-coated drug-loaded diatomite microparticles were collected and freeze-dried, and the mass of drug remaining in the samples prepared with different dopamine coating concentrations was tested by thermogravimetric analysis, and the mass ratio of PDA:DB was 1:2, 1:1, 3:2 and 2:1, respectively.

[0091] Step 104: Carboxymethyl chitosan was grafted onto the surface of polydopamine-coated drug-loaded diatomite microparticles.

[0092] Take 100 mg of carboxymethyl chitosan (CMCS, molecular weight: 240 kDa) and add it to 10 mL of phosphate buffer (pH = 7.4, 0.1 M) and apply vigorous mechanical stirring (450 rpm, 30 min) at room temperature until the CMCS is completely dissolved; take 100 mg of 1-ethyl-(3-dimethylaminopropyl) carbodiimide (EDC) and 50 mg of N-hydroxysuccinimide (NHS) as a carboxyl activator and add it to the above solution, continue stirring at room temperature for 20 min, form an activated solution A; during activation, take 100 mg of polydopamine coated drug-loaded diatomite microparticles (optimal coating mass ratio PDA:DB = 1:1) and add it to 10 mL of Tris-HCl buffer (pH 8.5, 10 mM) to form a mixed solution B; immediately after the activation is completed, add the mixed solution B to the activated solution A, and stir (200 rpm) at room temperature for 6 h to allow the carboxyl groups of the carboxymethyl chitosan and the amino groups of the polydopamine to crosslink to form amide bonds, and graft the carboxymethyl chitosan to the surface of the diatomite particles; filter and separate using a stainless steel screen with a pore size of 38 μm, wash with deionized water several times to remove excess crosslinking agent and unreacted substances, collect and freeze-dry to obtain carboxymethyl chitosan / polydopamine (CMCS / PDA) coated drug-loaded diatomite microparticles with bioadhesion.

[0093] The surface morphology and microstructure of the CMCS / PDA coated drug-loaded diatomite microparticles are characterized by scanning electron microscopy, and the components of the CMCS / PDA coated drug-loaded diatomite microparticles are quantitatively tested by a thermal gravimetric analyzer. The specific surface area and pore volume of the CMCS / PDA coated drug-loaded diatomite microparticles are determined by a porosity analyzer, the chemical groups of the CMCS / PDA coated drug-loaded diatomite microparticles are analyzed by Fourier transform infrared spectroscopy, the surface crystal structure of the CMCS / PDA coated drug-loaded diatomite microparticles is analyzed by an X-ray diffractometer, the bioadhesion of the CMCS / PDA coated drug-loaded diatomite microparticles is tested by a particle size analyzer, and the drug release capacity of the CMCS / PDA coated drug-loaded diatomite microparticles is tested by a UV-visible spectrophotometer.

[0094] Figure 2 The scanning electron microscope image of the diatom-based nasal drug delivery sustained-release carrier (PDA:DB mass ratio = 1:1) of the embodiment of the present application. The nasal drug delivery sustained-release carrier prepared in the embodiment of the present application is mainly obtained by the dipping method, self-polymerization coating and chemical coupling process, and the surface of the diatomite biological template is first adsorbed with a hydrophobic drug, then deposited with polydopamine (PDA), and then coated with carboxymethyl chitosan (CMCS). As shown in Figure 2As shown, the CMCS / PDA-coated diatom-based drug carrier presents a petri dish-shaped structure, both the concave and convex surfaces of which have nanoscale pore structures and are evenly coated with the CMCS / PDA double coating to form drug loading and encapsulation.

[0095] Figure 3 (a) is a graph showing drug encapsulation efficiency at different PDA coating concentrations according to an embodiment of the present invention. Figure 3 (b) is the thermogravimetric analysis of the diatom-based sustained-release carrier for nasal administration obtained at the optimal coating concentration (2.0 mg / mL). Figure 3 As shown in (a), compared with the original drug-loaded diatomaceous earth, the amount of encapsulated drug was lost after the PDA coating modification process (the optimal coating mass ratio was PDA:DB = 1:1, the drug encapsulation efficiency was 24.39±1.169%, and the unit mass loading was 121.95±5.845μg / mg). This is because budesonide has a certain water solubility, and some drugs adsorbed on the diatomaceous earth surface during the modification process will be dissolved due to the large concentration gradient. Figure 3 (b) is a thermogravimetric analysis diagram of the components of the diatom-based nasal sustained-release carrier obtained at the optimal ratio, and the final drug loading per unit mass is 91.8 μg / mg.

[0096] Figure 4 The N2 adsorption / desorption isotherms and pore size distribution diagram of the diatom-based sustained-release carrier for nasal administration (PDA:DB mass ratio = 1:1) in the embodiment of the present invention are shown. Figure 4 As shown in the figure, the purified diatomaceous earth, PDA-coated drug-loaded diatomaceous earth and CMCS / PDA-coated drug-loaded diatomaceous earth microparticles all showed typical type II isotherm characteristics. After drug loading, PDA coating and CMCS grafting, the specific surface area and pore volume of the diatom-based microparticles decreased successively. The surface drugs were successfully adsorbed onto the surface and pores of the diatomaceous earth microparticles, and the coating of PDA and CMCS further blocked the outlet of the pores to achieve the drug encapsulation effect.

