Composite nanoparticle for promoting transdermal absorption and preparation method and application of carrier of composite nanoparticle

Abstract

The invention discloses a preparation method and application of a composite nano-particle for promoting transdermal absorption and a carrier of the composite nano-particle. The aminated zein-gallic acid composite nano-particle carrier prepared by taking aminated zein and gallic acid as raw materials through an anti-solvent method is a smooth sphere with positive charges, has the minimum particle size of 95.6 nm, has relatively good dispersity and stability, can encapsulate hydrophobic substances, and can be used for preparing the nano-particles. According to the zein-gallic acid nano-particle, the water solubility and stability of a carried substance can be improved, the transdermal absorption performance of the carried substance can also be improved, the zein-gallic acid nano-particle can more quickly and deeply permeate into the skin cuticle, the pharmacological activity of the carried substance is exerted, and the problem that existing zein-gallic acid nano-particles are poor in transdermal performance is solved. The carrier provided by the invention can carry various active substances and promote transdermal absorption of the active substances, and provides more effective means and methods for preparation of a carrying system of hydrophobic active substances and a transdermal enhancer.

Description

Technical Field

[0001] The present invention belongs to the technical field of biomedicine and cosmetics, and more specifically, relates to a preparation method and application of composite nanoparticles and carriers thereof for promoting transdermal absorption. Background Art

[0002] At present, for drugs or preparations containing active ingredients for external use, compared with oral administration, injection and other routes of administration, transdermal administration can not only avoid the first-pass effect of the liver, but also prevent various adverse reactions caused by excessive release rate during drug release. In the current research on transdermal administration, it is found that the penetration and diffusion rate of drug molecules in human skin will be limited by the stratum corneum of the skin, and the stability of drug molecules will also affect the transdermal performance of drugs. Therefore, how to increase the transdermal performance of drug molecules and maintain the stability of drug molecules in preparations are problems that need to be solved in the current research on transdermal preparations.

[0003] At present, the methods for promoting the transdermal absorption of drug molecules include: (1) chemical methods, such as the use of permeation enhancers; (2) physical methods, including iontophoresis, electroporation, ultrasound introduction, microneedle technology, needle-free injection delivery, etc.; (3) nanotechnology, such as making liposomes, ethosomes, transfersomes, microemulsions, solid lipid nanoparticles, etc. However, due to the great differences in the properties and modes of action of different drugs, the degree of transdermal absorption after using the above methods is different.

[0004] Most active drugs not only play a role in medicine, but are also commonly used in the field of cosmetics. They work through transdermal absorption through skin application. The transdermal transmission of the effective ingredients of skin care products refers to the process in which the effective ingredients in skin care products pass through the skin, reach different skin layers and play a role. Most of the effective ingredients of skin care products on the market can reach the epidermis, accumulate and play a role in this part, and very few ingredients can enter the dermis to play a transdermal role. In the fields of medicine and cosmetics, most drugs with good anti-inflammatory, antioxidant, and anti-aging effects are fat-soluble substances. The fat-soluble substances can penetrate the semipermeable membrane of keratinocytes for absorption and are difficult to play a role deeply. For example, hydroxypinacolone retinoic acid ester, galangin, sanpinin, cinnamyl alcohol, etc. have poor stability and water solubility, which makes the active ingredients easy to precipitate from the product and difficult to penetrate the stratum corneum of the skin to play a role, limiting their application.

[0005] In recent years, there have been many studies on the use of natural permeation enhancers and the application of nano-transdermal technology. However, there are still many problems in the different nano-delivery systems prepared by them: for example, the disadvantage of using liposome nanoparticles is that the active substance loading capacity is not high enough, and the embedded active contents are easily excluded, and the transdermal absorption effect is not good; while the thermodynamics of the nanoemulsion system is unstable, and the active particles are distributed in both the aqueous phase and the oil phase, so the system stability and transdermal amount are further reduced. Existing studies all use nano-delivery systems to encapsulate hydrophobic substances or unstable active substances that are easily oxidized and photolyzed to improve their stability; for example, existing studies use nanoparticles prepared by zein and gallic acid, and modify them by adding gallic acid to increase the stability of Zein nanoparticles, but the transdermal performance of the nano-drug delivery system prepared by them is poor. At present, there is still a lack of drug delivery systems that can be used to deliver different active substances and promote transdermal absorption. Summary of the invention

[0006] The technical problem to be solved by the present invention is to overcome the defect of poor transdermal performance of existing fat-soluble active ingredients and the problem of poor transdermal performance of existing zein and gallic acid nanoparticles, and to provide a preparation method and application of composite nanoparticles and carriers thereof that promote transdermal absorption.

[0007] The purpose of the present invention is to provide a method for preparing a composite nanoparticle carrier for promoting transdermal absorption.

