WO2018054229A1

WO2018054229A1 - Micro-nucleic acid acne-removing composition

Info

Publication number: WO2018054229A1
Application number: PCT/CN2017/101023
Authority: WO
Inventors: 朱远源; 李铁军; 於如霞; 刘永红; 周宋峰; 孙云成
Original assignee: 百奥迈科生物技术有限公司
Priority date: 2016-09-21
Filing date: 2017-09-08
Publication date: 2018-03-29
Also published as: CN106420792A

Abstract

A micro-nucleic acid acne-removing composition consists of the following components by weight percent: 0.001-0.5% of micro-nucleic acid molecules and 99.50-99.999% of a cosmetic excipient or an externally applied agent excipient, wherein the micro-nucleic acid molecules have a length in a range of from 17-100 bp, and the source of the micro-nucleic acid molecules is Saccharomyces cerevisiae or Candida tropicalis or a synthetic double-chain RNA molecule. The micro-nucleic acid acne-removing composition can be used for removing and preventing facial acne.

Description

Small nucleic acid acne composition

Technical field

The present invention belongs to the field of cosmetics and pharmaceuticals, and in particular to an acne composition, in particular an acne cosmetic and an acne drug comprising a double-stranded small nucleic acid molecule.

Background technique

Acne, also known as acne, acne, etc., scientific name acne, is a chronic inflammatory skin disease of the hair follicle sebaceous gland unit, which occurs in adolescents, clinical manifestations of acne, papules, pustules, nodules, etc. Polymorphic lesions are characteristic. Acne is a hair follicle sebaceous gland disease caused by a combination of various factors. Acne is mainly related to factors such as excessive sebum secretion, hyperkeratosis of hair follicles, proliferation of P. acnes, and excessive immune response. After entering puberty, the level of androgen, especially testosterone, rises rapidly in the human body, promotes the development of sebaceous glands and produces a large amount of sebum. At the same time, the abnormal keratinization of the hair follicle sebaceous gland duct leads to clogging of the catheter and obstruction of sebum, which is the initial factor leading to acne. The desquamation is mixed with the filaments and the lipids to form a keratin plug, that is, micro-acne. A variety of microorganisms in the hair follicle, especially Propionibacterium acnes, multiply, and the lipase produced by Propionibacterium acnes decomposes sebum to form free fatty acids, and simultaneously oxidizes inflammatory cells and mediators, eventually inducing and aggravating the inflammatory response. According to the nature and severity of acne lesions, acne is divided into 3 degrees and 4 grades, specifically: grade 1 (mild), only acne; grade 2 (moderate), in addition to acne, there are some inflammatory Pimples; grade 3 (moderate), in addition to acne, there are more inflammatory papules or pustules; grade 4 (severe), in addition to acne, inflammatory papules and pustules, as well as nodules, cysts or scar.

At present, there are many drugs and methods for the treatment of hemorrhoids, but there is a lack of effective treatment methods, and some of the treatment methods use products with large side effects, ineffective effects, long course of treatment, easy recurrence, and palliative treatment. Common methods of acne treatment and many problems, including: (1) topical drugs, vitamin A acid, benzoyl peroxide, antibiotics (clindamycin, erythromycin, chloramphenicol, etc.), 壬二Acid, sulfur lotion, etc. However, the stability of retinoic acid itself is poor, and local stimuli are more common, with large side effects, which greatly limit its clinical application; benzoyl peroxide is a very effective topical antibiotic preparation, but due to traditional preparations It is prone to local irritation and limits its clinical application. (2) oral antibiotics, preferred tetracyclines (minocycline, doxycycline, etc.), followed by macrolides (erythromycin), although an effective means of treating acne, but appear during use The problem of drug resistance has become increasingly serious. (3) Oral isotretinoin, for severe acne. (4) anti-androgen therapy, such as oral contraceptive compound acetaminophen Progesterone tablets are suitable for women with moderate or severe acne, with high androgen levels (such as hairy, seborrhea, etc.) or polycystic ovary syndrome. (5) Oral glucocorticoids, mainly used for fulminant or polymeric acne. (6) Other treatments, such as photodynamic therapy (PDT), acid therapy, laser therapy, etc. At present, clinical treatment is mainly based on grade 3 and grade 4 of acne, but it is difficult to achieve the desired effect.

Although acne is a skin disease, it can seriously affect the patient's study, work, exercise, interpersonal relationship and social life, and damage the patient's physical and mental health. Therefore, there is an urgent need to develop a safe and effective acne product.

Summary of the invention

In order to overcome the above-mentioned drawbacks of the existing acne treatment techniques, the present invention applies small nucleic acids to cosmetics and external medicines, and develops novel skin care products and medicines which have remarkable and safe acne effects.

Accordingly, it is a first object of the present invention to provide an acne composition comprising a small nucleic acid molecule having a therapeutic effect on acne, including cosmetics and medicaments.

A second object of the present invention is to provide a method for preparing the above acne cosmetic and acne medication.

In order to achieve the above object, the present invention adopts the following technical solutions.

An acne composition consisting of the following components by weight: 0.001-0.5% of a small nucleic acid molecule, 99.50-99.999% of a cosmetic excipient or a topical excipient, wherein the length of the small nucleic acid molecule ranges from 17 to 100 bp.

Alternatively, the above small nucleic acid molecule is derived from Saccharomyces cerevisiae or Candida tropicalis, ie Saccharomyces cerevisiae total RNA or Candida tropicalis total RNA.

Alternatively, the small nucleic acid molecule described above is a synthetic double stranded RNA molecule.

Preferably, the artificially synthesized double-stranded RNA molecule is one selected from the two types of double-stranded RNA molecules described below, or a mixture of two or more thereof, consisting of a sense strand and an antisense strand, respectively:

The first type of small nucleic acid molecule,

Justice chain: 5'-UAGGUACUUCGAGAAGCCUAGUANn-3' (SEQ ID NO: 1),

Antisense strand: 5'-UACUAGGCUUCUCGAAGUACCUANn-3' (SEQ ID NO: 2);

The second type of small nucleic acid molecule,

The sense strand: 5'-AGGUACUUCGAGAAGCCNn-3' (SEQ ID NO: 3),

Antisense strand: 5'-GGCUUCUCGAAGUACCUNn-3' (SEQ ID NO: 4),

Wherein N is cytidine C, guanosine G, adenosine A, uridine U, deoxycytidine dC, deoxyguanosine dG, deoxyadenosine dA or deoxygenation Thymidine dT; n represents the number of N, and n is an integer of 0-2.

In a preferred embodiment, the first class of small nucleic acid molecules consists of the following sense and antisense strands:

Justice strand: 5'-UAGGUACUUCGAGAAGCCUAGUA-3' (SEQ ID NO: 5),

Antisense strand: 5'-UACUAGGCUUCUCGAAGUACCUA-3' (SEQ ID NO: 6); or

Justice strand: 5'-UAGGUACUUCGAGAAGCCUAGUAdTdT-3' (SEQ ID NO: 15),

Antisense strand: 5'-UACUAGGCUUCUCGAAGUACCUAdTdT-3' (SEQ ID NO: 16);

The second class of small nucleic acid molecules consists of the following sense and antisense strands:

The sense strand: 5'-AGGUACUUCGAGAAGCC-3' (SEQ ID NO: 7),

Antisense strand: 5'-GGCUUCUCGAAGUACCU-3' (SEQ ID NO: 8); or

The sense strand: 5'-AGGUACUUCGAGAAGCCdTdT-3' (SEQ ID NO: 13),

Antisense strand: 5'-GGCUUCUCGAAGUACCUdTdT-3' (SEQ ID NO: 14).

