CN114376964B

CN114376964B - Puerarin nanoparticle film-forming hydrogel preparation and preparation method thereof

Info

Publication number: CN114376964B
Application number: CN202111542983.8A
Authority: CN
Inventors: 赵文英; 朱庆书; 金青; 刘蓓; 许宁; 田龙奇
Original assignee: Qingdao University of Science and Technology
Current assignee: Qingdao University of Science and Technology
Priority date: 2021-12-16
Filing date: 2021-12-16
Publication date: 2023-06-16
Anticipated expiration: 2041-12-16
Also published as: CN114376964A

Abstract

The invention discloses a transdermal puerarin nanoparticle film-forming hydrogel preparation, which takes puerarin as a raw material, carbomer 940 as a gel, zein, hydroxypropyl methylcellulose and the like as film-forming agents, and sodium carboxymethyl cellulose as a modifier. The preparation process comprises the following steps: (1) preparing puerarin nano-particles by a subcritical water method; (2) adding puerarin nano-particles into a film forming agent; (3) Adding puerarin and film forming agent mixture into gel matrix, and adding modifier and menthol. The preparation process is simple, green and environment-friendly; the prepared gel is yellowish and transparent, the medicine is uniformly dispersed, and can be smeared on the skin part under eyes to form a transparent medicine film after 5-10 minutes, and the medicine can be released continuously; the film-forming hydrogel preparation has good transdermal performance, has microcirculation improving effect, and can be used for relieving or treating diabetic eye diseases.

Description

Puerarin nanoparticle film-forming hydrogel preparation and preparation method thereof

Technical field:

the invention relates to improvement of puerarin pharmaceutical dosage form, in particular to a puerarin nanoparticle film-forming hydrogel preparation for relieving or treating diabetic eye diseases by transdermal administration, and also provides a preparation method of the preparation, belonging to the technical field of pharmaceutical preparations.

The background technology is as follows:

diabetic patients often experience retinopathy, resulting in vision loss or blindness. It is counted that 50% of diabetes mellitus can occur in about 10 years and up to 80% of diabetes mellitus can occur in more than 15 years. The heavier the diabetes, the older the age and the higher the chance of onset.

Puerarin is an isoflavone component extracted from Pueraria lobata (Willd.) Ohwi of Leguminosae, has effects of promoting blood circulation, removing blood stasis, and improving microcirculation, and has good therapeutic effect for treating diabetic eye disease, and is widely studied and valued in medicine community. But puerarin has the characteristics of low water solubility, poor membrane permeability, short half-life period, P-gp efflux effect, first pass metabolism effect of intestinal enzymes and the like, has low oral bioavailability, needs large-dose administration, and limits the clinical application of the puerarin. In recent years, puerarin injection is clinically used for treating retinal vascular diseases, fundus diseases and the like to achieve better effects. However, as puerarin injections are widely used in clinic, adverse reaction reports are also increasing. The monitoring shows that the puerarin injection can cause acute intravascular hemolysis, is easy to accumulate drug, generates toxicity and damages liver and kidney.

In order to overcome the defects of low bioavailability of puerarin for oral administration, easy hemolysis, allergy and the like caused by clinical use of injection, the invention develops a film-forming hydrogel preparation for transdermal administration. The medicine generally suitable for preparing the percutaneous hydrogel needs to have proper oil-water distribution coefficient and good transdermal performance. But puerarin is slightly soluble in water and ethanol, has weak transdermal capability, and is not suitable for preparing common external hydrogel preparation. Currently, in order to improve the transdermal absorption capacity of puerarin drugs, the puerarin drugs are mostly prepared into nano-preparations in the literature. The puerarin is prepared into lipid cubic liquid crystal for nasal administration to treat brain diseases as Tan Ning [ a nasal nano preparation puerarin liquid crystal nanoparticle and a preparation method thereof, CN201710460975.6 ], the cubic crystal has high viscosity and high strength characteristics suitable for the warm and moist internal environment of the nasal cavity, but not suitable for transdermal administration exposed in the environment. Zhang Jifen A puerarin lyophilized Pickering emulsion and its preparation method, CN201910568549.3, adopt the high-pressure homogenization technique to prepare puerarin nanocrystal, then prepare into self-stabilization Pickering emulsion through the high-pressure homogenization, the process is complex, and emulsion in long-term preservation, the emulsion drops are apt to fuse, rupture, etc., cause medicament to separate out, the thermodynamic stability is bad. Other researches for improving the bioavailability of puerarin by using a nano technology are mostly used for oral administration, and the researches for treating diabetic eye diseases by external use have not been reported yet.