[0008] Another object of the present invention is to provide a composite nanoparticle carrier.

[0009] Another object of the present invention is to provide application of the composite nanoparticle carrier.

[0010] Another object of the present invention is to provide a product that promotes transdermal absorption.

[0011] Another object of the present invention is to provide composite nanoparticles.

[0012] The above-mentioned purpose of the present invention is achieved through the following technical solutions:

[0013] The present invention provides a method for preparing a composite nanoparticle carrier for promoting transdermal absorption, comprising the following steps:

[0014] S1. Preparation of an amino zein solution: dissolving zein in an organic solvent, adding 1.5 to 3% aminopropylene glycol, heating and stirring, dialyzing and freeze-drying after the reaction to obtain an amino zein; adding the amino zein to an organic solvent, ultrasonicating, stirring, and preparing an amino zein solution;

[0015] S2. Add gallic acid to the amino-zein solution, perform ultrasonic treatment, stir, and remove the alcohol solvent to obtain a composite nanoparticle carrier.

[0016] The aminated zein-gallic acid nanoparticle drug delivery system prepared by the anti-solvent method in the present application has more positive charges by using aminated zein, which can form more stable nanoparticles with negatively charged gallic acid, which can not only improve the stability of the nanoparticle carrier, but also improve the transdermal performance of the carrier. The composite nanoparticles prepared by the present invention are smooth spheres with positive charges, and the minimum particle size is 95.6nm, with good dispersibility and stability, and can be used for encapsulating fat-soluble active substances, such as hydroxypinacolone retinoic acid ester, galangin, shanjiangin, and cinnamyl alcohol, which improves the water solubility and stability of fat-soluble active ingredients, and improves the transdermal performance of its drugs, so that it can play a role in the stratum corneum, and penetrate to 20μm below the stratum corneum within 20min. It can penetrate into the stratum corneum of the skin faster and deeper, exert the pharmacological activity of its carrier, and solve the problem of poor transdermal performance of existing zein-gallic acid nanoparticles.

[0017] Preferably, the concentration of the amino-zein alcohol solution prepared in S1 is 20-60 mg / mL.

[0018] Preferably, the heating and stirring conditions in S1 are: temperature 30-40° C., 500-700 rpm, and 1-2 h.

[0019] Preferably, the ultrasonic conditions in S1 and S2 are: power 350-450 W, time 15-25 min, temperature 30-40° C.

[0020] More preferably, the preparation method of the amino zein in S1 is: adding the zein to an 80% ethanol aqueous solution, adding 2% aminopropylene glycol, stirring at 35° C. and 500 rpm for 1.5 h, dialyzing and freeze-drying to obtain the amino zein.

[0021] More preferably, the preparation method of the alcohol solution of amino-zein in S1 is: adding amino-zein to 80% ethanol aqueous solution, ultrasonicating at 400W for 20min at 35°C, and stirring at 600rpm for 30min.

[0022] Preferably, the material-liquid ratio of gallic acid to amino-zein solution in S2 is 10:1-2 mg / mL.

[0023] More preferably, the solid-liquid ratio of gallic acid to the amino-zein alcohol solution is 10:1 mg / mL.

[0024] The invention provides a composite nanoparticle carrier, which is prepared by the above method.

[0025] The invention provides application of a composite nanoparticle carrier in carrying a fat-soluble active substance and promoting its transdermal absorption effect.

[0026] The invention provides application of a composite nanoparticle carrier in preparing a product for promoting transdermal absorption.

[0027] The invention also provides a product for promoting transdermal absorption, which contains a composite nanoparticle carrier.

[0028] Preferably, when the product is a cosmeceutical or skin care product, the active substance can be embedded in the composite nanoparticles, and the embedding preparation can be carried out by conventional methods in the art.

[0029] Furthermore, the active substance is a fat-soluble substance, specifically hydroxypinacolone retinoic acid ester, galangin, sanpinin or cinnamyl alcohol.

[0030] In addition, the present invention provides a composite nanoparticle, which is prepared by encapsulating a fat-soluble active substance in a composite nanoparticle carrier; the nanoparticle is obtained by self-assembly of amino zein to encapsulate the fat-soluble active substance, and gallic acid is adsorbed on the surface of the nanoparticle.

[0031] Furthermore, the fat-soluble active substance is selected from one or more of hydroxypinacolone retinoic acid ester, galangin, sanpinin, and cinnamyl alcohol.