Preferably, the acne composition is an acne cosmetic, and the cosmetic auxiliary materials and the weight percentages are: 2.5-15% of the base oil, 1-20% of the humectant, 0.01-1.0% of the antioxidant, 0.1-2.0% of the thickener, Emulsifier 0.1-3%, plant oil control factor 1-5%, skin feel improver 0.1-2%, metal chelator 0-0.5%, fragrance 0.1-0.5%, the rest are water such as deionized water, purified water, distilled water Or double distilled water.

Alternatively, the acne composition is an acne drug, and the acne drug dosage form is an ointment, a cream, a cream, an expectorant, a liquid, a lotion, a gel, a film, or a spray, selected from the group consisting of pharmaceutically acceptable excipients for maintaining the stability of small nucleic acid molecules, skin permeation enhancers for promoting transdermal absorption of small nucleic acid molecules, and pharmaceutically acceptable fillings for the preparation of topical dosage forms And excipients, water such as deionized water, purified water, distilled water, or double distilled water, and the like.

Preferably, the above base fat is jojoba oil, dimethicone, or a mixture thereof.

Preferably, the humectant is glycerin, 1,3-butanediol, 1,2-pentanediol, 1,2-hexanediol/octylene glycol, hyaluronic acid, glyceryl octylate, or two or more thereof. mixture.

Preferably, the above antioxidant is oil soluble vitamin E acetate, sodium phytate, or a mixture thereof.

Preferably, the thickening agent is acryloyldimethyltaurylamine/VP copolymer, xanthan gum, or a mixture thereof.

Preferably, the above emulsifier is cetearyl garate, sorbitan olive oil, or a mixture thereof.

Preferably, the plant oil control factor is a horse chestnut seed extract, a sage leaf extract, a sage (Sage) extract, a coltsfoot leaf extract, a rosemary leaf extract, a mother chrysanthemum (European daisy) Extract, fragrant bee (melissa) leaf extract, or a mixture of two or more thereof.

Preferably, the above skin feel improving agent is a silicon elastomer.

Preferably, the above metal chelating agent is disodium edetate.

In a preferred embodiment, the acne cosmetic comprises the following components by weight: hojoba oil 1-10%, dimethyl silicone oil 1-5%, glycerin 2-10%, 1,3-butyl Diol 1-10%, 1,2-pentanediol 0.1-5%, 1,2-hexanediol/octylene glycol 0.1-2%, hyaluronic acid 0.01-5%, caprylic acid glyceride 0.1-1% , oil-soluble vitamin E acetate 0.1-0.5%, acryloyldimethyltaurylamine / VP copolymer 0.1-1.0%, xanthan gum 0.1-1%, sodium phytate 0.01-0.5%, cetyl stearin Alcohol oleate / sorbitan oleate 0.1-3%, plant oil control factor 1-5%, silica elastomer 0.1-2%, disodium edetate 0-0.5%, small nucleic acid 0.001-0.5 %, fragrance 0.1-0.5%, the balance is deionized water.

A method for preparing the above acne cosmetic comprising the following steps:

1) A phase mixture configuration, weigh the following components separately: jojoba oil, dimethicone, glycerin, 1,3-butanediol, 1,2-pentanediol, 1,2-hexanediol / Glycol, caprylic acid, oil soluble vitamin E acetate, acryloyldimethyltaurylamine/VP copolymer, xanthan gum, sodium phytate, cetearyl garate, sorbitan olive Oleic acid ester, disodium edetate, deionized water;

2) Mixture of phase B mixture, weigh the following components separately: hyaluronic acid, silicon elastomer, deionized water;

3) C-phase mixture configuration, weigh the following components separately: small nucleic acid, plant oil control factor, flavor, glycerin and deionized water;

4) Take the A-phase mixture and stir at 30-45 rpm and heat to 85 ° C, keep for 30 minutes, open the vacuum and stir to the maximum speed of 72 rpm, homogenize for 5 minutes, change the stirring speed to 30-45 rpm, cool down to 60 °C;

5) Add the B phase mixture, open the vacuum, stir at a maximum speed of 72 rpm, homogenize for 2 minutes, change the stirring speed to 30-45 rpm, and cool to 50 ° C;

6) Add the mixture of phase C, open the vacuum, stir at a maximum speed of 72 rpm, homogenize for 2 minutes, change the stirring speed to 30-45 rpm, continue cooling, change the stirring to low speed to 20-30 rpm after 3 minutes, and cool to 25-40 °C. , that is, get acne cosmetics.

The acne cosmetic of the present invention can be effectively used for the elimination and prevention of facial acne, skin oil control, moisturizing, nourishing and repair, and is safe and non-toxic; the acne medicine of the present invention can be effectively used for the treatment of facial acne.

DRAWINGS

Figure 1 is a view of Propionibacterium acnes observed under a microscope (magnification 1000 times) Gram stained photos.

Figure 2 is a photograph of a normal cultured Propionibacterium acnes (puncture anaerobic culture).

Figure 3 is a photo of small nucleic acid inhibition of P. acnes growth, in the figure "+" access strain; "-" no strain; a is untreated group (no small nucleic acid added); b is erythromycin; It is a small nucleic acid of 40 mg/mL; d is a small nucleic acid of 80 mg/mL; and e is a small nucleic acid of 120 mg/mL.

Figure 4 is a photo of small nucleic acid inhibition of P. acnes growth, "+" access strain; "-" no strain; a is untreated group (no small nucleic acid added); b is 80mg/mL small nucleic acid c is a small nucleic acid of 90 mg/mL; d is a small nucleic acid of 100 mg/mL; and e is a small nucleic acid of 110 mg/mL.

Figure 5 is a bar graph showing the growth of small nucleic acids inhibiting E. coli, wherein the ordinate is the OD ₆₀₀ relative absorbance and the abscissa is the different treatment groups.

Figure 6 is a photograph of the cup dish method for detecting the growth of small nucleic acids to inhibit E. coli growth.

Figure 7 is a gel electrophoresis diagram of S. cerevisiae M-dsRNA and Candida tropicalis M-dsRNA purified by RNase digestion, in which: M is RNA Marker; 1 is Candida tropicalis total RNA; 2 is RNase T1 Isolated Candida tropicalis small dsRNA; 3 is Saccharomyces cerevisiae total RNA; 4 is Saccharomyces cerevisiae small dsRNA isolated by RNase T1.

Figure 8 is a bar graph of real-time quantitative PCR detection of small nucleic acids affecting the expression of cytokines in sebaceous gland cells.

Fig. 9 is a photograph showing a trial effect I of a small nucleic acid acne cosmetic.

Fig. 10 is a photograph showing a trial effect II of a small nucleic acid acne cosmetic.

Fig. 11 is a photograph showing a trial effect III of a small nucleic acid acne cosmetic.

detailed description

The amounts, contents and concentrations of various substances are referred to herein, and the percentages thereof are all percentages by mass unless otherwise specified.

For convenience, hereinafter, the terms "double-stranded nucleic acid molecule", "small nucleic acid", "small nucleic acid molecule", "RNA", "RNA molecule", "double-stranded RNA", "double-stranded RNA molecule" or "dsRNA" "Can be interchanged, they mean the same meaning and scope. Among them, RNA is a double-stranded structure formed by annealing the sense strand and the antisense strand.

