CN103446030A - External skin preparation and applications thereof - Google Patents
External skin preparation and applications thereof Download PDFInfo
- Publication number
- CN103446030A CN103446030A CN2012101722311A CN201210172231A CN103446030A CN 103446030 A CN103446030 A CN 103446030A CN 2012101722311 A CN2012101722311 A CN 2012101722311A CN 201210172231 A CN201210172231 A CN 201210172231A CN 103446030 A CN103446030 A CN 103446030A
- Authority
- CN
- China
- Prior art keywords
- preparation
- venenum apis
- external preparation
- epidermis
- polypeptide
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Pending
Links
Images
Landscapes
- Medicines That Contain Protein Lipid Enzymes And Other Medicines (AREA)
- Cosmetics (AREA)
Abstract
The invention relates to an external skin preparation and applications thereof, and specifically provides an external preparation, which adopts bee venom polypeptide as the active component. The preparation is capable of inhibiting proliferation of epidermal cells, inducing apoptosis of epidermal cells, and promoting formation of granular layer epidermis, and therefore the preparation can regulate, control and cure skin diseases caused by epidermis proliferation and/or prosoplasia.
Description
Technical field
The invention belongs to field of medicaments, be specifically related to a kind of external skin preparation and application thereof.
Background technology
Epidermis is the first protective barrier of human body, and its chief component cell is epithelial cells (keratinocyte, claims again keratinocyte, sometimes also general term epidermis cell).In order to complete the barrier function of epidermis, epidermis cell must carry out complexity and be subject to the atomization of tight regulation and control, and final formation comprises basal layer, spinous layer, granular layer, clear layer and cuticular multilayer structure.
Wherein, granular layer (stratum granulosum) is positioned at the shallow-layer of prickle cell layer, the layer 2-3 cell, consists of, and its thickness changes with the thickness of cuticular layer.Granular layer extremely often cause epidermis proliferative disorder and dysdifferentiation, thereby cause multiple epidermis propagation or the relevant dermatopathy of disdifferentiation.
Epidermal growth regulation and control can cause various diseases extremely, and psoriasis is one of common skin diseases wherein, and its cause of disease just relates to the unbalance of epidermis cell homeostasis mechanism.
Psoriasis (psoriasis, PS) is a kind of erythroderma desquamativum of common chronic recurrent.Psoriasis is to take epidermal keratinocytes hyper-proliferative and differentiation is complete, granular layer minimizing or disappearance, high dermis Marjoram Extract the keratinization of epidermis dermatoses that is pathological characters with inflammatory reaction.The cytobiology phenomenon of a series of Growth of Cells disorders appears in silver bits skin lesion, except relevant with inducing of cell growth factor, and also closely related with the dysregulation of the apoptosis of epidermis cell and atomization.
Still lack at present satisfied Therapeutic Method, be difficult to radical cure, easily recurrence.Also there is no at present the psoriatic medicine of a kind of highly effective treatment.
Venenum apis is a kind of mixture of complicated component, and it also contains several protein and peptide class, enzyme, histamine, acids, aminoacid and trace element etc. except containing large quantity of moisture.Research and to apply maximum be the Venenum apis polypeptide in Venenum apis, the Venenum apis polypeptide accounts for the Venenum apis dry weight more than 50%, is major function material in Venenum apis.In prior art, the clinical practice of Venenum apis mainly contains following several: (1) connective tissue disease, as rheumatic and rheumatoid arthritis; (2) neuritis and neuralgia, as treatment sciatica, trigeminal neuralgia, occipital neuralgia, dorsal nerve root inflammation etc.; (3) cardiovascular disease, as hypertension of atherosclerosis disease, venous thrombosis, thromboangiitis obliterans and endarteritis; (4) allergic disease, as bronchial asthma.
Also without any technology or research, disclosed and Venenum apis or its main component can be applied to treat psoriasis at present.
Summary of the invention
One of purpose of the present invention is to provide a kind of new purposes of Venenum apis polypeptide.
