CN113797177B - Quercetin oral sustained-release preparation modified by ionic emulsifier chitosan nanoparticles and preparation method thereof - Google Patents

Quercetin oral sustained-release preparation modified by ionic emulsifier chitosan nanoparticles and preparation method thereof Download PDF

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CN113797177B
CN113797177B CN202111183969.3A CN202111183969A CN113797177B CN 113797177 B CN113797177 B CN 113797177B CN 202111183969 A CN202111183969 A CN 202111183969A CN 113797177 B CN113797177 B CN 113797177B
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quercetin
preparation
chitosan
oral
release
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CN113797177A (en
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姚京京
崔世华
段聪慧
胡洁
李留成
齐浩然
王欣
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Anhui University of Science and Technology
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    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K9/00—Medicinal preparations characterised by special physical form
    • A61K9/48—Preparations in capsules, e.g. of gelatin, of chocolate
    • A61K9/50—Microcapsules having a gas, liquid or semi-solid filling; Solid microparticles or pellets surrounded by a distinct coating layer, e.g. coated microspheres, coated drug crystals
    • A61K9/51—Nanocapsules; Nanoparticles
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    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
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    • A61K31/335—Heterocyclic compounds having oxygen as the only ring hetero atom, e.g. fungichromin
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    • A61K31/352—Heterocyclic compounds having oxygen as the only ring hetero atom, e.g. fungichromin having six-membered rings with one oxygen as the only ring hetero atom condensed with carbocyclic rings, e.g. methantheline 
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K9/00—Medicinal preparations characterised by special physical form
    • A61K9/48—Preparations in capsules, e.g. of gelatin, of chocolate
    • A61K9/50—Microcapsules having a gas, liquid or semi-solid filling; Solid microparticles or pellets surrounded by a distinct coating layer, e.g. coated microspheres, coated drug crystals
    • A61K9/51—Nanocapsules; Nanoparticles
    • A61K9/5107—Excipients; Inactive ingredients
    • A61K9/5123—Organic compounds, e.g. fats, sugars
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K9/00—Medicinal preparations characterised by special physical form
    • A61K9/48—Preparations in capsules, e.g. of gelatin, of chocolate
    • A61K9/50—Microcapsules having a gas, liquid or semi-solid filling; Solid microparticles or pellets surrounded by a distinct coating layer, e.g. coated microspheres, coated drug crystals
    • A61K9/51—Nanocapsules; Nanoparticles
    • A61K9/5107—Excipients; Inactive ingredients
    • A61K9/513—Organic macromolecular compounds; Dendrimers
    • A61K9/5161—Polysaccharides, e.g. alginate, chitosan, cellulose derivatives; Cyclodextrin
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P29/00—Non-central analgesic, antipyretic or antiinflammatory agents, e.g. antirheumatic agents; Non-steroidal antiinflammatory drugs [NSAID]
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P31/00—Antiinfectives, i.e. antibiotics, antiseptics, chemotherapeutics
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    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B82—NANOTECHNOLOGY
    • B82Y—SPECIFIC USES OR APPLICATIONS OF NANOSTRUCTURES; MEASUREMENT OR ANALYSIS OF NANOSTRUCTURES; MANUFACTURE OR TREATMENT OF NANOSTRUCTURES
    • B82Y30/00—Nanotechnology for materials or surface science, e.g. nanocomposites
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B82—NANOTECHNOLOGY
    • B82Y—SPECIFIC USES OR APPLICATIONS OF NANOSTRUCTURES; MEASUREMENT OR ANALYSIS OF NANOSTRUCTURES; MANUFACTURE OR TREATMENT OF NANOSTRUCTURES
    • B82Y40/00—Manufacture or treatment of nanostructures
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B82—NANOTECHNOLOGY
    • B82Y—SPECIFIC USES OR APPLICATIONS OF NANOSTRUCTURES; MEASUREMENT OR ANALYSIS OF NANOSTRUCTURES; MANUFACTURE OR TREATMENT OF NANOSTRUCTURES
    • B82Y5/00—Nanobiotechnology or nanomedicine, e.g. protein engineering or drug delivery
    • Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
    • Y02—TECHNOLOGIES OR APPLICATIONS FOR MITIGATION OR ADAPTATION AGAINST CLIMATE CHANGE
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    • Y02A50/30—Against vector-borne diseases, e.g. mosquito-borne, fly-borne, tick-borne or waterborne diseases whose impact is exacerbated by climate change

Abstract

The invention relates to an ion emulsifier chitosan nanoparticle modified quercetin oral sustained-release preparation and a preparation method thereof, and specifically comprises the following steps: the invention takes quercetin as a wrapped medicine, and prepares an anionic surfactant lauryl sodium sulfate with oral penetration promoting effect and chitosan with positive charge by an emulsifying ion cross-linking method, the preparation process is simple and convenient, the energy consumption is low, the operation is easy, and the quercetin oral sustained-release preparation with the size less than 200 nanometers and higher entrapment rate is obtained, so as to improve the water solubility of the quercetin, prolong the sustained-release effect and promote the penetration of the preparation to enhance the absorption of the quercetin so as to enhance the oral bioavailability.