[0032] The present invention has the following beneficial effects:

[0033] The invention uses aminated zein (Zein) and gallic acid (GA) as raw materials, and prepares the aminated zein-gallic acid composite nanoparticles through an anti-solvent method. The composite nanoparticles are smooth spheres with positive charge, and have a minimum particle size of 95.6 nm. The composite nanoparticles have good dispersibility and stability, and can be used to encapsulate hydrophobic active substances. The aqueous solubility and stability of the encapsulated active substance components can be improved, and the transdermal absorption performance of the carried substances can be improved. The carried substances can penetrate the stratum corneum of the skin faster, thereby exerting pharmacological activity. The carrier is suitable for a variety of substances with different activities, and can promote their transdermal absorption performance, thereby providing more effective means and methods for the preparation of the carrying system of hydrophobic active substances and transdermal enhancers. BRIEF DESCRIPTION OF THE DRAWINGS

[0034] Figure 1 is the encapsulation efficiency of Zein-gallic acid nanoparticles with different Zein to active ingredient mass ratios.

[0035] Figure 2These are the results of different nanoparticle sizes (A) and PDI (B) (a: Zein nanoparticles; b: hydroxypinacolone retinoic acid ester Zein-gallic acid nanoparticles; c: galangin Zein-gallic acid nanoparticles; d: galangin Zein-gallic acid nanoparticles; e: cinnamyl alcohol Zein-gallic acid nanoparticles).

[0036] Figure 3 These are the Zeta potential diagrams of different nanoparticles (a: Zein nanoparticles; b: hydroxypinacolone retinoic acid ester Zein-gallic acid nanoparticles; c: galangin Zein-gallic acid nanoparticles; d: galangin Zein-gallic acid nanoparticles; e: cinnamyl alcohol Zein-gallic acid nanoparticles).

[0037] Figure 4 These are SEM images of different nanoparticles (a: Zein nanoparticles; b: hydroxypinacolone retinoic acid ester Zein-gallic acid nanoparticles; c: cinnamyl alcohol Zein-gallic acid nanoparticles).

[0038] Figure 5 This is a comparison chart of Raman spectra of blank skin and skin applied with Alpinia officinalis.

[0039] Figure 6 This is a comparison of Raman spectra of blank skin and skin applied with galangin.

[0040] Figure 7 This is a comparison of Raman spectra of blank skin and skin applied with hydroxypinacolone retinoic acid ester.

[0041] Figure 8 This is a comparison of the Raman spectra of blank skin and skin applied with cinnamyl alcohol.

[0042] Fig. 9 These are the Raman spectra at different depths of the skin stratum corneum 20 minutes after applying ethanol solutions of different active ingredients (from left to right in the picture: Zein-gallic acid nanoparticles coated with galangin, galangin, hydroxypinacolone retinoic acid ester, and cinnamyl alcohol).

[0043] Fig.10 These are the Raman spectra at different depths of the skin stratum corneum 20 minutes after applying Zein-gallic acid nanoparticles with different active ingredients (from left to right in the figure: Zein-gallic acid nanoparticles applied with galangin, hydroxypinacolone retinoic acid ester, and cinnamyl alcohol). DETAILED DESCRIPTION

[0044] The present invention is further described below in conjunction with the accompanying drawings and specific examples, but the examples do not limit the present invention in any form. Unless otherwise specified, the reagents, methods and equipment used in the present invention are conventional reagents, methods and equipment in the art.

[0045] Unless otherwise specified, the reagents and materials used in the following examples are commercially available.

[0046] Hydroxypinacone Retinoate is a new generation of anti-aging ingredients in the vitamin A family. It combines the advantages of retinoic acid and retinoic acid esters. When the retinoic acid receptors in cells bind to HPR, it can promote cell proliferation, the synthesis of polysaccharides and extracellular proteins, thereby reducing wrinkles and anti-aging effects.

[0047] Galangin is a natural flavonol compound that can be extracted and separated from the rhizome of the traditional Chinese medicine Alpinia galanga and propolis in many regions. Due to its antioxidant, antiviral, antibacterial and anti-inflammatory effects, it has been widely used in the treatment of some cardiovascular and cerebrovascular diseases, anti-aging, and beauty care.

[0048] Alpinetin is a flavonoid found in herbal plants of the Leguminosae and Zingiberaceae families. It is the main component of the seeds of the Zingiberaceae plant Alpinia officinalis. It is also widely distributed in many families and genera of dicotyledonous plants. It has antibacterial, antioxidant, and anti-inflammatory effects, and is also characterized by low toxicity and high safety.

[0049] Cinnamic alcohol is an important unsaturated alcohol. Due to its unique aroma and anti-inflammatory, anti-tumor and antibacterial effects, it is widely used in the synthesis of flavors, fragrances, cosmetics, medicines and fungicides.

[0050] The zein used in the following examples was purchased from Bitart Pharmaceuticals with a purity of ≥12%; gallic acid with a purity of 99% was purchased from Weston Laboratory; hydroxypinacolone retinoate with a purity of 99% was purchased from MacLean Reagent; galangin with a purity of >98% was purchased from Chengdu Zhibiao Huachun Biotechnology Co., Ltd.; shanpinin with a purity of ≥98% was purchased from MacLean Reagent; and cinnamyl alcohol with a purity of ≥98% was purchased from Sichuan Weikeqi Biotechnology Co., Ltd.