For the sake of convenience, the acne cosmetics containing small nucleic acid molecules of the present invention are simply referred to as "small nucleic acid acne cosmetics" or "small nucleic acid cosmetics". The acne drug containing a small nucleic acid molecule of the present invention is simply referred to as a "small nucleic acid acne drug" or a "small nucleic acid drug".

Studies have found that RNA derived from Saccharomyces cerevisiae and Candida tropicalis, especially small nucleic acid molecules, have acne activity. Among the total RNA of Saccharomyces cerevisiae and Candida tropicalis, the acne activity of small nucleic acid molecules ranging in length from 17 to 100 bp is particularly remarkable, and they are all double-stranded RNA molecules.

Both the small nucleic acid acne cosmetic and the acne drug of the present invention may comprise a small nucleic acid molecule such as Saccharomyces cerevisiae total RNA, Candida tropicalis total RNA, the first type of small nucleic acid molecule, or the second type of small nucleic acid. a molecule; may also comprise a mixture of two or more small nucleic acid molecules, namely, S. cerevisiae total RNA, Candida tropicalis total RNA, the first type of small nucleic acid molecule, and any two or more of the second type of small nucleic acid molecules. Mixtures in any ratio.

In the small nucleic acid acne cosmetic and acne medicine of the present invention, the lower limit of the weight percentage of the small nucleic acid is about 0.001%, preferably about 0.005%, 0.01%, 0.015%, 0.02%, 0.025%, 0.03%, 0.035%. 0.04%, 0.045%, or 0.05%; the upper limit of the weight percentage is about 0.5%, preferably about 0.48%, 0.45%, 0.42%, 0.40%, 0.38%, 0.35%, 0.32% or 0.3%. If the weight percentage of the small nucleic acid is less than about 0.001%, the acne effect is not fully manifested; if the weight percentage of the small nucleic acid is higher than about 0.5%, the product cost is too high, which may affect product sales.

It will be understood that the term "about" or "approximately" as used herein means that the indicated number may have an error range or floating range of ±10%.

Preferably, the small nucleic acid acne medicament of the present invention may further comprise other pharmaceutically active ingredients which have synergistic effects with the above small nucleic acid molecules in acne but do not affect the stability and physiological activity of small nucleic acids, such as vitamin A acid, Benzoyl peroxide, antibiotics (clindamycin, erythromycin, chloramphenicol, etc.), sebacic acid, sulfur, etc., the amount of these active ingredients should be limited to the extent that it does not cause skin irritation.

As a topical drug, the dosage form should be suitable for maintaining the stability of small nucleic acid molecules and promoting percutaneous absorption of small nucleic acid molecules, that is, skin penetration, and is preferably an aqueous dosage form such as an ointment, a cream, a cream, an expectorant, a liquid, A lotion, gel, film, or spray. The manner in which these dosage forms are prepared is well known to those skilled in the art. One of skill in the art can select suitable pharmaceutically acceptable topical excipients, including fillers and excipients, depending on the dosage form.

In the above-mentioned aqueous preparations, elixirs, creams, ointments, lotions, gels, and the like, the main ingredient to be used is not particularly limited as long as it is a pharmaceutically acceptable main ingredient, and examples thereof include a water-soluble main ingredient, Oily main agent, emulsion main agent. One or two or more such main agents can be used as needed.

The water-soluble main component may, for example, be polyethylene glycol (macrogol), ethanol, glycerin or a pharmaceutically acceptable glycol (for example, propylene glycol or butylene glycol). Among them, from the viewpoint of the solubility of the compound of the present invention is good Preferred are pharmaceutically acceptable glycols.

The oily main agent may, for example, be a petrolatum or a paraffin wax derived from minerals, a plastibase obtained by gelatinizing a polyethylene resin with liquid paraffin, or beeswax derived from living organisms.

As the emulsion main agent, for example, lanolin, stearyl alcohol or the like can be exemplified. In the case of using an emulsion main agent, it is preferred to further add an emulsifier. Examples of such an emulsifier include polyoxyethylene hardened castor oil and glyceryl monostearate.

Ointments, creams, creams, gels and lotions generally comprise a base such as white petrolatum, yellow petrolatum, lanolin, white beeswax, cetyl alcohol, stearyl alcohol, stearic acid, hydrogenated oil, coagulation Gelated hydrocarbons, polyethylene glycol, liquid paraffin, squalane, etc.; antioxidants, such as tocopherol derivatives, L-ascorbic acid, dibutylhydroxytoluene, butylated hydroxyanisole; preservatives, such as Hydroxybenzoate, etc.; humectants such as glycerin, propylene glycol, sodium hyaluronate, etc.; surfactants such as polyoxyethylene derivatives, fatty acid glycerides, sucrose fatty acid esters, sorbitan fatty acid esters, propylene glycol Fatty acid esters, lecithin, etc.; and thickeners such as carboxyvinyl polymer, xanthan gum, carboxymethylcellulose, sodium carboxymethylcellulose, hydroxypropylcellulose, hydroxypropylmethylcellulose Wait. Further, stabilizers, preservatives, absorption enhancers, pH adjusters, and other suitable additives may be added as needed.

The preparation in the form of a solution or emulsion may contain a solvent, emulsifier or demulsifier as a carrier material such as water, ethanol, isopropanol, ethyl carbonate, ethyl acetate, benzyl alcohol, benzyl benzoate, propylene glycol, 1,3 - butyl glycol oil, fatty acid glyceride, polyethylene glycol or sorbitan fatty acid ester, and the like.

The preparation in the form of a suspension may contain a liquid diluent such as water, ethanol or propylene glycol as a carrier material; suspensions such as ethoxylated isosteatrylalcohol, polyoxyethylene sorbitan ester and polyoxyethylene sorbitan ester; And microcrystalline cellulose, aluminummetahydrate, bentonite, agar, tragacanth, and the like.

The coating agent may include a film-forming agent such as an alkyl methacrylate copolymer, hydroxypropylcellulose, hydroxypropylmethylcellulose, methylcellulose, ethylcellulose, or polyvinyl alcohol. The coating agent provided by the present invention is a liquid preparation for external use in which a small nucleic acid is dissolved or dispersed in a solvent containing a film-forming material, and a film is formed after coating the affected area.

Further, the preparation in the form of the above solution, emulsion or suspension may also be attached to a polymer material to prepare a mask.

The spray may comprise a base used in the preparation of ointments, creams, creams, gels, suspensions, liquids and lotions, such as white petrolatum, yellow petrolatum, lanolin, white beeswax, whales. Wax alcohol, stearyl alcohol, stearic acid, hydrogenated oil, gelled hydrocarbon, polyethylene glycol, liquid paraffin, squalane, etc.; antioxidants, such as tocopherol derivatives, L- Ascorbic acid, dibutylhydroxytoluene, butylhydroxyanisole, etc.; preservatives such as parabens; humectants such as glycerin, propylene glycol, sodium hyaluronate, etc.; surfactants such as polyoxyethylene derived , fatty acid glycerides, sucrose fatty acid esters, sorbitan fatty acid esters, propylene glycol fatty acid esters, lecithin, etc.; thickeners, such as carboxyvinyl polymer, xanthan gum, carboxymethyl cellulose, carboxymethyl Cellulose sodium, hydroxypropyl cellulose, hydroxypropyl methyl cellulose, and the like. Different stabilizers, buffers, flavoring agents, suspending agents, emulsifying agents, fragrances, preservatives, solubilizing agents, and other suitable additives may be added. In particular, propellants such as chlorofluorocarbons, propane/butane or dimethyl ether may also be included.