[0097] Figure 5 The following is a Fourier transform infrared spectrum of the diatom-based sustained-release carrier for nasal administration (PDA:B mass ratio = 1:1) according to an embodiment of the present invention. Figure 5 As shown, peaks corresponding to the characteristic groups of diatomaceous earth can be observed in purified diatomite, PDA-coated drug-loaded diatomite, and CMCS / PDA-coated drug-loaded diatomite microparticles. Furthermore, the amino shear vibration peak corresponding to polydopamine can be observed in the PDA-coated drug-loaded diatomite, while the characteristic peak of the amide group generated by the cross-linking reaction can be observed in the CMCS / PDA-coated drug-loaded diatomite microparticles, indicating that PDA and CMCS have been successfully coated on the surface of the diatomite microparticles.

[0098] Pig gastric mucin was selected as the main functional component in the mucous membrane mucus, and the bioadhesion of the CMCS / PDA coated drug-loaded diatomite microparticles was tested and analyzed through the mucin binding experiment. The mucin binding experiment of the CMCS / PDA coated drug-loaded diatom-based microparticles includes:

[0099] 100 mg of pig gastric mucin was weighed and added into 0.1 M PBS buffer (pH 5.0, 5.8, 6.5 and 7.4) and subjected to severe mechanical stirring (450 rpm, 2 h) at 37℃ until completely dissolved to obtain a mucin solution with a concentration of 1% w / v; the obtained solution was subjected to ultrasonic treatment for 30 min and centrifuged (4000 rpm, 20 min) to obtain a supernatant; a cellulose acetate filter membrane (pore size 0.8 μm) was used for vacuum suction filtration to obtain a submicron (300-400 nm) mucin-PBS solution for subsequent use. 0.5 mL of the above mucin-PBS solution was taken and placed in a 10 mL centrifuge tube, and the corresponding PBS solution with different pH was used for dilution to 5 mL, and 20 mg of drug-loaded diatomite (experimental group 1), PDA coated drug-loaded diatomite (experimental group 2), CMCS / PDA coated drug-loaded diatomite (experimental group 3) was added to each group of solutions; the mixture was gently mixed by vortex mixer for 2 h and then centrifuged (8000 rpm, 20 min) to obtain the supernatant, and the absorbance at 260 nm was detected by ultraviolet spectrophotometer, and the mucin binding rate of the microparticles was calculated according to the calibration results.

[0100] Figures 6-7 The surface zeta potential diagram and mucin binding experiment test results of the diatom-based nasal administration sustained-release carrier (PDA:DB mass ratio = 1:1) of the present application under different physiological pH conditions. As shown in Figures 6-7 the purified diatomite and the PDA coated drug-loaded diatomite microparticles all showed electronegativity at the tested pH points (pH = 5.0, 6.0 and 7.0), and the mucin binding amount decreased with the decrease of pH; however, for the CMCS / PDA coated drug-loaded diatomite microparticles, the surface zeta potential realized the charge reversal of pH responsiveness from negative to positive during the process of pH from 7.0 to 5.0, which confirmed that it could interact with mucin through electrostatic attraction, hydrogen bonding and hydrophobic interaction mechanisms, etc., and endow the CMCS / PDA coated drug-loaded diatomite microparticles with enhanced bioadhesion, so that it has the ability of nasal efficient adhesion to prolong the in-situ action of the drug.

[0101] The stimulus-responsive drug release characteristics of the CMCS / PDA coated drug-loaded diatomite microparticles under physiological (pH = 7.4, 35℃) and allergic rhinitis corresponding nasal microenvironment (pH = 5.0, 35℃) were explored. The stimulus-responsive release experiment of the CMCS / PDA coated drug-loaded diatom-based microparticles includes:

[0102] 8mg drug-loaded diatomite samples (pure drug-loaded diatomite, PDA-coated drug-loaded diatomite, CMCS / PDA-coated drug-loaded diatomite) were weighed and suspended in 50mL PBS (0.1M, pH=5.0 or 7.4) solution, and incubated in a 35℃ water bath shaker; 1.5mL buffer was sampled every 20min in the first 8h, and every 24h in the following 96h for absorbance measurement, and the same volume of liquid was supplemented; the absorbance of the sampled solution at 247nm was detected by UV-visible spectrophotometer, and the cumulative release rate of the drug was calculated according to the results.

[0103] Figure 8 The drug release test results of the diatom-based nasal drug delivery sustained-release carrier (PDA:DB mass ratio=1:1) of the embodiment of the present application under different physiological pH conditions. Budesonide, a typical anti-rhinitis drug, was selected as a typical drug molecule for nasal drug delivery, and the responsive release capacity of the diatom-based nasal drug delivery sustained-release carrier was tested by in vitro drug release experiment. Under physiological conditions (pH=7.4), the CMCS / PDA coating can effectively prevent drug leakage, and the release rate within 8h is only 22.3%; while in the corresponding microenvironment of allergic rhinitis (pH=5.0), the CMCS / PDA coating layer successively swells and hydrolyzes, so that the drug release rate is significantly accelerated, and the release rate within 8h reaches 51.5%, and the drug is continuously released within 96h.