[0051] Example 1 Preparation of Aminated Zein-Gallic Acid Composite Nanoparticles

[0052] 1. Preparation of aminated zein

[0053] The zein was added to 80% ethanol aqueous solution, and 2% aminopropylene glycol was added, and the mixture was stirred at 35° C. and 500 rpm for 1.5 hours. After the reaction, the mixture was dialyzed and freeze-dried to obtain the amino-zein.

[0054] 2.0 g of aminated zein was added to 50 mL of 80% ethanol aqueous solution, and ultrasonicated at a power of 400 W, a time of 20 min, a temperature of 35° C., and magnetically stirred at 600 rpm for 30 min to prepare a 40 mg / mL aminated zein solution.

[0055] 2. Preparation of aminated Zein-gallic acid

[0056] 500 mg of gallic acid was added to 50 mL of the aminated Zein solution, and ultrasonication was performed at a power of 400 W, a time of 20 min, and a temperature of 35° C. to prepare an aminated Zein-gallic acid solution with a gallic acid concentration of 10 mg / mL.

[0057] 3. Preparation of aminated Zein-gallic acid nanoparticle dispersion

[0058] Take 1 mL of the aminated Zein-gallic acid solution and quickly add it to 9 mL of pH = 4 deionized water. After magnetic stirring at 800 rpm for 30 minutes, remove the ethanol with a rotary evaporator, add 0.80 mL of pH = 4 deionized water to make up to 10 mL, let it stand and take the supernatant, which is the desired sample aminated Zein-gallic acid nanoparticle dispersion.

[0059] Example 2 Preparation of Zein-Gallic Acid Nanoparticle Dispersion for Carrying Active Substances

[0060] 1. Preparation of Hydroxypinacolone Retinoic Acid Zein-Gallic Acid Nanoparticle Dispersion

[0061] Take 5 mL of the aminated Zein-gallic acid solution prepared in Example 1, add 5 mg of hydroxypinacolone retinoic acid ester and ultrasonically treat for 30 min, power 400 W, time 20 min, temperature 35° C. Quickly add 1 mL of Zein-hydroxypinacolone retinoic acid ester-gallic acid solution to 9 mL of pH=4 deionized water, magnetically stir at 800 rpm for 30 min, remove ethanol with a rotary evaporator, add 0.80 mL of pH=4 deionized water to make up to 10 mL, let stand, and take the supernatant to obtain a dispersion of hydroxypinacolone retinoic acid ester Zein-gallic acid nanoparticles.

[0062] 2. Preparation of Zein-gallic acid nanoparticle dispersion

[0063] Take 5 mL of the aminated Zein-gallic acid solution prepared in Example 1, add 5 mg of galangin and ultrasonically treat for 30 min, the ultrasonic power is 400 W, the time is 20 min, and the temperature is 35° C. 1 mL of Zein-galangin-gallic acid solution is quickly added to 9 mL of pH = 4 deionized water, after magnetic stirring at 800 rpm for 30 min, the ethanol is removed by a rotary evaporator, 0.80 mL of pH = 4 deionized water is added to make up to 10 mL, and the supernatant is taken after standing to obtain the galangin Zein-gallic acid nanoparticle dispersion.

[0064] 3. Preparation of Zein-Gallic Acid Nanoparticle Dispersion

[0065] Take 5 mL of the aminated Zein-gallic acid solution prepared in Example 1, add 5 mg of shanjiangsu, and ultrasonically treat for 30 min, with an ultrasonic power of 400 W, a time of 20 min, and a temperature of 35° C. Quickly add 1 mL of the Zein-shanjiangsu-gallic acid solution to 9 mL of pH=4 deionized water, and after magnetic stirring at 800 rpm for 30 min, remove ethanol with a rotary evaporator, add 0.80 mL of pH=4 deionized water to make up to 10 mL, and take the supernatant after standing to obtain a dispersion of shanjiangsu Zein-gallic acid nanoparticles.

[0066] 4. Preparation of Zein-gallic acid nanoparticle dispersion

[0067] Take 5 mL of the aminated Zein-gallic acid solution prepared in Example 1, add 5 mg of cinnamyl alcohol and ultrasonically treat for 30 min, ultrasonic power 400 W, time 20 min, temperature 35 ° C. Quickly add 1 mL of Zein-cinnamyl alcohol-gallic acid solution to 9 mL of pH = 4 deionized water, magnetically stir at 800 rpm for 30 min, remove ethanol with a rotary evaporator, add 0.80 mL of pH = 4 deionized water to make up to 10 mL, let stand and take the supernatant to obtain cinnamyl alcohol Zein-gallic acid nanoparticle dispersion.