The invention comprises the following steps:

the invention provides a puerarin nanoparticle film-forming hydrogel preparation for external application and percutaneous absorption, which solves the problems of low oral bioavailability, short half-life, large individual difference of in vivo metabolism, easy hemolysis and allergy of injection, easy loss of common gel preparation after application and the like of puerarin.

The invention can effectively improve the percutaneous absorption rate of puerarin and improve the bioavailability of the medicine.

The invention also provides a preparation method of the puerarin nanoparticle film-forming hydrogel agent.

The invention provides a puerarin nanoparticle film-forming hydrogel preparation, which mainly comprises the following components in percentage by mass: 0.1-1.0% puerarin, 1.0-5.0% gel, 0-10% film forming agent, 0-10% modifier and 0.1-1.0% menthol, distilled water to 100% total.

The gel is carbomer 940;

the film forming matrix is one or a combination of zein and hydroxypropyl methylcellulose;

the modifier is sodium carboxymethyl cellulose.

The preparation method of the puerarin nano film-forming hydrogel preparation provided by the invention comprises the following steps:

(1) Weighing appropriate amount of gel, film forming agent, modifier and menthol, placing into a beaker, adding appropriate amount of solvent to swell or dissolve respectively, and keeping;

(2) Adding puerarin into a pressurized extractor, taking water as solvent, maintaining at a certain pressure and temperature for a certain time, and adding puerarin solution into film forming agent;

(3) Adding the prepared puerarin-film forming agent mixed solution into gel matrix, adding modifier and menthol, stirring, adding distilled water to 100% of the total amount, and stirring to obtain puerarin nanoparticle film-forming hydrogel.

The invention discloses a puerarin nanoparticle film-forming hydrogel preparation, which comprises the following components in percentage by mass: 0.5% puerarin, 0.5% carbomer 940,0.1% zein, 2.0% sodium carboxymethylcellulose, 0.1% menthol and the balance distilled water to a total of 100%.

The preparation method of the puerarin nanoparticle film-forming hydrogel preparation comprises the following steps:

(1) And adding 80% ethanol to dissolve the zein with proper amount to obtain zein solution.

(2) Carbomer 940 is weighed in a beaker, deionized water is added, the temperature is heated to 50 ℃, swelling is carried out for 4 hours, and a proper amount of sodium hydroxide is added to adjust the pH of the solution to be neutral, so that the hydrogel matrix is obtained.

(3) Adding puerarin into pressure extractor, and maintaining at a certain temperature and pressure for a certain time. And (2) spraying the puerarin solution into the zein solution obtained in the step (1) from a nozzle of an extractor to obtain a puerarin-zein mixed solution.

(4) Adding the puerarin-zein mixed solution prepared in the step (3) into the prepared hydrogel matrix, adding sodium carboxymethylcellulose and menthol, stirring, adding the rest distilled water to 100% of the total amount, and uniformly mixing to obtain the puerarin nanoparticle film-forming hydrogel.

The beneficial effects of the invention are as follows: the solubility of puerarin in water is increased by adopting subcritical water technology, and then puerarin aqueous solution is sprayed into zein solution, so that the two-phase solution is subjected to intense impact and fully mixed. The zein-puerarin composite nano-particles with the particle size within the range of 100-1000nm can be prepared by controlling the factors such as the spraying speed, the concentration of the two-phase solution, the volume ratio and the like. The method can continuously form nanoparticles, and can be used for industrial production.

The zein and puerarin are prepared into a nano-composite, which has a protective effect on puerarin, and can slowly and permanently release the medicine after percutaneous absorption, thereby overcoming the defects of short half-life and quick elimination of puerarin in vivo.