In one embodiment, the small nucleic acid acne drug of the present invention is a cream, which is a preparation in which a small nucleic acid is dissolved in an emulsion-type matrix. The preparation method is carried out by a method conventional in the art, for example, as follows:

(1) Formulating the oil phase: Weigh the oily matrix component and heat it to 50 ° C ~ 100 ° C for melting, stir well, and keep warm;

(2) Preparing the aqueous phase: weigh the surfactant, the transdermal enhancer dissolved in the aqueous matrix, and heat to 50 ° C ~ 100 ° C, stir well, keep warm;

(3) slowly adding the above oil phase to the above aqueous phase, and stirring for 10 to 90 minutes;

(4) After cooling to 45 ° C ~ 60 ° C, add small nucleic acids, continue to stir;

(5) After the temperature is lowered to room temperature, a cream is obtained.

Wherein the oily matrix component is selected from the group consisting of petrolatum, paraffin, liquid paraffin, silicone oil, beeswax, stearic acid, lanolin, glyceryl monostearate or a combination of two or more thereof in any ratio; The aqueous matrix component is selected from any one of glycerin, propylene glycol, ethanol, water, or a combination of two or more thereof in any ratio; the surfactant is selected from the group consisting of glyceryl monostearate, triethanolamine, sodium lauryl sulfate, Any combination of polysorbate, fatty acid sorbitan, poloxamer, lecithin, gelatin or any combination of two or more; the transdermal enhancer is selected from the group consisting of lauric acid, polyethylene glycol, and mint Either of the alcohols or a combination of two or more of any ratios.

In another embodiment, the small nucleic acid acne drug of the present invention is an ointment, which is a homogeneous semi-solid external preparation prepared by mixing a small nucleic acid with an oily or water-soluble base. The preparation method is carried out by a method conventional in the art, for example, as follows:

(1) Weigh the oily matrix component and heat it to 50 ° C ~ 100 ° C to melt, stir well, keep warm;

(2) Adding a small nucleic acid and a transdermal enhancer to the above oily base, and stirring well.

Wherein the oily matrix component is selected from the group consisting of petrolatum, paraffin, liquid paraffin, silicone oil, beeswax, stearic acid, lanolin or any combination of two or more thereof; the transdermal enhancer is selected from the group consisting of Any one or more of a combination of ketone, polyethylene glycol, and menthol.

In another embodiment, the small nucleic acid acne drug of the present invention is a gel, which is made of small nucleic acids and auxiliary materials. A thick liquid or semi-solid preparation having gel properties. The preparation method is carried out by a method conventional in the art, for example, as follows:

(1) soaking the gel matrix component with a solvent for 24 hours to fully swell; adding triethanolamine under stirring to adjust the pH to 6.0-8.0 to form a transparent substrate for use;

(2) otherwise, the small nucleic acid is dissolved in the aqueous matrix component, added to the above gel matrix, and stirred;

(3) Add a transdermal enhancer, add the aqueous matrix component to a sufficient amount, and mix well.

Wherein the gel matrix component is selected from any one of carbomer, cellulose derivative, alginate, tragacanth, gelatin, starch, or a combination of two or more thereof in any ratio; the aqueous matrix The component is selected from any one of glycerin, propylene glycol, water, ethanol, or a combination of two or more of any ratio; the transdermal enhancer is selected from any one of lauricone, polyethylene glycol, and menthol. Or a combination of two or more in any ratio.

In another embodiment, the small nucleic acid acne drug of the present invention is a spray, a solution or emulsion or suspension of a small nucleic acid and a suitable excipient, filled in a special device, and sprayed in a mist when used. The preparation method is carried out by a method conventional in the art, for example, as follows:

(1) Weighing the small nucleic acid with an aqueous matrix component, and stirring;

(2) Adding a transdermal enhancer, adding the aqueous base component to a sufficient amount, stirring it, and filling it into a spray device.

Wherein the aqueous matrix component is selected from any one of glycerin, propylene glycol, water, ethanol or a combination of two or more of any ratio; the transdermal enhancer is selected from the group consisting of lauric acid, polyethylene glycol, and mint Either of the alcohols or a combination of two or more of any ratios.

The present invention will be further described in detail below in conjunction with specific embodiments. It is understood that the following examples are merely illustrative of the invention and are not intended to limit the scope of the invention.

Example

The preparation and sequencing of the S. cerevisiae total RNA, Candida tropicalis total RNA, the first type of small nucleic acid molecule and the first type of small nucleic acid molecule in the present invention are all completed by Biomax Biotech Co., Ltd. The following numbers are given in the examples:

1. Candida tropicalis total RNA, number BMX;

2. Saccharomyces cerevisiae total RNA, number BMX01;

3. One of the first class of small nucleic acid molecules, number BM06, the sequence is:

Justice strand: 5'-UAGGUACUUCGAGAAGCCUAGUAdTdT-3' (SEQ ID NO: 15),

Antisense strand: 5'-UACUAGGCUUCUCGAAGUACCUAdTdT-3' (SEQ ID NO: 16);

4. The second type of small nucleic acid molecule, number BM05, the sequence is:

Justice strand: 5'-UAGGUACUUCGAGAAGCCUAGUA-3' (SEQ ID NO: 5),

Antisense strand: 5'-UACUAGGCUUCUCGAAGUACCUA-3' (SEQ ID NO: 6);

5. One of the second type of small nucleic acid molecules, number BM01, the sequence is:

The sense strand: 5'-AGGUACUUCGAGAAGCCdTdT-3' (SEQ ID NO: 13),

Antisense strand: 5'-GGCUUCUCGAAGUACCUdTdT-3' (SEQ ID NO: 14);

6. The second type of small nucleic acid molecule, number BM02, the sequence is:

The sense strand: 5'-AGGUACUUCGAGAAGCC-3' (SEQ ID NO: 7),

Antisense strand: 5'-GGCUUCUCGAAGUACCU-3' (SEQ ID NO: 8).

Among them, BM05 is the backbone sequence of the first type of small nucleic acid molecule; BM02 is the backbone sequence of the second type of small nucleic acid molecule.

Example 1

1. Experimental materials and instruments:

Propionibacterium acnes (ATCC 6919): Shanghai Fuxiang Bio; Modified GAM Broth Medium: Shanghai Biochemistry; Hemin, Vitamin K1: Shanghai Guyan Bio; 37 °C Constant Temperature Incubator: Hundred Omega Bio; Ultra Clean Workbench: Suzhou Purification Equipment Factory.

2. Experimental grouping:

The concentration of small nucleic acid BMX was: 40mg/mL, 80mg/mL, 120mg/mL; positive control: using commercially available erythromycin ointment as positive control, the concentration was 40mg/mL; sterile control: no inoculation of acne propionate Bacillus medium; untreated control: medium inoculated with P. acnes, but without small nucleic acid treatment.

3. Experimental steps:

After the nutrient broth medium is configured according to the instructions, it is autoclaved at 115 ° C / 15 min. When it is cooled to 50 ° C, hemin 5 μg / mL and 1 μg / mL of vitamin K1 are added, and 4 mL is taken and transferred to a glass test tube. After cooling to room temperature with a sterile membrane seal, the small nucleic acid added to the above treatment group was mixed, and the medium was solidified and used.