[0068] Example 3 Characterization of Aminated Zein-Gallic Acid Nanoparticles

[0069] 1. Determination of encapsulation efficiency

[0070] (1) Preparation of standard solution: Use an electronic balance to accurately weigh 2, 4, 6, 8, and 10 mg of the sample of active substance carrier prepared in Example 2 (hydroxypinacolone retinoic acid ester, galangin, sanpinin, and cinnamyl alcohol) and place them in a 100 mL volumetric flask. Add 80% ethanol aqueous solution to dissolve and dilute to the mark, shake well, then take out 1 mL of the solution and place it in a 10 mL volumetric flask, add 80% ethanol aqueous solution to dilute to the mark, shake well, and obtain 2, 4, 6, 8, and 10 μg / mL standard solutions.

[0071] (2) Preparation of reference solution: Take 2 mL of the amino Zein-gallic acid nanoparticle dispersion prepared in Example 1, add 8 mL of anhydrous ethanol and 3.3 mL of 80% ethanol aqueous solution, mix and stir for 10 min, centrifuge at 10000 rpm for 20 min, and take the supernatant to obtain the reference solution.

[0072] (3) Preparation of the test solution: Take 2 mL of the nanoparticle dispersion carrying active substances prepared in Example 2, add 8 mL of anhydrous ethanol and 3.3 mL of 80% ethanol aqueous solution, mix and stir for 10 min, let stand and take the supernatant to obtain the test solution.

[0073] (4) Determination of detection wavelength: The prepared 10 μg / mL standard solution was scanned in the full band range of 200-800 nm with 80% ethanol aqueous solution as blank, and the maximum absorption wavelength was selected as the detection wavelength.

[0074] (5) Determination of the standard curve of the standard substance: The standard solution of different concentrations prepared above was used as a blank with 80% ethanol aqueous solution, and the absorbance was measured at the detection wavelength. The absorbance A (ordinate, y) was linearly regressed against the concentration c (μg / mL, abscissa, x) to obtain the linear regression equation of the standard substance.

[0075] (6) Determination of encapsulation efficiency: The test solution and reference solution prepared above were used as blanks with 80% ethanol aqueous solution, and the absorbance was measured at the detection wavelength and recorded. The absorbance of the reference solution was subtracted from the absorbance of the test solution, and the obtained absorbance was the sample absorbance. The sample content W encapsulated in the nanoparticles was calculated according to the linear regression equation obtained in (5).

[0076] The encapsulation efficiency can be calculated according to the following formula:

[0077] Encapsulation efficiency (EE%) = W package (mg) / total amount of added sample (mg) × 100%

[0078] At the same time, the encapsulation efficiency of aminated Zein-gallic acid nanoparticles with different mass ratios of aminated Zein to active ingredient (20:1, 30:1, 40:1) was determined. The results are as follows: Figure 1As shown in the figure, it can be seen that with the increase of the mass ratio of aminated Zein to active ingredients, the encapsulation rate of aminated Zein-gallic acid nanoparticles for active substances has a significant increase, and the highest encapsulation rate can reach 84.01%. Aminated Zein-gallic acid nanoparticles prepared when the mass ratio of aminated Zein to active ingredients is 40:1 can effectively encapsulate these four hydrophobic active substances, and the encapsulation effect is good. Therefore, the aminated Zein-gallic acid nanoparticles prepared with a mass ratio of aminated Zein to active ingredients of 40:1 were used for subsequent characterization experiments.

[0079] 2. Measurement of Nanoparticle Size, Zeta Potential and PDI

[0080] Blank aminated Zein nanoparticles (without adding gallic acid) (a) and hydroxypinacolone retinoic acid ester Zein-gallic acid nanoparticles (b), galangin Zein-gallic acid nanoparticles (c), alpinia officinalis Zein-gallic acid nanoparticles (d), and cinnamyl alcohol Zein-gallic acid nanoparticles (e) with a mass ratio of aminated Zein: active ingredient of 40:1 were diluted ten times with deionized water at pH = 4, and the particle size, zeta potential and PDI of the nanoparticles were measured by a zeta potential meter.

[0081] The results of the test are as follows Figure 2 As shown in A, it can be seen that the particle size of each group of nanoparticles is less than 160nm, and compared with the blank amino Zein nanoparticles, the particle size of the nanoparticles carrying active substances is reduced after the Zein nanoparticles are modified by adding gallic acid. The smallest particle size of the cinnamyl alcohol Zein-gallic acid nanoparticles reaches 95.6nm, which shows that the modification of gallic acid can significantly reduce the particle size of Zein nanoparticles. The reduction of the particle size of nanoparticles can enhance the stability of nanoparticles during storage and reduce the phenomenon of large particle aggregation, so that they can be better transported and stored.