Another object of the present invention is to provide a kind of external preparation and method for making and purposes that the Venenum apis polypeptide is active component of take.
In first aspect present invention, a kind of purposes of Venenum apis polypeptide is provided, it is for the preparation of the compositions of the formation of (a) regulation and control granular layer of epidermis; (b) compositions of fibroblast and/or melanocyte hypertrophy in regulation and control skin; And/or (c) compositions of reduce wrinkle or whitening.
In another preference, described compositions is for the preparation of the compositions of the formation of regulation and control granular layer of epidermis.
In another preference, described compositions is for the preparation of the compositions of fibroblast and/or melanocyte hypertrophy in regulation and control skin.
In another preference, described compositions is for the preparation of the compositions of reduce wrinkle or whitening.
In another preference, described compositions comprises pharmaceutical composition and cosmetics.
In another preference, described cosmetics comprise: facial film, vanishing cream, crease-proof cream, fair complexion cream, freckle-removing cream, emulsifiable paste, eye sticker, emulsion, cosmetic water, soap, medicated soap, cleansing milk, perfume, body lotion or shampoo.
In another preference, described compositions is to be used for the treatment of psoriatic pharmaceutical composition.
In another preference, the formation of described regulation and control granular layer of epidermis is the formation that promotes granular layer of epidermis.
In another preference, described compositions also for: suppress epidermis cell propagation, promote apoptosis in epidermal cell or the abnormal relevant dermatopathy for the treatment of epidermal hyperplasia.
In another preference, described epidermis cell is normal epidermis cell.
In another preference, described fibroblast and melanocyte are normal cells.
In another preference, the external preparation that described compositions is percutaneous or transdermal.
In another preference, described Venenum apis polypeptide comprises: Venenum apis Copeptin (Melittin), Venenum apis gelatin (Apamin), mastocyte take off cell peptide (MCDP), spend in comfort and even up (Adolapin) or its combination.
In second aspect present invention, a kind of external preparation is provided, it comprises: (a) Venenum apis polypeptide; And (b) pharmaceutically acceptable carrier or excipient.
In another preference, in described preparation, the content of Venenum apis polypeptide is 0.05 μ g-500mg/ gram, is preferably 0.1 μ g-100mg/ gram, is more preferably 1-1000 μ g/ gram, 10-100 μ g/ gram best.
In another preference, described external preparation is cosmetic formulations or pharmaceutical preparation; Or
Described external preparation is unguentum, cream, patch, varnish, spray or micropin agent.
In another preference, described pharmaceutically acceptable carrier or excipient are to be selected from one or more of lower group: water, oils and fats, high molecular polymer and moisturizing isostearyl glyceryl pentaerythrityl ether; And/or
Described oils and fats is to be selected from one or more of lower group: Vegetable oil lipoprotein, animal oil and wax, mineral grease and wax and synthetic ester oil and wax; And/or
Described high molecular polymer is natural polymers or artificial-synthetic copolymer.
In another preference, described Vegetable oil lipoprotein is selected from lower group: lauric acid oils, Palmic acid oils, oleic acid oils, linoleic acid oils, linolenic acid oils, erucic acid oils, conjugate acid oils, hydroxy acid oils, or its combination.
In another preference, described animal oil and wax are selected from lower group: mink oil, shark liver oil, Adeps Bovis seu Bubali etc.; Animal wax comprises spermaceti, Cera Flava, lanoline or its combination.
In another preference, described mineral grease and wax are selected from lower group: liquid paraffin, vaseline, paraffin, microwax etc.; Described synthetic ester oil and wax comprise various lanolin derivatives, squalane, polysiloxanes or its combination.
In another preference, described high molecular polymer is selected from lower group: collagen protein, hyaluronic acid, cellulosic polymer, polyvinyl alcohol and derivant thereof, Polyethylene Glycol, polyethylene glycol oxide, polyvinylpyrrolidone, acrylic acid and derivant thereof or its combination.