After the dry powder of P. acnes is dissolved in sterile water, the cells are inoculated into the culture medium of the glass test tube by using the sterile inoculation needle, and after inoculation, about 1 mL of the liquid medium is covered on the top of the glass test tube medium. After cooling and solidification, it was placed in a 37 ° C incubator for 72 h and photographed.

4. Experimental results

1) Gram staining results shown in Figure 1 show that the growth pattern of bacteria conforms to P. acnes.

2) As shown in the culture results of Fig. 2, the anaerobic culture of P. acnes was grown in the medium along the puncture route.

3) As shown in the results in Figure 3, the positive control erythromycin completely inhibited the growth of P. acnes; when the small nucleic acid concentration was 80 mg/mL, it partially inhibited the growth of P. acnes; when the small nucleic acid concentration was 120 mg/mL , completely inhibited the growth of P. acnes.

The results indicate that the small nucleic acid molecule of the present invention has an activity of inhibiting P. acnes.

Example 2

1. Experimental materials and instruments:

Propionibacterium acnes (ATCC 6919): Shanghai Fuxiang Bio; Modified GAM Broth Medium: Shanghai Biochemistry; Hemin, Vitamin K1: Shanghai Guyan Bio; 37 °C Constant Temperature Incubator: Shanghai Xinmiao; ultra-clean workbench: Suzhou purification equipment factory.

2. Experimental grouping:

The concentration of small nucleic acid BMX was: 80mg/mL, 90mg/mL, 100mg/mL, 110mg/mL, 120mg/mL; positive control: using commercially available erythromycin ointment as a positive control, the concentration was 40mg/mL; sterile Control: medium in which P. acnes was not inoculated; untreated control: medium inoculated with P. acnes, but without small nucleic acid treatment.

3. Experimental steps:

The P. acnes dry powder is dissolved in sterile water, and then inoculated into the culture medium of the glass test tube by using the aseptically treated inoculation needle. After inoculation, the top of the glass test tube medium is covered with about 1 mL of the liquid medium. After cooling and solidifying, it was placed in a 37 ° C incubator for 72 h and photographed.

4. Experimental results

As shown in the results of Figure 4, 1) the antibacterial effect of the RNA molecule BMX is concentration-dependent, that is, the higher the concentration, the stronger the antibacterial effect. 2) The RNA molecule BMX concentration completely inhibited the growth of P. acnes when it was 110 mg/mL.

Example 3

1. Experimental materials and instruments:

Escherichia coli: Biomaike bio; LB liquid medium: Biomax bio; 37 °C constant temperature shaker: Shanghai Zhicheng; ultra-clean workbench: Suzhou purification equipment factory.

2. Experimental grouping:

The concentration of small nucleic acid BMX was: 40mg/mL, 80mg/mL, 120mg/mL; positive control: ampicillin as positive control, concentration was 100μg/mL; sterile control: medium without E. coli; untreated Control: medium inoculated with E. coli, but without the addition of small nucleic acids.

3. Experimental steps:

After LB liquid medium (1% (W/V) Tryptone (tryptone), 0.5% (W/V) Yeast (yeast extract), 1% (W/V) NaCl, pH=7.0), After autoclaving at 121 ° C / 20 min, 4 mL was transferred to a glass test tube, and after cooling to room temperature with a sterile membrane seal, the small nucleic acid added to the above treatment group was mixed and used.

The Escherichia coli strain was inoculated into a culture medium of a glass test tube, and cultured overnight at 37 ° C under a constant temperature shaker, and the OD ₆₀₀ absorbance was measured by an ultraviolet spectrophotometer at a wavelength of 600 nm, and the treatment results were counted.

4. Experimental results

As shown in Figure 5, 1) different concentrations of small nucleic acid BMX have the effect of inhibiting E. coli. 2) The growth of E. coli was completely inhibited when the small nucleic acid concentrations were 80 and 120 mg/mL.

Example 4

1. Experimental materials and instruments:

Escherichia coli: Biomaxco Biotechnology Co., Ltd.; LB solid medium: Biomax Biotech Co., Ltd.; 37 °C constant temperature shaker: Shanghai Zhicheng; ultra-clean workbench: Suzhou Purification Equipment Factory.

2. Experimental grouping:

3. Experimental steps:

Configure LB solid medium (1% (W/V) Tryptone (tryptone), 0.5% (W/V) Yeast (yeast extract), 1% (W/V) NaCl, 1.5% (W/V) Agar (Agar), pH=7.0), autoclave at 121 ° C / 20 min, inject 10-15 mL into a glass sterile plate, and allow it to naturally set before use. The E. coli strains were diluted and uniformly coated on the surface of the LB solid medium. After the bacteria liquid was absorbed, four Oxford cups were uniformly placed symmetrically on the surface of each medium. The above different concentrations of small nucleic acids were added to the Oxford Cup and placed in a 37 ° C incubator for 24 h. Observe the growth of the bacteria.

4. Experimental results

As shown in the results of Figure 6, 1) different concentrations of small nucleic acid BMX have the effect of inhibiting the growth of E. coli. 2) The growth of E. coli was completely inhibited when the small nucleic acid concentrations were 80 and 120 mg/mL.

Example 5

Yeast RNA isolation and identification

1. Total yeast RNA: Saccharomyces cerevisiae total RNA (Beaucome Biotechnology Co., Ltd.), Candida tropicalis total RNA (Beaucom Biotech Co., Ltd.).

2. Main reagents and materials: RNase T1 (Fermentas, USA), RNA Marker (RL1000, TaKaRa), RNase-free glycogen (Shanghai Ruiqi Biotechnology), and other biochemical reagents were purchased from Shanghai Biotech.

3, the main instruments: ultra-clean workbench (SW-CJ-2F Suzhou purification equipment factory); low temperature refrigerator (-80 ° C, DF-U4086S, Japan Sanyo); electrophoresis instrument (Bio-Rad, USA); gel imaging system (Biosens, USA); UV spectrophotometer and analytical workstation (Shimadzu Shimadazi, Japan) and so on.

4. The dsRNA was isolated by treating total yeast RNA with RNaseT1. The reaction was as follows: 200 μL of the solution contained 20 μL (20 μg) of RNA; 10 μL (10 U) of RNase T1; 20 μL of 10×TE Buffer (10 mM Tris-HCl, 1 mM EDTA, pH=8.0), 150 μL of sterile water. The reaction was carried out at 37 ° C for 30 min, 20 μL of EDTA (250 mM) was added to stop the reaction, and the enzyme was inactivated by heating at 72 ° C for 20 min. Take 5 μL of electrophoresis analysis. One μL of yeast dsRNA was added to 99 μL of 1×TE Buffer (100-fold dilution), and the ratio of OD ₂₆₀ and OD _{280 was} measured by ultraviolet spectrophotometer (this value was between dsRNA between 1.8 and 2.0).

5. Electrophoresis analysis of digestion products:

The isolated yeast dsRNA was detected by 15% polyacrylamide gel electrophoresis. As shown in Fig. 7, the isolated yeast small dsRNA was mainly distributed below 100 bp, and no DNA and other single-stranded RNA were observed. In the figure: M is RNA Marker; 1 is total RNA of Candida tropicalis; 2 is small Candida tropicalis dsRNA isolated by RNase T1; 3 is total RNA of Saccharomyces cerevisiae; 4 is Saccharomyces cerevisiae small dsRNA isolated by RNase T1.