Description

Quercetin oral sustained-release preparation modified by ionic emulsifier chitosan nanoparticles and preparation method thereof
Technical Field
The invention relates to a quercetin oral sustained-release preparation modified by an ionic emulsifier chitosan nanoparticle and a preparation method thereof, belonging to the technical field of quercetin nanometer oral sustained-release preparations.
Background
Quercetin is a natural polyhydroxy flavonoid compound, widely exists in melons, fruits, vegetables and various Chinese herbal medicines, has various biological activities and pharmacological actions such as anti-inflammation, anti-allergy, anti-free radical oxidation, anti-cancer, anti-HIV and the like, has the advantages of wide sources, low price, no toxic or side effect, high biocompatibility and the like, and can have great development value in clinical research and application. But the clinical application effect is not ideal due to the defects of low solubility, small internal activity, poor gastrointestinal absorption, low bioavailability and the like, thereby seriously limiting the application of the compound in the field of medicine. Therefore, a strategy for increasing the solubility and oral bioavailability of quercetin is urgently needed at present, so that the effective development and medical application values of quercetin resources are improved.
The nanometer technology is a very effective method for enhancing the solubility of the medicine and improving the curative effect, and compared with the traditional medicine, the nanometer preparation changes the pharmacokinetics, the tissue distribution and the internal pharmacological activity of the medicine by reducing the particle size, increasing the specific surface area, improving the solubility and the like so as to achieve the effects of reducing the adverse reaction and improving the medicine effect and the bioavailability. At present, researchers have conducted extensive research on quercetin nano-formulations, and have attempted to prepare various nano-formulations, such as quercetin nanocrystals, quercetin nanoliposomes, quercetin nanoemulsions, and the like. However, how to ensure the stability of the quercetin nanocrystals is still a difficult problem faced by researchers in the preparation process, and meanwhile, due to the defects of easy deterioration, poor stability, unsatisfactory controlled-release effect and the like of the quercetin nanoliposomes, the application of the quercetin nanocrystals in clinical practical treatment is limited. In addition, the quercetin nanoemulsion technology also has the defects of high pharmaceutical cost, short shelf life, influence on standard detection and the like.
The polymer nanoparticle drug carrying system is used for dissolving or dispersing drugs into nanoparticles, and the system is proved to be capable of improving drug solubility and biological activity by being used as a drug carrier at present, and has the advantages of stable drug carrying system, high drug encapsulation efficiency and the like. Among many drug carriers, polybutylcyanoacrylate and chitosan are attracting attention due to their good biocompatibility and biodegradability, however, polybutylcyanoacrylate has the disadvantage of being expensive. Chitosan, as a natural polymer with biocompatibility and positive charge after ionization in solution, has good permeability and drug absorption enhancing performance, is widely applied to form a carrier structure of a nano-composite, can obviously improve the bioavailability of drugs, but is difficult to dissolve in water and is difficult to form a stable nano-preparation by single use. It has been reported that stable chitosan nanoparticles are prepared by using sodium tripolyphosphate as a cross-linking agent, the preparation cost is increased by using sodium tripolyphosphate, the industrialization is not facilitated, and the application of the prepared chitosan nanoparticles is limited due to poor stability, large particle size, early drug release caused by particle degradation, intestinal malabsorption and the like.
Disclosure of Invention
Aiming at the defects of low stability, burst release and low gastrointestinal absorption rate of the existing chitosan nano technology, the invention provides a quercetin nano oral sustained-release preparation which is prepared by combining chitosan and sodium dodecyl sulfate and has the advantages of small particle size, strong dispersibility, good stability, slow drug release, effective promotion of drug cell permeation and intestinal absorption and the like.
In order to solve the technical problems, the invention adopts the following technical scheme: an oral slow-release quercetin preparation modified by chitosan nanoparticles as ionic emulsifier comprises coated quercetin, coating material comprising chitosan and sodium laurylsulfate.
Further, the particle size of the quercetin oral sustained-release preparation modified by the ionic emulsifier chitosan nanoparticle is 160-180 nanometers, and the inventor finds that the oral absorption efficiency of the quercetin oral sustained-release preparation is better under the particle size.
Furthermore, the encapsulation rate of the quercetin oral sustained-release preparation modified by the ionic emulsifier chitosan nanoparticle is 71.97-75.83%, and in the implementation of the process of the invention, the inventor finds that the oral bioavailability of the quercetin sustained-release preparation is higher under the encapsulation rate, wherein the quercetin is a coated medicament, and chitosan and sodium dodecyl sulfate are coating materials.
The invention provides a preparation method of the ionic emulsifier chitosan nanoparticle modified quercetin oral sustained-release preparation, which comprises the following steps:
(1) Preparation of raw materials
Preparing a cation crosslinking raw material: uniformly dispersing chitosan in a glacial acetic acid aqueous solution;
preparing an anionic emulsifying crosslinking raw material: dispersing an anionic emulsifier or oral penetration enhancer uniformly in ultrapure water;
preparing stable reaction raw materials: uniformly dispersing a stabilizer in ultrapure water;
(2) Crosslinking reaction
The crosslinking reaction adopts an emulsifying ion crosslinking reaction;
the emulsifying ionic crosslinking reaction process comprises the following steps: adding quercetin into cation crosslinking raw material which is stirred overnight and filtered by a filter membrane, carrying out ultrasonic treatment to fully dissolve the quercetin, adjusting pH to be neutral by using sodium hydroxide to obtain a mixture I, and then dropwise and slowly adding anion crosslinking raw material into the mixture I to carry out emulsification ion crosslinking reaction.