In another preference, described moisturizing isostearyl glyceryl pentaerythrityl ether is selected from lower group: fatty acid ester, polyol fatty acid ester, triglyceride, fatty acid, fatty alcohol and fatty alcohol-polyoxyethylene ether, silicone oil, wax ester, hydrocarbon ils and chloroflo, phospholipid or its combination.
In another preference, described external preparation has following one or more effects:
(i) formation or the treatment psoriasis of regulation and control (as promoted) granular layer of epidermis;
(ii) hypertrophy of fibroblast or melanocyte in regulation and control (as suppressed) skin;
(iii) reduce wrinkle or whitening;
(iv) suppress epidermal cell proliferation, promote apoptosis in epidermal cell or the abnormal relevant dermatopathy for the treatment of epidermal hyperplasia.
In third aspect present invention, a kind of method for making as the described external preparation of second aspect present invention is provided, comprise step: mix (a) Venenum apis polypeptide and (b) pharmaceutically acceptable carrier or excipient, thereby make the described external preparation of second aspect present invention.
In fourth aspect present invention, a kind of Therapeutic Method is provided, comprise step: the object that the Venenum apis polypeptide is applied to the needs treatment.
In another preference, described object is mammal, comprises the people.
In another preference, described method reaches following one or more effects:
(i) the formation treatment psoriasis of regulation and control (as promoted) granular layer of epidermis;
(ii) hypertrophy of fibroblast or melanocyte in regulation and control (as suppressed) skin;
(iii) reduce wrinkle or whitening;
(iv) suppress epidermal cell proliferation, promote apoptosis in epidermal cell or the abnormal relevant dermatopathy for the treatment of epidermal hyperplasia.
In another preference, described method reaches following effect: suppress other histiocyte propagation or promote other histiocytic apoptosis, described other histiocytes are mesenchymal cell or chondrocyte.
In another preference, during treatment, the activity of Venenum apis polypeptide is 1-1000 μ g/mL, is preferably 10-200 μ g/mL.
In should be understood that within the scope of the present invention, above-mentioned each technical characterictic of the present invention and can combining mutually between specifically described each technical characterictic in below (eg embodiment), thus form new or preferred technical scheme.As space is limited, tire out and state no longer one by one at this.
The accompanying drawing explanation
Fig. 1 has shown the impact of Melittin on epidermal cell proliferation.
Fig. 2 has shown the impact of Venenum apis polypeptide on epidermal cell proliferation.
Fig. 3 has shown the impact of Melittin on apoptosis in epidermal cell.
Fig. 4 has shown the impact of Venenum apis polypeptide on apoptosis in epidermal cell.
Fig. 5 has shown the impact that Melittin forms Mus tail epithelium granular layer.
Fig. 6 has shown the impact that the Venenum apis polypeptide forms Mus tail epithelium granular layer.
Fig. 7 has shown the pigmentation inhibitory action to guinea epidermis containing Venenum apis polypeptide preparation for external application to skin.
The specific embodiment
The inventor is by long-term and deep research, be surprised to find that the Venenum apis polypeptide not only can suppress the propagation of epidermis cell and the apoptosis of promotion epidermis cell, effectively participate in epidermis cell Equilibrium mechanism, can also promote granular layer of epidermis to form, antagonism cuticular layer keratinization not exclusively, prickle cell layer thickening, epidermal thickness increase, thereby can regulate and control and treat epidermis propagation and/or the relevant dermatopathy of disdifferentiation.Test shows, the Venenum apis polypeptide to psoriasis occur, development has effective intervention effect.On this basis, the inventor has completed the present invention.
The Venenum apis polypeptide
Venenum apis (bee venom) is become to be grouped into by number of chemical, and wherein the polypeptide class mainly comprises Venenum apis Copeptin (Melittin), Venenum apis gelatin (Apamin), the de-cell peptide (MCDP) of mastocyte, spends in comfort and even up (Adolapin) etc.; Enzyme comprises hyaluronidase (Hyaluronidase), phospholipase A2 (Phospholipase A2) etc.; Amine (as histamine, dopamine etc.) and other materials.