6. Ultraviolet spectrophotometer detection: The ratio of OD ₂₆₀ to OD _{280 of} the isolated small Candida tropicalis and S. cerevisiae dsRNA was 1.91 and 1.89, respectively, which was confirmed to be dsRNA.

Example 6

Effects of small nucleic acids on TLR family and inflammatory factors after stimulating sebaceous gland cells

1. Experimental materials and instruments

Human sebaceous gland cells: Biomax Biotech Co., Ltd., Poly (I: C): Sigma-Aldrich, USA; small nucleic acid BM01, BM06, BMX: Biomax Biotech Co., Ltd.; 6-well cell culture plate: USA Corning Corporation; RISO ^TM RNA extraction reagent: one hundred Aomai Ke biotechnology Co., ^{Ltd; EzOmics TM One-Step qPCR Kit} : one hundred Aomai Ke biotechnology Co., Ltd; StepOnePlus ABI real-time PCR instrument (ABI, USA).

2. Experimental grouping:

Small nucleic acid treatment group: BM01, 50 nM; BM06, 50 nM; Poly (I: C), 10 ng/ml; BMX, 2.5 μg;

3. Cell treatment: 6-well plates were plated, and the cell confluence was about 50% before transfection, and the cells were treated with the above small nucleic acids.

4. Real time quantitative PCR (RT-qPCR) detection

After 48h cells were treated with RISO ^TM RNA extraction reagent according to the instructions extracted total cellular RNA, mRNA expression levels detected by RT-qPCR cells the gene of interest.

Gene-specific primers were used to detect the expression level of gene mRNA in the sample, and the housekeeping gene GAPDH was amplified as an internal reference control. Each sample simultaneously amplified the gene of interest and the internal reference gene GAPDH, and each reaction was made in 3 parallels. Quantitative reactions were performed using the One-Step qPCR kit to establish the following reaction system: 2 μL RNA template, 12.5 μL of 2×Master Mix, 5′ end primer (10 μM) and 3′ end primer (10 μM) each 0.5 μL, 0.5 μL of 50 ×SYBR Green I Solution, treated with DEPC to make up the water to 25 μL. After mixing well, the reaction was placed on a real-time quantitative PCR machine. The required primers were synthesized by Biomax Biotechnology Co., Ltd. The sequence is as follows:

IL4 detection primers:

Upstream primer: 5'-ATGAGAAGGACACTCGCTGC-3' (SEQ ID NO: 9);

Downstream primer: 5'-CCAACGTACTCTGGTTGGCT-3' (SEQ ID NO: 10).

GAPDH Primer:

Upstream primer: 5'-GAAGGTGAAGGTCGGAGTC-3' (SEQ ID NO: 11);

Downstream primer: 5'-GAAGATGGTGATGGGATTTC-3' (SEQ ID NO: 12).

Reaction conditions: reverse transcription at 42 ° C for 30 min, pre-denaturation at 95 ° C for 5 min, denaturation at 95 ° C for 20 sec, annealing at 58 ° C for 30 sec, extension at 72 ° C for 30 sec, and 45 cycles. The dissolution curve reaction was carried out: 95 ° C / 5 min, 58 ° C / 5 min, and the temperature was raised to 95 ° C at 0.5 ° C / 5 sec.

5, statistical processing

The data were analyzed by SPSS 14.0 statistical software. The measurement data were expressed by the mean ± SD. The difference significance test between the two groups was analyzed by one-way analysis of variance. The t test was used for comparison between the two groups. P<0.05 showed that the difference was statistically significant.

6, the experimental results

As shown in Figure 8, the expression level of the anti-inflammatory cytokine IL4 was significantly up-regulated by the action of small nucleic acids compared to the untreated group (P < 0.05).

Example 7

A small nucleic acid acne cosmetic comprising the following weight percentage components: jojoba oil 2%, dimethicone 1%, glycerol 7.5%, 1,3-butanediol 5%, 1,2- Pentyl glycol 2%, 1,2-hexanediol/octyl alcohol 0.5%, hyaluronic acid 0.02%, caprylic acid glyceride 0.3%, oil-soluble vitamin E acetate 0.2%, acryloyldimethyl taurate /VP copolymer 0.65%, xanthan gum 0.15%, sodium phytate 0.1%, cetearyl oleate / sorbitan olive oil 1%, plant oil control factor 2%, silicon elastomer 0.5%, Small nucleic acid (BMX) 0.25%, fragrance 0.15%, the balance is deionized water.

Example 8

A small nucleic acid acne cosmetic comprising the following weight percentage components: jojoba oil 2%, dimethicone 1%, glycerol 7.5%, 1,3-butanediol 5%, 1,2- Pentyl glycol 2%, 1,2-hexanediol/octyl alcohol 0.5%, hyaluronic acid 0.02%, caprylic acid glyceride 0.3%, oil-soluble vitamin E acetate 0.2%, acryloyldimethyl taurate /VP copolymer 0.65%, xanthan gum 0.15%, sodium phytate 0.1%, cetearyl oleate / sorbitan olive oil 1%, plant oil control factor 2%, silicon elastomer 0.5%, Small nucleic acid (BMX01) 0.5%, fragrance 0.15%, the balance is deionized water.

Example 9

A small nucleic acid acne cosmetic comprising the following weight percentage components: jojoba oil 2%, dimethicone 1%, glycerol 7.5%, 1,3-butanediol 5%, 1,2- Pentyl glycol 2%, 1,2-hexanediol/octyl alcohol 0.5%, hyaluronic acid 0.02%, caprylic acid glyceride 0.3%, oil-soluble vitamin E acetate 0.2%, acryloyldimethyl taurate /VP copolymer 0.65%, xanthan gum 0.15%, sodium phytate 0.1%, cetearyl oleate / sorbitan olive oil 1%, plant oil control factor 2%, silicon elastomer 0.5%, Small nucleic acid (BM01) 0.003%, fragrance 0.15%, the balance is deionized water.

Example 10

A small nucleic acid acne cosmetic comprising the following weight percentage components: jojoba oil 2%, dimethicone 1%, glycerol 7.5%, 1,3-butanediol 5%, 1,2- Pentyl glycol 2%, 1,2-hexanediol/octyl alcohol 0.5%, hyaluronic acid 0.02%, caprylic acid glyceride 0.3%, oil-soluble vitamin E acetate 0.2%, acryloyldimethyl taurate /VP copolymer 0.65%, xanthan gum 0.15%, sodium phytate 0.1%, cetearyl oleate / sorbitan olive oil 1%, plant oil control factor 2%, silicon elastomer 0.5%, Small nucleic acid (BM06) 0.001%, fragrance 0.15%, the balance is deionized water.

Example 11

A small nucleic acid acne cosmetic comprising the following weight percentage components: jojoba oil 2%, dimethicone 1%, glycerol 7.5%, 1,3-butanediol 5%, 1,2- Pentyl glycol 2%, 1,2-hexanediol/octyl alcohol 0.5%, hyaluronic acid 0.02%, caprylic acid glyceride 0.3%, oil-soluble vitamin E acetate 0.2%, acryloyldimethyl taurate /VP copolymer 0.65%, xanthan gum 0.15%, sodium phytate 0.1%, cetearyl oleate / sorbitan olive oil 1%, plant oil control factor 2%, silicon elastomer 0.5%, Small nucleic acid (BM06) 0.003%, fragrance 0.15%, the balance is deionized water.