[0104] In summary: the diatom-based nasal drug delivery sustained-release carrier of the present application can realize the deposition and coating of drug molecules, polydopamine and carboxymethyl chitosan on the surface of diatomite particles by bio-forming method, and can make the drug be loaded and effectively encapsulated in amorphous form, has enhanced bioadhesion, can realize responsive release under nasal pathological conditions, and has application potential in the field of intelligent drug delivery and the like.

[0105] Compared with the prior art, the diatom-based nasal drug delivery sustained-release carrier of the present application has the following beneficial effects:

[0106] (1) The diatom-based nasal drug delivery sustained-release carrier of the present application can load hydrophobic drugs in amorphous form and has stable drug encapsulation effect, has good bioadhesion and stimulus-responsive release characteristics, can realize drug triggering and long-term release under specific pathological conditions, and has the potential to quickly relieve symptoms and prolong the therapeutic effect in the nasal environment.

[0107] (2) The preparation method described in the application uses natural culture dish-shaped diatomite as a template, realizes efficient loading and coating of drug molecules, polydopamine and carboxymethyl chitosan coating through physical adsorption, self-assembly and chemical coupling methods, is stable in structure, simple in process, and can realize controllable and batch production of diatom-based nasal cavity drug delivery sustained-release carriers.

[0108] The above only describes the preferred embodiments of the present application, and does not limit the present application in any form. It should be noted that, for those skilled in the art, without departing from the principles of the present application, a number of improvements and refinements can be made, and these improvements and refinements should also be considered as the protection scope of the present application.

Claims

1. A diatom-based, sustained-release nasal delivery vehicle, characterized in that, The porous diatomite microparticle is coated with a polydopamine coating and a carboxymethyl chitosan adhesion coating from inside to outside. The porous diatomite microparticle is in the shape of a culture dish, and has a diameter of 40-80 μm and a thickness of 2-4 μm. The mass ratio of the porous diatomite microparticle, the polydopamine coating and the carboxymethyl chitosan adhesion coating is 1:0.5-2:

1. The drug loaded is budesonide. The preparation method of the diatom-based nasal drug delivery sustained-release carrier comprises the following steps: The diatomite microparticle is washed and purified to obtain a porous diatomite microparticle. The porous diatomite microparticle is loaded with a drug to obtain a drug-loaded diatomite microparticle. The drug-loaded diatomite microparticle and a dopamine solution are mixed to obtain a drug-loaded diatomite microparticle coated with polydopamine. The drug-loaded diatomite microparticle coated with polydopamine and a Tris-HCl buffer solution are mixed to obtain a dispersion liquid. Carboxymethyl chitosan is mixed with a phosphate buffer solution to obtain a carboxymethyl chitosan solution. The carboxymethyl chitosan solution, 1-ethyl-(3-dimethylaminopropyl) carbodiimide and N-hydroxysuccinimide are mixed to obtain an activation solution. The dispersion liquid is added to the activation solution to perform a cross-linking reaction to obtain the diatom-based nasal drug delivery sustained-release carrier.

2. The diatom-based, intranasal, slow release delivery vehicle of claim 1, wherein, The mass ratio of the porous diatomite microparticle, the polydopamine coating and the carboxymethyl chitosan adhesion coating is 2:1:2, 1:1:1, 2:3:2 or 1:2:

1.

3. The method of claim 1 or 2 for the preparation of a diatom-based sustained release nasal delivery vehicle, characterized in that, The preparation method comprises the following steps: The diatomite microparticle is washed and purified to obtain a porous diatomite microparticle. The porous diatomite microparticle is loaded with a drug to obtain a drug-loaded diatomite microparticle. The drug-loaded diatomite microparticle and a dopamine solution are mixed to obtain a drug-loaded diatomite microparticle coated with polydopamine. The drug-loaded diatomite microparticle coated with polydopamine and a Tris-HCl buffer solution are mixed to obtain a dispersion liquid. Carboxymethyl chitosan is mixed with a phosphate buffer solution to obtain a carboxymethyl chitosan solution. The carboxymethyl chitosan solution, 1-ethyl-(3-dimethylaminopropyl) carbodiimide and N-hydroxysuccinimide are mixed to obtain an activation solution. The dispersion liquid is added to the activation solution to perform a cross-linking reaction to obtain the diatom-based nasal drug delivery sustained-release carrier.

4. The production method according to claim 3, characterized by, The average diameter of the drug-loaded diatomite microparticle is 51.8±0.16 μm.

5. The preparation method according to claim 3, characterized in that The molecular weight of the carboxymethyl chitosan is 240 kDa.

6. The preparation method according to claim 3, characterized in that The mass ratio of the 1-ethyl-(3-dimethylaminopropyl) carbodiimide and N-hydroxysuccinimide is 2:

1.

7. The diatom-based nasal drug delivery sustained-release carrier of claim 1 or 2 for use in the preparation of a drug for treating allergic rhinitis.

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