The Venenum apis polypeptide is the several polypeptides matter contained in Venenum apis, is most important functional mass in Venenum apis.In the Venenum apis polypeptide, that wherein content is maximum is Melittin, accounts for the 40-50% of Venenum apis polypeptide total amount; Except Melittin, other main polypeptide compositions also comprise: MCDP accounts for 2-3%; Apamin accounts for 2-3%; Adolapin accounts for 0.5-1%, also comprises some micro-polypeptide (as Secapine, Pamine, Minimine, Procamine, Tertiapine etc.).
Wherein, Melittin is a kind of typical cationic polypeptide (cationic antimicrobial peptide, CAP), by 26 amino acid residues, formed, molecular weight is 2840D, the primary structure sequence be GIGAVLKVLTTGLPALISWIKRKRQQ (as shown in SEQ ID NO.:1, No. CAS: 20449-79-0).Secondary structure is mainly amphipathic (amphiphilic) alpha-helix.Melittin has very high biological activity, and research shows that it has the multiple effects such as antiinflammatory, analgesia, inhibition platelet aggregation and antitumor.
Research has found that Melittin can suppress the propagation of multiple cultured tumor cells in vitro or apoptosis-induced (as Toxicol Appl Pharmacol.2012:p72; J Clin Invest.2009:p2830; Life Sci.2007:p364), relevant Melittin is to normal somatic cell, and particularly the research of epidermal keratinocytes propagation and Apoptosis has no report.
External preparation
" active component " of the present invention is the Venenum apis polypeptide.Active component of the present invention has the multiple beneficial effect, for example, the formation of regulation and control granular layer of epidermis (forming as promoted granular layer of epidermis), the treatment psoriasis, the propagation of inhibition epidermis cell, promote apoptosis in epidermal cell, the abnormal relevant dermatopathy for the treatment of epidermal hyperplasia, fibroblastic hypertrophy in regulation and control (as promoted) skin, the hypertrophy of melanocyte in regulation and control (as suppressed) skin, or the effects such as reduce wrinkle and whitening.The pharmaceutical composition (as external preparation) that can be used for the abnormal relevant disease of preparation treatment skin hyperplasia.
External preparation of the present invention comprises acceptable excipient or carrier on the interior Venenum apis polypeptide of safe and effective weight range and pharmacology.Wherein " safe and effective amount " refers to: the amount of active component is enough to obviously improve the state of an illness, and is unlikely to produce serious side effect.Usually, external preparation contains 0.05 μ g-500mg Venenum apis polypeptide/agent, more preferably, contains 0.1 μ g-100mg Venenum apis polypeptide/agent.Preferably, described " potion " is a patch.Preferably, when described external preparation is liquid preparation, in described external preparation, the concentration of Venenum apis polypeptide is 0.05 μ g-500mg/mL, preferably 0.1 μ g-100mg/mL, more preferably 1-1000 μ g/mL.When described external preparation is solid preparation, in described external preparation, the content of Venenum apis polypeptide is 0.05 μ g-500mg/ gram, preferably 0.1 μ g-100mg/ gram, more preferably 1-1000 μ g/ gram.
" pharmaceutically acceptable carrier " refers to: one or more compatibility solids or liquid filler or gelatinous mass, they are suitable for the people uses, and enough purity and enough low toxicity must be arranged." compatibility " referred to herein as each component energy and active component of the present invention and blending mutually between them in preparation, and the drug effect of not obvious reduction active component.Pharmaceutically acceptable carrier part example has cellulose and derivant (as sodium carboxymethyl cellulose, ethyl cellulose sodium, cellulose ethanoate etc.) thereof, gelatin, Talcum, kollag (as stearic acid, magnesium stearate), calcium sulfate, vegetable oil (as Oleum Glycines, Oleum sesami, Oleum Arachidis hypogaeae semen, Fructus Canarii albi wet goods), polyhydric alcohol (as propylene glycol, glycerol, mannitol, sorbitol etc.), emulsifying agent
wetting agent (as sodium lauryl sulphate), coloring agent, flavoring agent, stabilizing agent, antioxidant, antiseptic, apirogen water etc.