[0082] The PDI of nanoparticles carrying active substances is greater than that of amino Zein nanoparticles, such as Figure 2 As shown in B, the overall distribution uniformity decreases after the nanoparticles encapsulate the hydrophobic active substance, but the PDI of all nanoparticles carrying the active substance is still less than 0.2, which shows that the dispersion properties of the particle dispersion are still very good. Figure 3It can be seen that compared with blank Zein nanoparticles, the surface potential of nanoparticles modified with gallic acid that carry active substances has increased. The highest potential of Zein-gallic acid nanoparticles reached +37.9mV. Generally speaking, nanoparticles can be considered stable if their absolute value of surface potential is greater than 20mV, which indicates that nanoparticles that carry hydrophobic active substances have better stability.

[0083] 3. SEM test

[0084] The blank aminated Zein nanoparticles, hydroxypinacolone retinoic acid ester Zein-gallic acid nanoparticles and cinnamyl alcohol Zein-gallic acid nanoparticles were further diluted with deionized water at pH = 4, and a small amount of the diluted sample solution was taken with a rubber-tipped dropper and dropped onto the conductive tape, dried at room temperature, and observed under SEM after gold spraying in high vacuum mode with an accelerating voltage of 5.0 kV.

[0085] The results are as follows Figure 4 As shown in the figure, it can be seen that the size of blank amino Zein nanoparticles is about 150nm, while the size of nanoparticles carrying active substances modified with gallic acid is about 100nm. The main morphology of the nanoparticles is spherical. The observed particle size is consistent with the measurement results of the zeta potential instrument, and has good structural density and stability.

[0086] At the same time, the prepared nanoparticles were characterized by using Fourier infrared spectrometer, UV-visible spectrophotometer and field emission scanning electron microscope. The results of infrared and ultraviolet showed that gallic acid and amino Zein were mainly combined by non-covalent bonds and hydrophobic interactions, thereby improving the performance of Zein nanoparticles. The amino-modified zein has more positive charges and can form more stable nanoparticles with negatively charged gallic acid, adsorbing more gallic acid, further enhancing the performance of amino Zein-gallic acid nanoparticles. The addition of gallic acid allows amino zein to interact with gallic acid through non-covalent binding, and the surface charge of amino zein increases, thereby enhancing the electrostatic repulsion between nanoparticles. Electrostatic repulsion plays a vital role in maintaining the stability of the nanosystem, so the enhancement of electrostatic repulsion will make the entire nanoparticle system carrying active substances more stable, which is more conducive to the transportation, storage and use of nanoparticles.

[0087] In summary, the particle size of the nanoparticles carrying active substances prepared in the present application is about 100 nm, the minimum particle size is 95.6 nm, the PDI is less than 0.20, the zeta potential is greater than +30 mV, the highest potential can reach +37.9 mV, the encapsulation rate of hydrophobic active substances is greater than 70%, and the highest encapsulation rate can reach 84.01%. The nanoparticle system has good stability.

[0088] Example 4 Transdermal Performance Analysis

[0089] 1. Preparation of active substance ethanol solution

[0090] Weigh 4 mg of hydroxypinacolone retinoic acid ester, galangin, sanpinin, and cinnamyl alcohol respectively into a 10 mL volumetric flask, add anhydrous ethanol to dissolve to the scale line, and shake well to obtain a 0.4 mg / mL active substance ethanol solution.

[0091] 2. Raman detection

[0092] The subjects were randomly selected healthy women aged 35 to 45. Before the test, they washed the backs of their hands with clean water and selected a 2cm×2cm area on the backs of the left and right hands as the test area. 3mL of active substance ethanol solution was evenly applied to the test area. After 20 minutes of application, the confocal Raman spectrometer was used to measure the laser wavelength of 785nm, the exposure time of 60s, and the detection range of 400-2000cm -1 The penetration of active substances in the skin was tested under the conditions of scanning depth range of 0 to 20 μm, step length of 5 μm, laser power of 100 mV and 10x eyepiece.

[0093] After the test is completed, clean the test area with clean water in time, and then evenly apply 3mL of Zein-gallic acid nanoparticle solution carrying different active substances on the test area. The test steps are the same as above. The subsequent active substance testing process is the same as above until the penetration of the four active substance ethanol solutions and Zein-gallic acid nanoparticle solution in the skin is tested. After testing with a confocal Raman spectrometer and processing with data processing software, the Raman spectrum results of each sample are obtained.

[0094] The Raman spectra of blank skin and skin applied with different active substances are shown in Figure 5-8 As shown, the characteristic peak of shanjiangsu is 1224 cm -1 galangin is characterized by a peak at 1406 cm -1 The characteristic peak of hydroxypinacolone retinoic acid ester is 1501cm -1 The characteristic peak of cinnamyl alcohol is 1435 cm -1 .