Example 12

The preparation method of the small nucleic acid acne cosmetics of Examples 7-11 is as follows:

1) A phase mixture configuration, weigh the following components separately: jojoba oil 2%, dimethicone 1%, glycerol 7%, 1,3-butanediol 5%, 1,2-pentanediol 2 %, 1,2-hexanediol/octyl glycol 0.5%, caprylic acid glyceride 0.3%, oil-soluble vitamin E acetate 0.2%, acryloyldimethyltaurylamine/VP copolymer 0.65%, xanthan gum 0.15%, sodium phytate 0.1%, cetearyl oleate / sorbitan olive oil 1%, the balance is deionized water;

2) Mixture of phase B mixture, weigh the following components separately: hyaluronic acid 0.02%, deionized water 3%, silicon bomb body 0.5%;

3) Mixture of phase C mixture, weigh the following components separately: glycerin 0.5%, small nucleic acid 0.001-0.5%, deionized water 3%, plant oil control factor 2%, flavor 0.15%;

4) Take phase A and stir at 30-45 rpm and heat to 85 ° C, keep for 30 minutes, open vacuum and stir to maximum speed 72 rpm, homogenize for 5 minutes, change stirring speed to 30-45 rpm, cool down to 60 ° C ;

5) Add phase B, open vacuum, stir at a maximum speed of 72 rpm, homogenize for 2 minutes, change the stirring speed to 30-45 rpm, and cool to 50 ° C;

6) Add phase C, open vacuum, stir at maximum speed 72 rpm, homogenize for 2 minutes, change stirring speed to 30-45 rpm, continue cooling, change stirring at low speed to 20-30 rpm after 3 minutes, cool to 25-40 °C discharge , that is, a small nucleic acid acne cosmetics.

Example 13

The small nucleic acid acne cosmetics of Examples 7-11 of the present invention were tested, as follows:

1. Subjects: Men and women aged 15 to 33 years (mean age 22.9 years). There are different degrees of hemorrhoids on the face and neck. The causes are hormone secretion, endocrine disorders, taking drugs, improper use of cosmetics, etc.

2, the experimental method: evenly apply a small nucleic acid acne cosmetics on the affected area every morning and evening, or evenly apply the entire face or neck. Two weeks is an observation period.

3. Divide acne according to severity (according to the internationally accepted three-level four-level classification of acne):

1) I (mild): acne is the main damage, there may be a small amount of papules and pustules, the total number of lesions is less than 30;

2) II (moderate): there are acne, and there are a moderate number of papules and pustules, the total number of lesions is between 31-50;

3) III (sub-severity): There are a large number of papules and pustules, the total number of lesions is between 51-100, occasionally there is large inflammatory damage, less than 3 nodules;

4) IV (severe): mainly nodules, cysts or polymeric acne. The total number of lesions is more than 100, the number of lesions is more than 100, and the number of nodules or cysts is more than 3.

4, efficacy evaluation grading (refer to the Ministry of Health dermatological drugs clinical research guidelines): "-" (invalid), skin lesions subsided <20% or increased; "+" (effective), skin loss subsided > 20% ~ 60% "++" (significant effect), skin lesions subsided >60%; "+++" (healing), skin lesions subsided.

5, the treatment effect is as follows:

祛痘化妆品Acne cosmetics	例数Number of cases	痊愈get well	显效Significant effect	有效effective	无效invalid	总有效率Total efficiency
实施例7小核酸祛痘化妆品Example 7 Small nucleic acid acne cosmetics	3030	1212	88	66	44	86.67％86.67%
实施例8小核酸祛痘化妆品Example 8 small nucleic acid acne cosmetics	4747	1616	1717	55	99	80.85％80.85%
实施例9小核酸祛痘化妆品Example 9 small nucleic acid acne cosmetics	3131	55	99	1212	55	83.87％83.87%
实施例10小核酸祛痘化妆品Example 10 Small nucleic acid acne cosmetics	3030	33	1010	1212	55	83.33％83.33%
实施例11小核酸祛痘化妆品Example 11 Small nucleic acid acne cosmetics	3333	66	55	1919	33	90.91％90.91%

6. Figure 9-11 shows a representative photo of the patient's acne effect (compared before and after use). As can be seen from these photographs, after applying the acne cosmetics of the present invention for two weeks, the symptoms of the original acne site were alleviated, and the acne of the patient almost disappeared.

Example 14

A small nucleic acid acne medicine, which is a cream, consists of the following components: small nucleic acid BMX 1g, stearic acid 40g, glyceryl monostearate 40g, petrolatum 20g, liquid paraffin 70g, glycerol 50g, triethanolamine 3 g, 3 g of sodium lauryl sulfate, 2 g of azoxystrobin, and an appropriate amount of purified water, the total amount of 1000 g.

The method for preparing the small nucleic acid acne medicine is as follows:

(1) Oil phase: Accurately weigh stearic acid, glyceryl monostearate, petroleum jelly, liquid paraffin on a water bath, heat and melt, stir well, keep warm at around 75 °C;

(2) Aqueous phase: Accurately weigh glycerin, triethanolamine, lauryl ketone, sodium lauryl sulfate and water, stir and dissolve in a water bath, and heat to 75 °C.

(3) The above oil phase was slowly added to the aqueous phase, and after stirring for 15 minutes, stirring was maintained.

(4) Cool down to about 55 °C, add a small amount of BMX nucleic acid, and continue to stir.

(5) After the temperature is lowered to room temperature, a cream is obtained.

Example 15

A small nucleic acid acne drug, which is a cream consisting of the following components: small nucleic acid BM06 1 g, stearic acid 40 g, glyceryl monostearate 40 g, petrolatum 20 g, liquid paraffin 70 g, glycerol 50 g, triethanolamine 3 g, 3 g of sodium lauryl sulfate, 2 g of azoxystrobin, and an appropriate amount of purified water, the total amount of 1000 g.

The method for preparing the small nucleic acid acne medicine is as follows:

(4) Cool down to about 55 °C, add a small amount of nucleic acid BM06, and continue to stir.

(5) After the temperature is lowered to room temperature, a cream is obtained.

Example 16

A small nucleic acid acne medicine, which is a gelling agent, consists of the following components: small nucleic acid BM01 1g, carbomer 10g, propylene glycol 50g, glycerol 50g, triethanolamine 10ml, ethanol 20ml, laurel azide 5g, The amount of purified water is 1000 g in total.

The method for preparing the small nucleic acid acne medicine is as follows:

Add glycerin and propylene glycol to the carbomer to make it wet, add about 500ml of purified water, let stand for 24h to fully swell; add triethanolamine under stirring, adjust the pH to 6.5-7.5, form a transparent matrix for use; Weigh the small nucleic acid BM01, add it to the above gel, stir well; finally add laurel azide, add purified water to a sufficient amount, and stir well to obtain a gel.

Example 17

The trials were carried out using the small nucleic acid acne drugs of Examples 14-16 of the present invention, as follows:

1. Subjects: Men and women aged 18 to 33 years (mean age 23.5 years). There are different degrees of hemorrhoids on the face and neck. The causes are hormone secretion, endocrine disorders, taking drugs, improper use of cosmetics, etc.