Pharmaceutically acceptable carrier of the present invention or excipient are preferably from one or more of lower group: water, oils and fats, high molecular polymer and moisturizing isostearyl glyceryl pentaerythrityl ether.
Wherein, described oils and fats includes, but is not limited to: Vegetable oil lipoprotein, animal oil and wax, mineral grease and wax and synthetic ester oil and wax.Described Vegetable oil lipoprotein includes, but is not limited to: lauric acid oils, Palmic acid oils, oleic acid oils, linoleic acid oils, linolenic acid oils, erucic acid oils, conjugate acid oils, hydroxy acid oils, or its combination.Described animal oil and wax include, but is not limited to: mink oil, shark liver oil, Adeps Bovis seu Bubali etc.; Animal wax comprises spermaceti, Cera Flava, lanoline or its combination.Described mineral grease and wax include, but is not limited to: liquid paraffin, vaseline, paraffin, microwax etc.; Described synthetic ester oil and wax include, but is not limited to: various lanolin derivatives, squalane, polysiloxanes or its combination.
Wherein, described high molecular polymer can be natural polymers or artificial-synthetic copolymer, for example includes, but is not limited to: collagen protein, hyaluronic acid, cellulosic polymer, polyvinyl alcohol and derivant thereof, Polyethylene Glycol, polyethylene glycol oxide, polyvinylpyrrolidone, acrylic acid and derivant thereof or its combination.
Wherein, described moisturizing isostearyl glyceryl pentaerythrityl ether can be preferably the moisturizing isostearyl glyceryl pentaerythrityl ether from lower group: fatty acid ester, polyol fatty acid ester, triglyceride, fatty acid, fatty alcohol and fatty alcohol-polyoxyethylene ether, silicone oil, wax ester, hydrocarbon ils and chloroflo, phospholipid or its combination.
The method of application of Venenum apis polypeptide or external preparation is not particularly limited, and representational method of application comprises that (but being not limited to) is for topical, transdermal administration and percutaneous dosing.The dosage form that is used for the representative Venenum apis polypeptide of above-mentioned administering mode comprises unguentum, cream, patch, varnish, spray, micropin agent etc.Active component under aseptic condition with physiologically acceptable carrier and any antiseptic, buffer agent, or the propellant that may need in case of necessity is mixed together.
The Venenum apis polypeptide can be individually dosed, or with other pharmaceutically acceptable medication combined administrations.
While using external preparation, the Venenum apis polypeptide of safe and effective amount to be applicable to need the mammal (as the people) for the treatment of, the effective dosage of dosage for pharmaceutically thinking while wherein using, for the people of 60kg body weight, day dosage is generally 0.05～1000mg, preferably 1～500mg.Certainly, concrete dosage also should be considered the factors such as route of administration, patient health situation, and these are all within the skilled practitioners skill.
Major advantage of the present invention:
1. in test, confirm in vitro for the first time: the Venenum apis polypeptide of finite concentration content suppresses epidermal cell proliferation, induces the apoptosis of hyper-proliferative epidermis cell.
2. in vivo test is found: the external preparation containing finite concentration Venenum apis polypeptide can promote granular layer of epidermis to form, can resist the cuticular layer keratinization not exclusively, prickle cell layer thickening, epidermal thickness increase.
3. find first that the Venenum apis polypeptide has effective intervention effect to psoriasis generation, development.
4. the external skin preparation containing the Venenum apis polypeptide has the effect that reduces wrinkle.
5. the external preparation containing the Venenum apis polypeptide has the pigmentation that inhibition UVB causes.
Visible, the Venenum apis polypeptide can be intervened propagation, the differentiation and apoptosis of epidermis cell, effectively participates in epidermis cell Equilibrium mechanism, regulates and controls and treat the dermatopathy of relevant epidermis propagation and/or disdifferentiation; And have and reduce wrinkle, white-skinned face function.