Furthermore, the emulsifier, the ionic crosslinking agent and the oral penetration enhancer are all sodium dodecyl sulfate, and in the implementation process of the invention, the inventor finds that the sodium dodecyl sulfate can be used as an anionic surfactant to ensure that the sodium dodecyl sulfate can be subjected to emulsification ionic crosslinking reaction with chitosan to form chitosan nanoparticles with small particle size, and the sodium dodecyl sulfate is an oral preparation excipient approved by FDA, and can also be an effective penetration enhancer and gastrointestinal protease inhibitor in an oral preparation, so that the prepared chitosan nanoparticles can effectively penetrate intestinal epithelial cells, the drug absorption is promoted, the oral bioavailability is improved, and the curative effect of quercetin is further improved.
Furthermore, 0.3 to 0.6 g of chitosan is dispersed in 0.8 to 1.1 percent of glacial acetic acid aqueous solution per 40 ml of cation emulsification crosslinking raw material. In the process of implementing the invention, the inventor finds that the encapsulation rate of the prepared chitosan nanoparticles to quercetin is not high below or above the range, mainly because the amount of the chitosan subjected to the emulsifying ion crosslinking reaction is not enough below the range, and above the range, the crosslinking reaction is incomplete, chitosan raw materials are wasted, and the quercetin which is difficult to dissolve in water is ingeniously dissolved by using the glacial acetic acid in the range, so that the use of toxic organic reagents for dissolving the quercetin is avoided.
Furthermore, 0.3 to 0.6 g of anionic emulsifier is dispersed in each 40 ml of ultrapure water solution of the anionic emulsifying crosslinking raw material. In the process of implementing the invention, the inventor finds that in the range, the prepared chitosan nanoparticles are small in size, uniform in distribution, highest in encapsulation efficiency and highest in oral bioavailability. Mainly because the sodium dodecyl sulfate is used as the emulsifier and the anionic cross-linking agent in the invention, the particle diameter is larger and easy to aggregate and is unstable when the particle diameter is lower than the range, and the penetration promoting effect of the nanoparticles is poor. Above this range, the electromotive force of the formed nanoparticles is lower than 30 mv, making the whole nano-system very unstable and the encapsulation rate of quercetin not high.
Furthermore, stable reaction raw materials are prepared, and 0.8-1.1 g of stabilizing agent is dispersed in each 20 ml of ultrapure water solution, and in the process of implementing the invention, the inventor finds that the prepared chitosan nanoparticles have high stability and slow release effect in the range.
Furthermore, the optimized dosage-volume ratio of the cation and anion emulsification crosslinking raw materials in the emulsification ion crosslinking reaction is 1:1, and the inventor finds that the reaction efficiency is higher, the reaction can be fully carried out and the operation is convenient by adopting the ratio.
Further, the inventor finds that the application of the range can ensure that the encapsulation rate of the obtained quercetin is 71.97-75.83% and the utilization rate of the quercetin is ensured. Filtering the cation crosslinking raw material by using a 0.45 micron filter membrane, and ensuring that a 160-180 nanometer slow release preparation can be obtained. Adjusting the mixture to neutral to ensure that the pH of the prepared quercetin nanometer preparation is close to physiological pH value absorbed by small intestine.
Further, in the process of the emulsification ion crosslinking reaction, magnetic stirring is carried out at the rotating speed of 900-1100 revolutions per minute. In the process of the invention, the inventor finds that the stirring speed can ensure that the prepared nanoparticles have smaller particle size and more uniform distribution.
Further, the reaction time of the emulsifying ionic crosslinking reaction is 1.5 to 2.5 hours. In the process of implementing the invention, the inventor finds that the range can ensure that the anion-cation crosslinking reaction is more complete, and more sufficient quercetin-loaded chitosan nanosuspension is obtained.
Further, the present invention also has the steps of:
(3) Stable reaction
The stable reaction process is as follows: slowly dropwise adding stabilizer dispersed in ultrapure water into the solution prepared by crosslinking reaction, stirring for stabilizing reaction, and obtaining polymer-coated quercetin nanometer sustained-release oral preparation suspension with high stability, or preparing lyophilized powder of quercetin nanometer sustained-release oral preparation by freeze drying technology.
Further, the stabilizer is dextran-70, although this kind of raw materials are the common nanometer preparation stabilizer, it is different in nature that its adding order is different to the improvement stability effect of nanometer preparation in implementing the process of the invention, when it is added finally, chitosan nanoparticle has already formed, it can combine with prepared nanometer through the hydrogen bond to form a layer of protective housing, make the prepared quercetin nanometer preparation more stable, in the course of drug delivery from long stomach to small intestine, guarantee the skeleton of nanoparticle not gather, not stand, until it reaches its penetrating cell of destination and slowly release the drug, exert the therapeutic effect.
Further, in the step (3), a magnetic stirrer with the rotation speed of 900-1100 revolutions per minute is adopted for stirring treatment, and the time is 0.5-1 hour. By implementing the process of the invention, the inventor finds that the quercetin nanometer preparation obtained in the range has better stability slow-release effect and oral bioavailability.