According to the invention, carbomer 940 is taken as a gel matrix, and zein or (and) hydroxypropyl methylcellulose are added, so that the prepared hydrogel preparation has film forming property, and the drug release is more gentle and the use is more convenient; sodium carboxymethyl cellulose is added, so that the gel has thixotropic property and film forming property, and the physical state of the gel, the moisture resistance, the water resistance, the tensile strength, the transparency and the like of the film can be improved; menthol is added to obtain the pharmacological auxiliary effect of clearing liver and improving vision and the pharmacological effect of correcting taste and promoting permeation.

The invention has the main advantages that: the preparation process is simple, the cost is low, and the preparation is green and safe; the medicine is administrated transdermally in the form of hydrogel, which is more convenient and comfortable than the patch, and the prepared gel has the advantages of light and thin film, good concealment, no skin tightening feeling and improved medication compliance of patients after the film is formed; the hydrogel exists in a film form after being smeared, the medicine is released slowly for a long time, the preparation is not easy to run off, and the medicine effect is not reduced.

Description of the drawings:

FIG. 1 shows the gel prepared in example 1 applied to skin to form a film (1. Film; 2. Dry; 3. Form after film formation);

FIG. 2 is a plot of cumulative release rate versus time for the preparation of example 1 for extra-gel transdermal;

FIG. 3 is a plot of cumulative release rate versus time for the preparation of example 2 for extra-gel transdermal;

FIG. 4 is a plot of cumulative release rate versus time for the preparation of example 3 for extra-gel transdermal;

FIG. 5 is a plot of cumulative release rate versus time for the preparation of example 4 for extra-gel transdermal;

FIG. 6 is a plot of cumulative release rate versus time for the preparation of example 5 for extra-gel transdermal;

FIG. 7 is a plot of cumulative release rate versus time for the preparation of example 6 for extra-gel transdermal;

FIG. 8 is a graph showing the cumulative rate of release versus time for the direct preparation of a gel from puerarin drug substance.

The specific embodiment is as follows:

the specific embodiments of the present invention are illustrated by the following examples, but the scope of the present invention is not limited thereto. The puerarin treatment method comprises the steps of taking water as a solvent in a pressurized extractor, and preserving heat for 10min at the temperature of 125 ℃ under the pressure of 0.5 MPa.

Example 1

Gels (in 100 g) were prepared according to the following recipe:

puerarin 0.5g, carbomer 940.0 g, zein 0.1g, sodium carboxymethylcellulose 2.0g, menthol 0.l and distilled water in balance.

Swelling carbomer 940 with deionized water, adjusting pH to neutrality with NaOH to obtain gel matrix, dissolving zein with 80% ethanol, swelling sodium carboxymethylcellulose with deionized water; spraying the prepared puerarin solution into zein solution; then adding puerarin-zein mixed solution into gel matrix, adding sodium carboxymethylcellulose and menthol, adding water to 100g, stirring until gel is uniform and transparent, and preserving at room temperature. The in vitro transdermal cumulative release rate versus time curve is shown in figure 2.

Example 2

A gel (in terms of I00 g) was prepared according to the following recipe:

puerarin 0.5g, carbomer 940.0 g, hydroxypropyl methylcellulose 10.0g, sodium carboxymethylcellulose 4.0g, menthol 0.l and distilled water in balance.

Swelling carbomer 940 with deionized water, adjusting pH to neutrality with NaOH to obtain gel matrix, dissolving hydroxypropyl methylcellulose with 50% ethanol, swelling sodium carboxymethylcellulose with deionized water; dripping the prepared puerarin solution into hydroxypropyl methyl cellulose solution; then adding puerarin-hydroxypropyl methyl cellulose mixed solution into gel matrix, adding sodium carboxymethyl cellulose and menthol, adding water to 100g, stirring until gel is uniform and transparent, and preserving at room temperature. The in vitro transdermal cumulative release rate versus time curve is shown in figure 3.

Example 3

A gel (in terms of I00 g) was prepared according to the following recipe:

puerarin 0.5g, carbomer 940 1.0g, zein 0.1g, sodium carboxymethylcellulose 6.0g, menthol 0.l and the balance of distilled water.