Below in conjunction with concrete enforcement, further set forth the present invention.Should be understood that these embodiment only are not used in and limit the scope of the invention for the present invention is described.The experimental technique of unreceipted actual conditions in the following example, usually according to normal condition, such as people such as Sambrook, molecular cloning: laboratory manual (New York:Cold Spring Harbor Laboratory Press, 1989) condition described in, or the condition of advising according to manufacturer.Unless otherwise indicated, otherwise percentage ratio and umber calculate by weight.Agents useful for same of the present invention or instrument unless otherwise noted, equal commercially available obtaining.
The Melittin (its sequence is with SEQ ID NO.:1) used in the present invention's experiment and Venenum apis (bee venom) are purchased from Sigma company; the Venenum apis polypeptide adopts existing method (Banks BE; Dempsey CE; Pearce FL; Vernon CA, Wholley TE.New methods of isolating been venom peptides.Anal Biochem.1981; 116 (1): the classical way separation of 48-52) reporting makes, the method mainly adopts chromatography method, separating the Venenum apis polypeptide obtained is a kind of mixture, and its composition is as consistent as document report, mainly contains Melittin, MCDP, Apamin, Adolapin etc.
The In vitro culture of embodiment 1. epidermis cell
Material and instrument
Trypsin is purchased from (GIBCO, USA);
0.53mM EDTA is purchased from (GIBCO, USA);
Pancreatin inhibitor is purchased from (GIBCO, USA);
Trypan blue is purchased from (Sigma, USA).
Experimental procedure:
1) draw materials
The Chinese Fetal Foreskin Fibroblasts of excision, insert containing in antibiotic keratinocyte culture fluid, brings cell culture chamber into.
2) separate epidermis
Carefully cut off subcutaneus adipose tissue and part corium with shears, and be trimmed to the strip of wide about 1-2mm, use without calcium magnesium PBS rinsing twice, epidermis side is dipped in 0.24U/ml neutral protease (Dispase II, Sigma, USA) 4 ℃ upward and spends the night.
3) digestion
After 18 hours, with two ophthalmology tweezers, epidermis is torn from corium, be cut into fragment, then digest 15 minutes at 37 ℃ with 0.05% trypsin-0.53mM EDTA, 10ml pancreatin inhibitor (10mg/ml) stops digestion.Through 150 order stainless steel filtering nets, filter, with 1000r/min centrifugal 10 minutes, supernatant discarded, added keratinocyte culture fluid 10ml, mixes Trypan Blue counting, cell viability>85%.
4) former culture
Make single cell suspension after counting, by 3 * 10
6/ 75cm
2the density inoculated and cultured, serum-free keratinocyte culture medium (GIBCO, USA), at 37 ℃, 5%CO
2, in the incubator of 100% relative humidity, to cultivate, culture fluid changes once in 3 days.
5) cultivation of going down to posterity
When cell fusion arrives 60%-75% density, with 10ml, without calcium magnesium PBS, rinse, digest 5-10 minute through 0.05% pancreatin-37 ℃ of 0.53mM EDTA, add the 10ml pancreatin inhibitor to stop digestion, in the immigration centrifuge tube, centrifugal 10 minutes of 1000r/min.Abandoning supernatant, add keratinocyte culture fluid 10ml, mixes, and counting, in the cultivation of going down to posterity of the ratio of 1:3.
Change culture fluid 2-3 day one time, until cell fusion is during to 60%-75% density, then the cultivation of going down to posterity.
The impact of embodiment 2.Melittin on epidermal cell proliferation
Material and instrument
The K-SFM culture fluid is purchased from GIBCO, USA;
([3-(4,5-dimethylthiazole-2-yl)-5-(3-carboxylic carbomethoxy)-2-(4-sulfophenyl)-2H-tetrazolium inner salt], purchased from Promega for MTS;
Automatically microplate reader is purchased from BioRad, USA.