The invention also provides an oral medicament. Comprises a quercetin sustained-release preparation and auxiliary materials. The oral medicine can be prepared into common oral medicine preparations such as capsules, tablets, pills, powder, granules, syrup and the like, and belongs to the protection scope of the invention. The preparation method can be prepared according to conventional methods in the pharmaceutical field, and the used auxiliary materials can be selected from common auxiliary materials in the pharmaceutical field according to different dosage forms. In specific implementation, proper auxiliary materials in the medical or edible field can be directly added into the prepared suspension and freeze-dried powder of the quercetin oral sustained-release preparation to prepare any dosage form or material for medical or food functions.
The invention has the beneficial effects that:
the ionic emulsifier chitosan nanoparticle modified quercetin oral sustained-release preparation provided by the invention utilizes the advantages of low toxicity, high biocompatibility, biodegradability and the like of chitosan and lauryl sodium sulfate to prepare nanoparticles which are non-toxic, harmless, good in biocompatibility and good in biodegradability, and the lauryl sodium sulfate is an oral preparation excipient approved by FDA and has the effect of promoting small intestine permeation and absorption, so that the prepared nanoparticle preparation can slowly release quercetin in the oral process, can effectively enter intestinal epithelial cells to promote absorption and has the effect of enhancing curative effect.
The quercetin sustained-release preparation provided by the invention is an oral preparation, can greatly improve the compliance of patients compared with injection preparations and suppositories, is easier to promote patients to take for treating various diseases, and thus obtains wider biomedical application.
The quercetin nanometer sustained-release preparation provided by the invention is a nanometer preparation, can utilize the advantage of small particle size of nanometer size, increases the contact area of the preparation on intestinal epithelial cells, further promotes the preparation to enter the intestinal epithelial cells more, promotes the drug absorption, enhances the treatment effect of quercetin, and provides a more convenient prospect for the full development and application of quercetin.
In the preparation method of the quercetin nanometer sustained-release preparation provided by the invention, both quercetin and chitosan are ingeniously dissolved in glacial acetic acid, and an organic solvent is not used, so that the problems of post-treatment for removing the solvent and solvent residue are solved, the process is simple, the cost is low, the requirement on reaction conditions is low, the industrialization is easy, the economic benefit and the environmental protection significance are brought, the social benefit is very high, and the application prospect is very high.
The preparation method of the ionic emulsifier chitosan nanoparticle modified quercetin oral sustained-release preparation can prepare milky suspension or white freeze-dried powder, the shape of the prepared suspension is spherical or ellipsoidal, the particle size of the suspension is less than 200 nanometers, the dispersion index of the suspension in ultrapure water is 0.128-0.213, the electromotive force is 30.10-45.01 millivolts, the encapsulation rate is 71.97-75.83%, the sustained-release period can reach 72 hours, and the preparation method has the advantages of small particle size, uniform dispersion, no adhesion, good stability, permeation promotion, easy absorption, controlled sustained release and the like.
The chitosan and the lauryl sodium sulfate have the effect of promoting absorption at the same time, and the emulsifying property of the lauryl sodium sulfate increases the solubility of the quercetin and other nano preparations, so that the activity of the quercetin can be inhibited and the penetration promoting effect can be achieved, the damage of degradation and damage of the long-term delivery of the quercetin from stomach to small intestine can be effectively protected, the chitosan nanoparticles can be further enhanced to penetrate into intestinal cells, and the quercetin can be promoted to exert the therapeutic activity in the cells.
In the preparation method for preparing the ionic emulsifier chitosan nanoparticle modified quercetin oral sustained-release preparation, the stable reaction is finally carried out, and the hydroxyl of the dextran-70 is combined with the hydroxyl of the chitosan nanoparticle through hydrogen bonds to form a barrier for preventing the wrapping crisp skin nanoparticle from degrading, so that the quercetin nanoparticle preparation is better prevented from degrading in the gastrointestinal severe environment delivery process, the stability of the oral preparation is improved, and the medicinal effect action time of the preparation is prolonged, the medicinal effect action is increased, and the bioavailability is increased.
Drawings
Fig. 1 is a flowchart of a preparation method of an ionic emulsifier chitosan nanoparticle modified quercetin oral sustained-release preparation prepared in example 1.
FIG. 2 is a transmission electron microscope image of the spheroidic and particle size less than 200 nm of an ionic emulsifier chitosan nanoparticle modified quercetin oral sustained-release preparation prepared in example 1;
FIG. 3 is Zeta potential diagram showing the high efficiency of improving the stability of quercetin oral sustained-release preparation modified with chitosan nanoparticles as an ionic emulsifier according to example 1;
FIG. 4 is a UV scanning spectrum of an ionic emulsifier chitosan nanoparticle modified quercetin oral sustained-release preparation prepared in example 1, demonstrating that quercetin is effectively encapsulated;
FIG. 5 is a graph of the example 6 demonstrating that the ionic emulsifier chitosan nanoparticle modified quercetin oral sustained-release preparation significantly improves the in-vitro sustained-release of quercetin;
FIG. 6 is a graph illustrating that the in vivo oral blood concentration-time curve of the ionic emulsifier chitosan nanoparticle modified quercetin oral sustained-release preparation is significantly higher than the oral absorption and effective bioavailability of quercetin alone in example 7.