[0095] Raman spectra at different depths of the stratum corneum of the skin 20 minutes after applying ethanol solutions of different active ingredients are shown in Figure 2. Fig. 9 As shown, galangin and shanjiangin do not have obvious characteristic peaks at 5-20μm, which means that most of the galangin in the ethanol solution still stays on the surface of the skin stratum corneum and does not penetrate deep into the stratum corneum, indicating that the transdermal performance of galangin and shanjiangin is weak and cannot penetrate into the stratum corneum within 20 minutes. Hydroxypinacolone retinoic acid ester is more evenly distributed between 0-5μm in the stratum corneum, and no obvious characteristic peaks appear at 10-20μm, which means that hydroxypinacolone retinoic acid ester cannot penetrate here at this time. Cinnamyl alcohol has medium-intensity characteristic peaks at 5-15μm, and the intensity difference is not large. No obvious characteristic peaks are found at 20μm, which means that the cinnamyl alcohol in the ethanol solution is more evenly distributed at 5-15μm in the skin stratum corneum.

[0096] Raman spectra of different depths of the stratum corneum of the skin 20 minutes after applying Zein-gallic acid nanoparticles with different active ingredients are shown in Figure 2. Fig.10 As shown, the encapsulated shanjiangsu can partially penetrate to 5 μm below the stratum corneum in the same time, while most of the shanjiangsu that has not been encapsulated remains on the surface of the stratum corneum; the encapsulated galangin can partially penetrate to 10 μm below the stratum corneum in the same time, while most of the galangin that has not been encapsulated remains on the surface of the stratum corneum; the encapsulated hydroxypinacolone retinoic acid ester mostly penetrates to 5 μm below the stratum corneum in the same time, while most of the hydroxypinacolone retinoic acid ester that has not been encapsulated is distributed between 0-5 μm in the stratum corneum; the encapsulated cinnamyl alcohol mostly penetrates to 20 μm below the stratum corneum in the same time, while most of the cinnamyl alcohol that has not been encapsulated is distributed between 5-15 μm in the stratum corneum.

[0097] The above results prove that the transdermal performance of active substances encapsulated by Zein-gallic acid nanoparticles has been significantly improved, and they can penetrate the stratum corneum of the skin faster to exert pharmacological activity.

[0098] Example 5 Optimization of preparation conditions for amino-zein-gallic acid composite nanoparticles

[0099] In Example 1, the preparation of Zein solution and Zein-gallic acid solution involves ultrasonic treatment. The ultrasonic conditions are now optimized. The specific conditions are set as shown in Table 1 below. The preparation methods of Examples 1 and 2 are used to prepare Zein-gallic acid nanoparticles, respectively, and the Raman spectroscopy detection method of Example 4 is used for detection, and the skin penetration depth of the prepared nanoparticles is counted.

[0100] The results are shown in Table 1 below, which shows that ultrasonic conditions affect the binding between amino-zein and gallic acid, and after encapsulating galangin with amino-zein-gallic acid composite nanoparticles, its transdermal performance is significantly improved compared with that before encapsulation.

[0101] Table 1 Optimization of preparation conditions of amino-zein-gallic acid composite nanoparticles

[0102] Group Ultrasonic power Ultrasound time temperature Skin penetration depth 1 400W 20min 35℃ 5.0μm below the stratum corneum 2 350 W 20min 35℃ 4.5μm below the stratum corneum 3 450 W 20min 35℃ 4.8μm below the stratum corneum 4 400 W 15min 35℃ 4.8μm below the stratum corneum 5 400 W 25min 35℃ 5.0μm below the stratum corneum 6 400 W 20min 30℃ 4.4μm below the stratum corneum 7 400 W 20min 40℃ 4.6μm below the stratum corneum

[0103] Comparative Example 1

[0104] The difference from Example 1 is that zein is not amino-modified and gallic acid is not added. The specific method is as follows: zein is added to 80% ethanol aqueous solution, ultrasonicated at 400W for 20min, at 35°C, and stirred at 600rpm for 30min to obtain a zein solution. Take 5mL of zein, add 5mg of shanjiangsu and ultrasonicate for 30min, ultrasonic power of 400W, time of 20min, and temperature of 35°C. 1mL of Zein-shanjiangsu solution is quickly added to 9mL of pH=4 deionized water, magnetically stirred at 800rpm for 30min, ethanol is removed by rotary evaporator, 0.80mL of pH=4 deionized water is added to make up to 10mL, and the supernatant is taken after standing to obtain a shanjiangsu nanoparticle dispersion.