Experimental technique:
(1) epidermal keratinocytes (Epidermal Keratinocytes, EK) that was cultured to for the 3rd generation according to the described method of embodiment 1 is selected in experiment,
(2) 4000, every hole cell is inoculated in 96 orifice plates, 37 ℃, 5%CO
2, in the K-SFM culture fluid, cultivate 12 hours.
After (3) 12 hours, culture fluid is changed in every hole, again adds the culture fluid that contains variable concentrations Melittin, and the ultimate density of Melittin in culture fluid is respectively 1,5,10,20,50,200 μ g/mL.Continue to hatch 48 hours.
After (4) 48 hours, every hole adds 20 μ l MTS, then continues to hatch 2 hours.
(5) measure the absorbance (Absorbance) in every hole by automatic microplate reader.
Experimental result:
Each organizes absorbance in Table 1 and Fig. 1.From result, can find out, each group all has inhibitory action in various degree, when the Melittin final concentration is 10 μ g/mL, to epidermis cell inhibited proliferation obvious (P<0.05); When the Melittin final concentration is 20,50,200 μ g/mL, to the inhibited proliferation of epidermal keratinocytes with matched group comparing difference highly significant (p<0.01).
The impact of table 1Melittin on epidermal cell proliferation
| Concentration | The absorbance average | Standard deviation |
| 1μg/ml | 0.9924 | 0.02312 |
| 5μg/ml | 0.9729 | 0.02701 |
| 10μg/ml | 0.9495 | 0.03564 |
| 20μg/ml | 0.9327 | 0.02438 |
| 50μg/ml | 0.9131 | 0.01673 |
| 200μg/ml | 0.8944 | 0.01480 |
The impact of embodiment 3. Venenum apis polypeptide on epidermal cell proliferation
This tests concrete experimental technique referring to embodiment 2, and only embodiment 2-a-(3) step, substitute Melittin with the Venenum apis polypeptide, and wherein the Venenum apis polypeptide makes by conventional method.Venenum apis polypeptide final concentration still is respectively 1,5,10,20,50,200 μ g/mL.
Result is as described in Fig. 2 and table 2.From result, can find out, each group all has inhibitory action in various degree, when Venenum apis polypeptide final concentration is 20 μ g/mL, to epidermis cell inhibited proliferation obvious (P<0.05); When Venenum apis polypeptide final concentration is 50,200 μ g/mL, to the inhibited proliferation of epidermal keratinocytes with matched group comparing difference highly significant (p<0.01).
The impact of table 2 Venenum apis polypeptide on epidermal cell proliferation
| Concentration | The absorbance average | Standard deviation |
| 1μg/ml | 0.9934 | 0.03708 |
| 5μg/ml | 0.9829 | 0.02522 |
| 10μg/ml | 0.9620 | 0.03978 |
| 20μg/ml | 0.9571 | 0.02475 |
| 50μg/ml | 0.9157 | 0.01828 |
| 200μg/ml | 0.9084 | 0.02161 |
The impact of embodiment 4.Melittin on apoptosis in epidermal cell
Material and instrument
RNase(Sigma,USA)；
Propidium iodide (PI) (Sigma, USA);
Flow cytometer: Beckman Coulter-Epics Altra, USA.
Experimental technique:
The epidermal keratinocytes in the 3rd generation of the Melittin Processing Example 2 experimental technique steps (a) that are 10,20,50,100 μ g/mL with final concentration, the cell do not processed with same generation is (CTRL) as a control group.After 48h, collect and respectively organize cell, adjust cell concentration to 1 * 10
7/ ml, respectively get 100 μ l; The binding buffer liquid 100 μ l that add pre-cooling, re-suspended cell, after 4 ℃, 24h, add RNase 0.1g/L, and 37 ℃ of digestion 30min, add propidium iodide (PI) 50mg/L, 4 ℃ of dyeing 1h, nylon net filter, flow cytometer is analyzed.
B. result:
Result, as shown in Fig. 3 and table 3, is compared with matched group, and when the Melittin final concentration is 10,20 μ g/mL, the apoptosis rate of cell rises to some extent, but it is not obvious to rise; Final concentration is that the 3rd of 50,100 μ g/mL Melittin processing represent that cornu cutaneum matter forms apoptosis rate and obviously rises, and the effect of 100 μ g/mL processed group is (p<0.01) especially significantly.