Detailed Description
The present invention will be described in detail with reference to examples.
Example 1
As shown in FIG. 1, the present embodiment provides a method for preparing an oral sustained-release quercetin preparation modified with chitosan nanoparticles as an ionic emulsifier
The preparation method comprises the following steps:
(1) Preparation of raw materials
Preparing a cation crosslinking raw material: uniformly dispersing 0.6 g of chitosan in 40 ml of 1.1% glacial acetic acid aqueous solution;
preparing an anionic emulsifying crosslinking raw material: dispersing 0.6 g of sodium dodecyl sulfate in 40 ml of ultrapure water solution;
preparing stable reaction raw materials: dextran-70, 1.1 grams, was uniformly dispersed in 20 milliliters of ultrapure water solution.
(2) Emulsion ionic crosslinking reaction
0.04 g quercetin was precisely weighed and added to the cation cross-linked raw material stirred overnight at 1000 rpm at room temperature and filtered through a 0.45 μm filter membrane, and the pH was adjusted to 6.9 with sodium hydroxide to obtain mixture I. And then, slowly adding the anion cross-linking raw material into the mixture I dropwise at a magnetic stirring speed of 1100 revolutions per minute to carry out an emulsifying ion cross-linking reaction for 2.5 hours in total, and preliminarily obtaining the quercetin chitosan-loaded nano suspension.
(3) Stable reaction
The stable reaction process comprises the following steps: slowly dropwise adding dextran-70 uniformly dispersed in ultrapure water into the suspension prepared by the crosslinking reaction, stirring at 1100 r/min for stabilizing the reaction for 1 hour, and obtaining the chitosan-coated quercetin nanometer oral preparation with higher stability.
The flow chart of the preparation of the ionic emulsifier chitosan nanoparticle modified quercetin oral sustained-release preparation prepared in the embodiment is shown in figure 1, the transmission electron microscope of the prepared preparation is shown in figure 2, the zeta potential is shown in figure 3, and the ultraviolet scanning spectrum is shown in figure 4.
It can be seen that the ionic emulsifier chitosan nanoparticle modified quercetin oral sustained-release preparation prepared by the embodiment has the advantages of simple preparation process, easy operation, green and low energy consumption, and the used raw materials are economical and easily available, so that the preparation process is easy to industrialize, and the prepared preparation is spherical, has small particle size of 188.73 +/-5.26 nanometers, uniform distribution, good dispersion, no particle adhesion phenomenon and encapsulation rate of 75.83%.
In fig. 3, 0.04 g of quercetin was fully dissolved in 100 ml of glacial acetic acid aqueous solution to obtain a solution, a blank nanoparticle solution prepared in the same preparation process as in this example without adding quercetin and an ionic emulsifier chitosan nanoparticle modified quercetin oral sustained release preparation solution prepared in this example were diluted ten times, and 1 ml of the sample was measured by a particle size analyzer. From the results, the Zeta potential of the quercetin oral sustained-release preparation modified by the ionic emulsifier chitosan nanoparticles prepared in the embodiment is about +36 mv, and the quercetin oral sustained-release preparation has good stability. The Zeta potential of the quercetin oral sustained-release preparation modified by the ionic emulsifier chitosan nanoparticles prepared by the embodiment is a positive value, so that the preparation is easy to combine with intestinal cells with negative charges, the affinity of the nanoparticles and the intestinal cells is increased, and the preparation is easy to permeate the intestinal epithelial cells to enhance the oral absorption capacity of the quercetin.
In fig. 4, the results of the spectrograms of the three samples of the solution obtained by fully dissolving 0.04 g of quercetin in 100 ml of glacial acetic acid aqueous solution, the blank nanoparticle solution prepared by the same preparation process as that of the present example without adding quercetin, and the ionic emulsifier chitosan nanoparticle modified quercetin oral sustained-release preparation solution prepared by the present example were subjected to the spectroscopic measurement. The maximum absorption wavelength of quercetin is known to be 374 nm, so the ultraviolet scanning spectrum results of three samples can be known as follows: the chitosan nano oral sustained-release preparation prepared by the embodiment can effectively wrap quercetin and increase the solubility of the quercetin.
Example 2
Preparation of ionic emulsifier chitosan nanoparticle modified quercetin oral sustained-release preparation
The preparation method comprises the following steps:
(1) Preparation of raw materials
Preparing a cation crosslinking raw material: uniformly dispersing 0.3 g of chitosan in 40 ml of 0.8% glacial acetic acid aqueous solution;
preparing an anionic emulsifying crosslinking raw material: dispersing 0.3 g of sodium dodecyl sulfate in 40 ml of ultrapure water solution;
preparing stable reaction raw materials: 0.8 g of dextran-70 was uniformly dispersed in 20 ml of ultrapure water solution.
(2) Emulsion ionic crosslinking reaction
0.10g quercetin was precisely weighed and added to the cation crosslinked material stirred overnight at a magnetic stirring rate of 800 rpm at room temperature and filtered through a 0.45 μm filter membrane, and the pH was adjusted to 6.6 with sodium hydroxide to obtain a mixture I. Then, under the magnetic stirring speed of 900 revolutions per minute, the anion cross-linking raw material is gradually added into the mixture I drop by drop to carry out the emulsification ion cross-linking reaction, the total reaction time is 1.5 hours, and the nano-suspension of the quercetin carried in the chitosan is preliminarily obtained.