5, the treatment effect is as follows:

祛痘药物Acne drug	例数Number of cases	痊愈get well	显效Significant effect	有效effective	无效invalid	总有效率Total efficiency
实施例14小核酸祛痘药物Example 14 Small nucleic acid acne drug	2525	1111	99	22	33	88.00％88.00%
实施例15小核酸祛痘药物Example 15 Small nucleic acid acne drug	1919	99	77	11	22	89.47％89.47%
实施例16小核酸祛痘药物Example 16 Small nucleic acid acne drug	2020	88	88	11	33	85.00％85.00%

In summary, the acne cosmetics and acne drugs of the present invention can be effectively used for the removal of facial acne, and are safe and non-toxic, and have broad commercial prospects.

Claims

An acne composition consisting of the following components by weight: 0.001-0.5% of a small nucleic acid molecule, 99.50-99.999% of a cosmetic excipient or a topical excipient, wherein the small nucleic acid molecule is selected from the total RNA of Saccharomyces cerevisiae, tropical silk Yeast total RNA, one or a mixture of two or more of the following two types of double-stranded RNA molecules, consisting of a sense strand and an antisense strand, respectively:

The first type of small nucleic acid molecule,

Justice chain: 5'-UAGGUACUUCGAGAAGCCUAGUANn-3' (SEQ ID NO: 1),

Antisense strand: 5'-UACUAGGCUUCUCGAAGUACCUANn-3' (SEQ ID NO: 2);

The second type of small nucleic acid molecule,

The sense strand: 5'-AGGUACUUCGAGAAGCCNn-3' (SEQ ID NO: 3),

Antisense strand: 5'-GGCUUCUCGAAGUACCUNn-3' (SEQ ID NO: 4),

Wherein N is cytidine C, guanosine G, adenosine A, uridine U, deoxycytidine dC, deoxyguanosine dG, deoxyadenosine dA or deoxygenation Thymidine dT; n represents the number of N, and n is an integer of 0-2.
The acne composition according to claim 1, wherein the first type of small nucleic acid molecule consists of the following sense and antisense strands:

Justice strand: 5'-UAGGUACUUCGAGAAGCCUAGUA-3' (SEQ ID NO: 5),

Antisense strand: 5'-UACUAGGCUUCUCGAAGUACCUA-3' (SEQ ID NO: 6); or

Justice strand: 5'-UAGGUACUUCGAGAAGCCUAGUAdTdT-3' (SEQ ID NO: 15),

Antisense strand: 5'-UACUAGGCUUCUCGAAGUACCUAdTdT-3' (SEQ ID NO: 16); and the second class of small nucleic acid molecules consists of the following sense and antisense strands:

The sense strand: 5'-AGGUACUUCGAGAAGCC-3' (SEQ ID NO: 7),

Antisense strand: 5'-GGCUUCUCGAAGUACCU-3' (SEQ ID NO: 8); or

The sense strand: 5'-AGGUACUUCGAGAAGCCdTdT-3' (SEQ ID NO: 13);

Antisense strand: 5'-GGCUUCUCGAAGUACCUdTdT-3' (SEQ ID NO: 14).
The acne composition according to claim 1 or 2, wherein the acne composition is an acne cosmetic, and the cosmetic auxiliaries and weight percentage are: base fat 2.5-15%, moisturizer 1-20 %, anti Oxidizer 0.01-1.0%, thickener 0.1-2.0%, emulsifier 0.1-3%, plant oil control factor 1-5%, skin feel improver 0.1-2%, metal chelator 0-0.5%, flavor 0.1-0.5 %, the rest is water; or

The acne composition is an acne drug, and the acne drug dosage form is an ointment, a cream, a cream, an expectorant, a water, a lotion, a gel, a film, or a spray. The exogenous adjuvant is selected from the group consisting of pharmaceutically acceptable excipients for maintaining the stability of small nucleic acid molecules, skin permeation enhancers for promoting percutaneous absorption of small nucleic acid molecules, pharmaceutically acceptable fillers and excipients for the preparation of external pharmaceutical forms. ,water.
The acne composition according to claim 3, wherein the base fat is jojoba oil, dimethicone, or a mixture thereof.
The acne composition according to claim 3, wherein the humectant is glycerin, 1,3-butanediol, 1,2-pentanediol, 1,2-hexanediol/octanol , hyaluronic acid, caprylic acid glyceride, or a mixture of two or more thereof.
The acne composition according to claim 3, wherein the antioxidant is oil-soluble vitamin E acetate, sodium phytate, or a mixture thereof.
The acne composition according to claim 3, wherein the thickener is acryloyldimethyltaurylamine/VP copolymer, xanthan gum, or a mixture thereof.
The acne composition according to claim 3, wherein the emulsifier is cetearyl garate, sorbitan oligoate, or a mixture thereof;

The plant oil control factor is a horse chestnut seed extract, a sage leaf extract, a buckwheat (sage) extract, a coltsfoot leaf extract, a rosemary leaf extract, a mother chrysanthemum (European chrysanthemum) extract. , fragrant bee flower (melissa) leaf extract, or a mixture of two or more thereof;

The skin feel improving agent is a silicon elastomer;

The metal chelating agent is disodium edetate.
The acne composition according to claim 3, wherein the cosmetic is composed of the following components by weight: hojoba oil 1-10%, dimethicone 1-5%, glycerin 2-10%, 1 , 3-butanediol 1-10%, 1,2-pentanediol 0.1-5%, 1,2-hexanediol/octylene glycol 0.1-2%, hyaluronic acid 0.01-5%, caprylic glyceride 0.1-1%, Oil-soluble vitamin E acetate 0.1-0.5%, acryloyldimethyltaurylamine/VP copolymer 0.1-1.0%, xanthan gum 0.1-1%, sodium phytate 0.01-0.5%, cetearyl alcohol Olive oil ester / sorbitan olive oil 0.1-3%, plant oil control factor 1-5%, silica elastomer 0.1-2%, disodium edetate 0-0.5%, small nucleic acid 0.001-0.5% The fragrance is 0.1-0.5%, and the balance is deionized water.
A method of preparing the acne cosmetic of claim 3, comprising the steps of:

1) A phase mixture configuration, weigh the following components separately: jojoba oil, dimethicone, glycerin, 1,3-butanediol, 1,2-pentanediol, 1,2-hexanediol / Glycol, caprylic acid, oil soluble vitamin E acetate, acryloyldimethyltaurylamine/VP copolymer, xanthan gum, sodium phytate, cetearyl garate, sorbitan olive Oleic acid ester, disodium edetate and deionized water;

2) Mixture of phase B mixture, weigh the following components separately: hyaluronic acid, silicon elastomer, deionized water;

3) C-phase mixture configuration, weigh the following components separately: small nucleic acid, plant oil control factor, flavor, glycerin and deionized water;

4) Take the A-phase mixture and stir at 30-45 rpm and heat to 85 ° C, keep for 30 minutes, open the vacuum and stir to the maximum speed of 72 rpm, homogenize for 5 minutes, change the stirring speed to 30-45 rpm, cool down to 60 °C;

5) Add the B phase mixture, open the vacuum, stir at a maximum speed of 72 rpm, homogenize for 2 minutes, change the stirring speed to 30-45 rpm, and cool to 50 ° C;

6) Add the mixture of phase C, open the vacuum, stir at a maximum speed of 72 rpm, homogenize for 2 minutes, change the stirring speed to 30-45 rpm, continue cooling, change the stirring to low speed to 20-30 rpm after 3 minutes, and cool to 25-40 °C. , that is, get acne cosmetics.