Swelling carbomer 940 with deionized water, adjusting pH to neutrality with NaOH to obtain gel matrix, dissolving zein with 80% ethanol, swelling sodium carboxymethylcellulose with deionized water; spraying the prepared puerarin solution into zein solution; then adding puerarin-zein mixed solution into gel matrix, adding sodium carboxymethylcellulose and menthol, adding water to 100g, stirring until gel is uniform and transparent, and preserving at room temperature. The in vitro transdermal cumulative release rate versus time curve is shown in figure 4.

Example 4

A gel (in terms of I00 g) was prepared according to the following recipe:

puerarin 0.5g, carbomer 940.0 g, sodium carboxymethylcellulose 6.0g, menthol 0.l and distilled water in balance.

Adding deionized water into carbomer 940 for swelling, adjusting pH to neutrality by NaOH to obtain gel matrix, and swelling sodium carboxymethylcellulose with deionized water for use; spraying the prepared puerarin solution into a proper amount of deionized water solution, adding into a gel matrix, adding sodium carboxymethylcellulose and menthol, adding water to 100g, stirring until the gel is uniform and transparent, and preserving at room temperature. The in vitro transdermal cumulative release rate versus time curve is shown in figure 5.

Example 5

A gel (in terms of I00 g) was prepared according to the following recipe:

puerarin 0.5g, carbomer 940.0 g, zein 0.5g, menthol 0.l and distilled water in balance.

Swelling carbomer 940 with deionized water, adjusting pH to neutrality with NaOH to obtain gel matrix, and dissolving zein with 80% ethanol; spraying the prepared puerarin solution into zein solution; then adding puerarin-zein mixed solution into gel matrix, adding menthol, adding water to 100g, stirring until gel is uniform and transparent, and preserving at room temperature. The in vitro transdermal cumulative release rate versus time curve is shown in figure 6.

Example 6

A gel (in terms of I00 g) was prepared according to the following recipe:

puerarin 0.5g, carbomer 940.0 g, zein 0.2g, menthol 0.l and distilled water in balance.

Swelling carbomer 940 with deionized water, adjusting pH to neutrality with NaOH to obtain gel matrix, and dissolving zein with 80% ethanol; spraying the prepared puerarin solution into zein solution; then adding puerarin-zein mixed solution into gel matrix, adding menthol, adding water to 100g, stirring until gel is uniform and transparent, and preserving at room temperature. The in vitro transdermal cumulative release rate versus time curve is shown in figure 7.

Examination of film Forming Effect of hydrogel formulations

The hydrogel preparation was applied to human skin and the film formation time and state were recorded, and the results are shown in Table 1.

TABLE 1 film Forming Effect of hydrogels on skin

The hydrogel preparation of example 1 is yellowish and semitransparent in color, and forms a layer of film which is hardly visible to naked eyes on the skin after being smeared, and the film is tightly attached to the skin, has certain elasticity, does not influence the normal movement of the skin, and does not pollute clothes.

Skin allergy test (example 1)

According to the conventional method, 4 adult healthy rabbits are taken, ear fur with the area of 4cm multiplied by 4cm is removed by 8% sodium sulfide before the test, and no damage is observed after 24 hours. The treated rabbits were randomly divided into 2 groups. Group 1 is experimental group (coated hydrogel preparation), group 2 is blank group (distilled water is supplied), and skin allergy of rabbit is observed at 0h, 24h, 48h and 72h respectively.

Experimental results show that after administration, the skin of the ear of the rabbit does not have the changes of erythema, edema and the like, and the rabbit also does not have the systemic allergic reactions of asthma, unstable standing, shock and the like. Indicating that the hydrogel preparation has no allergic reaction after skin administration.

Acute toxicity test (example 1)

According to the conventional method, 4 adult healthy rabbits are taken, and the fur of the two sides of the spine of the rabbits is removed by about 3cm multiplied by 3cm before administration. The treated rabbits are randomly divided into 2 groups, and the experiment groups rub the skin by the sterilized frosted paper until the skin is damaged by blood seepage; the control group was not treated and was intact skin. Two groups of rabbits were treated with the hydrogel formulation, respectively. After 24 hours, the medicines were wiped off with warm water, observed, and then the spraying of the same medicines was continued for 7 days. Daily observations and recordings of changes in animal body weight, skin, hair, eyes and mucous membranes, respiration, limb movement and other toxic manifestations were made. The results are shown in Table 2

TABLE 2 Experimental results of acute toxicity at the back of rabbits

The results showed that the hydrogel formulation was not irritating to normal intact skin and damaged skin of rabbits over multiple skin irritation periods of 24, 48 and 72 hours.