(3) Stable reaction
The stable reaction process is as follows: slowly dropwise adding dextran-70 uniformly dispersed in ultrapure water into the suspension prepared by the crosslinking reaction, stirring at 900 rpm for stable reaction for 0.5 hour, and obtaining the chitosan-coated quercetin nano oral preparation with high stability.
The transmission electron microscope image, zeta potential map and ultraviolet scanning spectrogram of the quercetin oral sustained-release preparation modified by the ionic emulsifier chitosan nanoparticles prepared in the embodiment are similar to those of the embodiment 1, and are not shown again for the sake of brevity.
The particle size of the quercetin oral sustained-release preparation modified by the ionic emulsifier chitosan nanoparticle prepared by the embodiment is 162.31 +/-5.48 nanometers, and the encapsulation rate is 80.15%.
Example 3
Preparation of ionic emulsifier chitosan nanoparticle modified quercetin oral sustained-release preparation
The preparation method comprises the following steps:
(1) Preparation of raw materials
Preparing a cation crosslinking raw material: uniformly dispersing 0.4 g of chitosan in 40 ml of 0.9% glacial acetic acid aqueous solution;
preparing an anionic emulsifying crosslinking raw material: dispersing 0.4 g of sodium dodecyl sulfate in 40 ml of ultrapure water solution;
preparing stable reaction raw materials: 0.9 g of dextran-70 was uniformly dispersed in 20 ml of ultrapure water solution.
(2) Emulsion ionic crosslinking reaction
0.08 g quercetin was precisely weighed and added to the cation-crosslinked material stirred overnight at 900 rpm at room temperature and filtered through a 0.45 μm filter membrane, and the pH was adjusted to 6.7 with sodium hydroxide to obtain mixture I. And then, slowly adding the anion cross-linking raw material into the mixture I dropwise at a magnetic stirring speed of 900 revolutions per minute to carry out an emulsifying ion cross-linking reaction, wherein the total reaction time is 1.5 hours, and preliminarily obtaining the quercetin chitosan-loaded nano suspension.
(3) Stable reaction
The stable reaction process is as follows: dextran-70 evenly dispersed in ultrapure water is slowly and dropwise added into the suspension prepared by the cross-linking reaction, and stirring and stabilizing reaction is carried out for 0.5 hour per minute at 1000 revolutions, so as to obtain the chitosan-coated quercetin nanometer oral preparation with higher stability.
The transmission electron micrograph, zeta potential map and ultraviolet scanning spectrogram of the quercetin oral sustained-release preparation modified by the ionic emulsifier chitosan nanoparticles prepared in the embodiment are similar to those in the embodiment 1, and are not shown for brevity.
The particle size of the quercetin oral sustained-release preparation modified by the ionic emulsifier chitosan nanoparticles prepared by the embodiment is 172.56 +/-4.83 nanometers, and the encapsulation efficiency is 73.57%.
Example 4
Preparation of ionic emulsifier chitosan nanoparticle modified quercetin oral sustained-release preparation
The preparation method comprises the following steps:
(1) Preparation of raw materials
Preparing a cation crosslinking raw material: uniformly dispersing 0.5 g of chitosan in 40 ml of 1.0% glacial acetic acid aqueous solution;
preparing an anionic emulsifying crosslinking raw material: dispersing 0.5 g of sodium dodecyl sulfate in 40 ml of ultrapure water solution;
preparing stable reaction raw materials: 1.0 g of dextran-70 was uniformly dispersed in 20 ml of an ultrapure aqueous solution.
(2) Emulsion ionic crosslinking reaction
0.06 gram of quercetin was precisely weighed and added to the cation-crosslinked raw material stirred overnight at 900 rpm at room temperature and filtered through a 0.45 micron filter membrane, and the pH was adjusted to 6.8 with sodium hydroxide to obtain mixture I. And then, slowly adding the anion cross-linking raw material into the mixture I drop by drop at a magnetic stirring speed of 900 revolutions per minute to carry out an emulsifying ion cross-linking reaction, wherein the total reaction time is 2.0 hours, and preliminarily obtaining the quercetin-loaded chitosan nano suspension.
(3) Stabilization of the reaction
The stable reaction process comprises the following steps: dextran-70 evenly dispersed in ultrapure water is slowly and dropwise added into the suspension prepared by the cross-linking reaction, and stirring is carried out for 1000 revolutions per minute to stably react for 1.0 hour, so as to obtain the chitosan-coated quercetin nanometer oral preparation with higher stability.
The transmission electron microscope image, the Zeta potential image and the ultraviolet scanning spectrogram of the quercetin oral sustained-release preparation modified by the ionic emulsifier chitosan nanoparticles prepared in the embodiment are similar to those of the embodiment 1, and are not shown again for the sake of brevity.
The particle size of the quercetin oral sustained-release preparation modified by the ionic emulsifier chitosan nanoparticle prepared by the embodiment is 179.62 +/-4.67 nanometers, and the encapsulation efficiency is 74.92%.