[0105] Comparative Example 2

[0106] The difference from Example 1 is that zein is not amino-modified. The specific method is as follows: zein is added to 80% ethanol aqueous solution, ultrasonicated at 400W for 20min, 35°C for 30min, and stirred magnetically at 600rpm for 30min to prepare zein solution. Gallic acid is then added to the rice alcohol-soluble protein solution and ultrasonicated to prepare a rice alcohol-soluble protein-gallic acid solution. 5mL of rice alcohol-soluble protein-gallic acid solution is taken, 5mg of shanjiangsu is added and ultrasonicated for 30min, ultrasonic power of 400W, time of 20min, and temperature of 35°C. 1mL of Zein-shanjiangsu solution is quickly added to 9mL of pH=4 deionized water, magnetically stirred at 800rpm for 30min, ethanol is removed by rotary evaporator, 0.80mL of pH=4 deionized water is added to make up to 10mL, and the supernatant is taken after standing to obtain a shanjiangsu nanoparticle dispersion.

[0107] Comparative Example 3

[0108] The difference from Example 1 is that gallic acid is not added. The specific method is as follows: 5 mL of amino zein is added, 5 mg of zein is ultrasonically treated for 30 min, the ultrasonic power is 400 W, the time is 20 min, and the temperature is 35° C. 1 mL of Zein-zein solution is quickly added to 9 mL of pH=4 deionized water, and after magnetic stirring at 800 rpm for 30 min, ethanol is removed by a rotary evaporator, 0.80 mL of pH=4 deionized water is added to make up to 10 mL, and the supernatant is taken after standing to obtain Zein nanoparticle dispersion.

[0109] Referring to the Raman spectroscopy detection method of Example 4, the shanjiangsu nanoparticles prepared in Comparative Examples 1-3 were tested, and the results are shown in Table 2 below, showing that the amino-zein-gallic acid composite nanoparticles prepared by the present invention not only improve the stability of the drug delivery system, but also significantly improve the transdermal absorption effect of the drug delivery system, and are used to encapsulate active drugs with poor stability and water solubility, and can penetrate the stratum corneum of the skin to exert their effects. For example, for encapsulating hydroxypinacolone retinoic acid ester, galangin, shanjiangsu, cinnamyl alcohol, etc., the transdermal absorption effect of the active ingredients can be significantly improved, and they can penetrate into the stratum corneum to exert their effects.

[0110] Table 2 Skin penetration depth of nanoparticles in comparative examples 1-3

[0111] Group Skin penetration depth Example 1 5.0μm below the stratum corneum Comparative Example 1 0.5μm below the stratum corneum Comparative Example 2 4.3μm below the stratum corneum Comparative Example 3 0.9μm below the stratum corneum

[0112] The above embodiments are preferred implementation modes of the present invention, but the implementation modes of the present invention are not limited to the above embodiments. Any other changes, modifications, substitutions, combinations, and simplifications that do not deviate from the spirit and principles of the present invention should be equivalent replacement methods and are included in the protection scope of the present invention.

Claims

1. A method for preparing a composite nanoparticle carrier for promoting transdermal absorption, characterized in that: The following steps are involved: S1. Preparation of an amino zein solution: dissolving zein in an organic solvent, adding 1.5 to 3% aminopropylene glycol, heating and stirring, dialyzing and freeze-drying after the reaction to obtain an amino zein; adding the amino zein to an organic solvent, ultrasonicating, stirring, and preparing an amino zein solution; S2. Add gallic acid to the amino-zein solution, perform ultrasonic treatment, stir, and remove the alcohol solvent to obtain a composite nanoparticle carrier.

2. The method according to claim 1, characterized in that: The concentration of the amino-zein alcohol solution prepared in S1 is 20-60 mg / mL.

3. The method according to claim 1, characterized in that: The heating and stirring conditions in S1 are: temperature 30-40°C, 500-700 rpm, 1-2 h.

4. The method according to claim 1, characterized in that: The ultrasonic conditions in S1 and S2 are: power 350-450 W, time 15-25 min, temperature 30-40 °C.

5. The method according to claim 1, characterized in that: The solid-liquid ratio of gallic acid to amino-zein solution in S2 is 10:1-2 mg / mL.

6. A composite nanoparticle carrier, characterized in that: The invention is prepared by the method according to any one of claims 1 to 5.

7. Use of the composite nanoparticle carrier according to claim 6 in delivering fat-soluble active substances and promoting their transdermal absorption effect.

8. Use of the composite nanoparticle carrier according to claim 6 in the preparation of products for promoting transdermal absorption.

9. A product for promoting transdermal absorption, characterized in that: Containing the composite nanoparticle carrier according to claim 6.

10. A composite nanoparticle, characterized in that: The nanoparticle carrier according to claim 6 is prepared by encapsulating a fat-soluble active substance; the fat-soluble active substance is selected from one or more of hydroxypinacolone retinoic acid ester, galangin, sanpinin, and cinnamyl alcohol.

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