Hydrogel formulation in vitro transdermal experiments

With Na ₂ S, removing abdominal hair of a healthy mouse, cleaning, killing by neck breakage, immediately cutting off the abdominal skin, removing the peeled adipose tissue and mucus tissue by a blade, cleaning by normal saline, and storing in a refrigerator for standby.

Gels were prepared as in examples 1-6. Taking out mouse skin from refrigerator, recovering to room temperature, and repeating with normal salineThe mouse skin was rinsed and gently wiped dry with filter paper. Franz diffusion cell (receiving cell volume 5.3mL, inner diameter 1cm, effective area 0.785cm was used ² ) The skin is secured between the supply chamber and the receiving chamber with the stratum corneum facing the supply chamber. Precisely weighing the gel, placing into a supply chamber, and spreading to closely contact with skin. 5ml (pH 7.4, L0 mmol/L) of Phosphate Buffer (PBS) was added to the receiving chamber, the water bath temperature was (32.+ -. 0.5) ℃, the constant speed was 300r/min, 5ml was sampled at regular intervals at L h, 2h, 4h, 6h, 8h, 10h, 12h and 24h, and 5ml of fresh medium was immediately replenished, filtered through a 0.45 μm microporous filter membrane, and the drug concentration in the receiving liquid was determined by HPLC.

The HPLC conditions were: XDB C18 column 5 μm,4.5mm×250mm, column temperature 30 ℃, detection wavelength 253nm, mobile phase 0.2% acetic acid-methanol (70:30), flow rate 0.8ml/min.

The results of 6 batches of gel transdermal delivery are shown below (Table 3), where D is the puerarin or puerarin-zein complex particle size, jss is the steady state drug transdermal rate, and Q is the 24 hour cumulative drug transdermal dose.

TABLE 3 results of gel transdermal experiments

Evaluation of the efficacy of hydrogel formulations in improving microcirculation (example 1 is taken as an example)

A bullfrog is taken, the bullfrog brain and spinal cord are destroyed, then the bullfrog is fixed on a bullfrog plate, a notch is cut at the side part of the abdomen, a small intestine is pulled out, the mesentery is unfolded, and the bullnose is fixed on the bullfrog plate. And dropwisely adding wilt liquid thereon to prevent drying. Observing the blood flow conditions in arterioles, venules and capillaries under a 100-fold mirror, and distinguishing the flow velocity, direction and characteristics of the arterioles, venules and capillaries; then, the hydrogel is diluted by adding a proper amount of any test solution, and is lightly dripped on a mesenteric blood vessel to observe the change of the caliber of the blood vessel and the blood flow velocity. The results are shown in Table 4.

TABLE 4 results of improved microcirculation experiments with hydrogel formulations

After the hydrogel preparation is added, the blood flow rate in the mesenteric circulation is accelerated, which indicates that the hydrogel preparation can relax blood vessels and promote the frog mesenteric microcirculation.

Claims

1. The puerarin nanoparticle film-forming hydrogel preparation is characterized by comprising the following raw material components in percentage by mass and volume:

0.5% puerarin, 0.5% carbomer 940,0.1% zein, 2.0% sodium carboxymethylcellulose, 0.1% menthol and the balance distilled water to a total of 100%.

2. The method for preparing the puerarin nanoparticle film-forming hydrogel preparation according to claim 1, which is characterized by comprising the following steps:

(1) Weighing carbomer 940, zein, sodium carboxymethylcellulose, and Mentholum, placing into a beaker, and adding appropriate amount of solvent to swell or dissolve respectively;

(2) Adding puerarin into a pressurized extractor, taking water as a solvent, preserving heat for a certain time under a certain pressure and temperature, and then spraying puerarin solution into zein solution;

(3) Adding the obtained puerarin-zein mixed solution into carbomer matrix, adding sodium carboxymethylcellulose, menthol and distilled water to total amount of 100%, and stirring.