Example 5
Preparation of ionic emulsifier chitosan nanoparticle modified quercetin oral sustained-release preparation
The preparation method comprises the following steps:
(1) Preparation of raw materials
Preparing a cation crosslinking raw material: uniformly dispersing 0.5 g of chitosan in 40 ml of 1.0% glacial acetic acid aqueous solution;
preparing an anionic emulsifying crosslinking raw material: dispersing 0.4 g of sodium dodecyl sulfate in 40 ml of ultrapure water solution;
preparing stable reaction raw materials: 0.9 g of dextran-70 was uniformly dispersed in 20 ml of an ultrapure aqueous solution.
(2) Emulsion ionic crosslinking reaction
0.08 g of quercetin was precisely weighed and added to the cation-crosslinked raw material stirred overnight at a magnetic stirring rate of 800 rpm at room temperature and filtered through a 0.45 μm filter membrane, and the pH was adjusted to 6.7 with sodium hydroxide to obtain a mixture I. And then, slowly adding the anion cross-linking raw material into the mixture I drop by drop at a magnetic stirring speed of 900 revolutions per minute to carry out an emulsifying ion cross-linking reaction, wherein the total reaction time is 2.0 hours, and preliminarily obtaining the quercetin chitosan-loaded nano suspension.
(3) Stable reaction
The stable reaction process is as follows: slowly dropwise adding dextran-70 uniformly dispersed in ultrapure water into the suspension prepared by the crosslinking reaction, stirring at 1100 r/min for stabilizing the reaction for 0.5 h to obtain the chitosan-coated quercetin nanometer oral preparation with high stability.
The transmission electron microscope image, the Zeta potential image and the ultraviolet scanning spectrogram of the quercetin oral sustained-release preparation modified by the ionic emulsifier chitosan nanoparticles prepared in the embodiment are similar to those of the embodiment 1, and are not shown again for the sake of brevity.
The particle size of the quercetin oral sustained-release preparation modified by the ionic emulsifier chitosan nanoparticle prepared by the embodiment is 171.83 +/-5.82 nanometers, and the encapsulation rate is 74.11 percent.
Observed under a transmission electron microscope, the quercetin oral sustained-release preparations prepared in examples 1 to 5 have small particle size, spherical particles, uniform distribution, good dispersion, and no particle adhesion phenomenon. The quercetin oral sustained release preparation products prepared in examples 1 to 5, which were analyzed by a particle size analyzer, all had a size of less than 200 nm, a dispersion index of 0.128 to 0.213, and an electromotive force of 30.10 to 45.01 mv, and obtained an ultraviolet absorption spectrum similar to that of fig. 4, were sufficiently encapsulated in quercetin of the above examples.
Example 6
Release test of ionic emulsifier chitosan nanoparticle modified quercetin oral sustained-release preparation
Phosphate Buffered Saline (PBS) with the pH value of 7.4 is taken as a release medium, 4 mg of quercetin is accurately weighed and is uniformly mixed with 20 ml of the release medium to obtain a control quercetin release solution, and then the quercetin nano-preparation suspension prepared in any implementation bar is uniformly mixed with the release medium to obtain a quercetin oral sustained-release preparation release experimental group with the total volume of 20 ml. Then, they were transferred into a dialysis bag soaked in distilled water, both ends of the dialysis bag were tied to the slurry, placed in a dissolution flask containing 250 ml of release medium, respectively, and shaken at constant speed in a constant temperature (37 ℃) shaker (120 rpm).
Respectively sampling 2 ml of samples at set time, storing in a refrigerator at-4 ℃, supplementing 2 ml of release medium solution after each sampling, centrifuging the sampled samples for 15 minutes at-4 ℃ at 10000 rpm by a high-speed low-temperature centrifuge, taking 1 ml of supernatant, filtering by a 0.45-micron microporous filter membrane, and taking the supernatant.
Measuring the content of quercetin in the sample supernatant at different time points by high performance liquid chromatography, sampling 20 microliter each time, measuring the peak area of quercetin, calculating the concentration of quercetin according to a standard curve regression equation, obtaining the cumulative quercetin release percentage, and drawing a drug release curve, wherein the experimental result is shown in figure 5.
The results of FIG. 5 show that the ionic emulsifier chitosan nanoparticles can slowly control the release of quercetin, and the release time is as long as 3 days, which effectively proves that the quercetin modified by the ionic emulsifier chitosan nanoparticles has the sustained and controlled release effect.
Example 7
Oral bioavailability test of ionic emulsifier chitosan nanoparticle modified quercetin oral sustained-release preparation
Adult healthy SD male rats 12 were prepared and divided into two groups on average: in groups A and B, after fasting for 12 hours, blood is taken from retroorbital sinus as a blank control sample, and then 1 ml of liquid medicine is given to each 100 g of the weight of the rat to irrigate the stomach of 12 rats. Wherein, 0.04 g of quercetin is fully dissolved in 100 ml of glacial acetic acid (0.1%) water solution is given to group A, and the quercetin nanometer oral sustained-release preparation solution prepared in the embodiment is given to group B, and different gastric lavage time is respectively recorded.
At 2 hours, 4 hours, 6 hours, 8 hours, 10 hours, 12 hours, 14 hours, 16 hours, 18 hours, 20 hours, 22 hours and 24 hours after gastric lavage of the rats in the two groups, 0.5 ml of blood is taken from the retroorbital sinus of the rats and is placed in a 5 ml anticoagulant-containing vacuum blood collection tube. Then, 3 ml of methanol is added to precipitate blood protein components, the mixture is vortexed for 10 minutes, the centrifugation is carried out for 10 minutes at 1000 revolutions per minute, the supernatant is filtered by a 0.45 micron filter membrane, 20 microliters of filtrate is precisely absorbed, and the concentration of quercetin in the filtrate is measured by adopting high performance liquid chromatography. And a blood concentration-time curve of quercetin and quercetin nano-formulations was plotted as shown in fig. 6.
The results of the analysis of figure 6 show that the plasma peak concentration of the quercetin oral sustained-release preparation solution modified by the ionic emulsifier chitosan nanoparticle is higher than that of the quercetin solution and the peak reaching time is prolonged, and meanwhile, the area under the curve of the blood concentration-time curve of the quercetin oral sustained-release preparation modified by the ionic emulsifier chitosan nanoparticle is obviously larger than the area under the curve of the quercetin. The result proves that the quercetin oral sustained-release preparation modified by the ionic emulsifier chitosan nanoparticle can obviously improve the bioavailability of quercetin and can achieve the purpose of slowly and continuously taking oral drugs for absorption.
The results of the quercetin in vitro release test and the oral bioavailability (blood concentration-time curve) test of the ionic emulsifier chitosan nanoparticle modified quercetin oral sustained-release preparation prepared in examples 1 to 5 are similar to those of the quercetin sustained-release oral preparation prepared in example 1, and are not shown again for the sake of brevity.
The foregoing is a detailed description of the invention with reference to specific embodiments, and the practice of the invention is not to be construed as limited thereto. Various modifications and changes may be made by those skilled in the art without departing from the spirit and scope of the invention, and any such modifications, equivalents, improvements, etc. are intended to be included within the scope of the invention.

Claims (5)

1. An ion emulsifier chitosan nanoparticle modified quercetin oral sustained-release preparation is characterized in that: comprises wrapped medicine quercetin, wrapping materials chitosan and lauryl sodium sulfate, the particle diameter of the quercetin nano sustained-release preparation is 160-180 nm, the encapsulation rate is 71.97-75.83%, and the preparation method comprises the following steps:
(1) Preparation of raw materials
Preparing a cation crosslinking raw material: evenly dispersing chitosan in glacial acetic acid aqueous solution, wherein 0.3-0.6 g of chitosan is dispersed in 0.8-1.1% glacial acetic acid aqueous solution per 40 ml of cation crosslinking raw material;
preparing an anionic emulsifying crosslinking raw material: dispersing anionic emulsifier or oral permeation enhancer in ultrapure water, wherein 0.3-0.6 g of anionic emulsifier is dispersed in each 40 ml of ultrapure water solution in the anionic crosslinking raw material;
preparing stable reaction raw materials: dispersing stabilizer in ultrapure water uniformly, and dispersing 0.8-1.1 g of stabilizer in each 20 ml of ultrapure water solution in the stable reaction raw material;
(2) Crosslinking reaction
The crosslinking reaction adopts an emulsifying ion crosslinking reaction;
the emulsifying ionic crosslinking reaction process comprises the following steps: adding quercetin into cation crosslinking raw material which is stirred overnight and filtered by a filter membrane, carrying out ultrasonic treatment to fully dissolve the quercetin, adjusting pH to be neutral by using sodium hydroxide to obtain a mixture I, and then dropwise and slowly adding anion crosslinking raw material into the mixture I to carry out emulsification ion crosslinking reaction.
(3) Stable reaction
The stable reaction process is as follows: the stabilizing agent dispersed in the ultrapure water is slowly and dropwise added into the solution prepared by the cross-linking reaction to be stirred for stabilizing the reaction, and the polymer-coated quercetin nanometer preparation with higher stability can be obtained.
2. The ionic emulsifier chitosan nanoparticle modified quercetin oral sustained release formulation as claimed in claim 1, characterized in that: the emulsifier, the anionic cross-linking agent and the oral penetration enhancer are all sodium dodecyl sulfate, and the stabilizer is dextran-70.
3. The ionic emulsifier chitosan nanoparticle modified quercetin oral sustained-release formulation as set forth in claim 1 or 2, wherein: in the process of the emulsification ion crosslinking reaction, the mass concentration of the quercetin in the mixture I is 1.0-2.5 g/L, and the pH value of the solution is 6.6-6.9 after the pH value of the solution is adjusted to be slightly neutral by sodium hydroxide.
4. The ionic emulsifier chitosan nanoparticle modified quercetin oral sustained-release formulation as set forth in claim 1 or 2, wherein: the emulsion ion crosslinking reaction time is 1.5-2.5 hours, and the stable reaction time is 0.5-1.0 hour.
5. An oral medicament, comprising: an oral sustained-release formulation of quercetin comprising the ionic emulsifier chitosan nanoparticle modified according to any one of claims 1 to 4 and an adjuvant.
CN202111183969.3A 2021-10-11 2021-10-11 Quercetin oral sustained-release preparation modified by ionic emulsifier chitosan nanoparticles and preparation method thereof Active CN113797177B (en)

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