The impact of table 3Melittin on apoptosis in epidermal cell
| Concentration | The apoptosis rate average | Standard deviation |
| 0μg/ml(C TRL) | 3.66 | 0.5221 |
| 10μg/ml | 3.8883 | 0.9112 |
| 20μg/ml | 3.8683 | 1.0277 |
| 50μg/ml | 4.5766 | 0.7000 |
| 100μg/ml | 8.9733 | 0.5288 |
The impact of embodiment 5. Venenum apis polypeptide on apoptosis in epidermal cell
This tests concrete experimental technique referring to embodiment 4, with the Venenum apis polypeptide, substitutes Melittin.
Venenum apis polypeptide final concentration still is respectively 10,20,50,100 μ g/mL.The cell do not processed with same generation is (CTRL) as a control group.
Result is as described in table 4 or Fig. 4.
The impact of table 4 Venenum apis polypeptide on apoptosis in epidermal cell
| Concentration | The apoptosis rate average | Standard deviation |
| 0μg/ml(C TRL) | 3.66 | 0.5221 |
| 10μg/ml | 3.6861 | 0.7380 |
| 20μg/ml | 3.7198 | 0.8712 |
| 50μg/ml | 4.4570 | 0.8144 |
| 100μg/ml | 5.2041 | 1.1433 |
Result shows, respectively organizes inhibitory action is in various degree all arranged, and when Venenum apis polypeptide final concentration is 10,20 μ g/mL, the apoptosis rate of cell rises to some extent, but rises not obvious (P<0.05); When Venenum apis polypeptide final concentration is 50,100 μ g/mL, the apoptosis rate of cell is with matched group comparing difference highly significant (p<0.05).
Embodiment 6. is containing the preparation of Melittin external skin preparation
Raw material: eugenol ester 20g, glyceryl monostearate 70g, stearic acid 100g, glycerol 85g, white vaseline 85g, sodium lauryl sulphate 10g, para hydroxybenzene tetracid ethyl ester 1g, Melittin 50mg, distilled water 1000ml.
Method for making: glyceryl monostearate, stearic acid, the white vaseline of getting above-mentioned weight, be placed in the vessel in heating fusing, keep 80 ℃, the sodium lauryl sulphate, glycerol, the Melittin that separately get above-mentioned weight add in the 1000ml distilled water, be heated to 80 ℃, add the para hydroxybenzene tetracid ethyl ester of above-mentioned weight, slowly add above-mentioned oil phase after dissolving, be stirred to by same direction the fine and smooth paste that is white in color, condensation and get final product.
Embodiment 7. is containing the preparation of Venenum apis polypeptide external skin preparation
Raw material: eugenol ester 20g, glyceryl monostearate 70g, stearic acid 100g, glycerol 85g, white vaseline 85g, sodium lauryl sulphate 10g, para hydroxybenzene tetracid ethyl ester 1g, Venenum apis polypeptide 50mg, distilled water 1000ml.
Method for making: glyceryl monostearate, stearic acid, the white vaseline of getting above-mentioned weight, be placed in the vessel in heating fusing, keep 80 ℃, sodium lauryl sulphate, glycerol, the Venenum apis polypeptide of separately getting above-mentioned weight add in the 1000ml distilled water, be heated to 80 ℃, add the para hydroxybenzene tetracid ethyl ester of above-mentioned weight, slowly add above-mentioned oil phase after dissolving, be stirred to by same direction the fine and smooth paste that is white in color, condensation and get final product.
Percentage ratio by accounting for the formula gross weight, take or measure following each composition.
Get 5% polyacrylic acid resin, 1% sodium carboxymethylcellulose pyce, 0.03% dihydroxyaluminum aminoacetate and 0.02%EDTA, be scattered in above composition successively in 20% glycerol and mix, and obtains A liquid.
Getting 0.05% polyvinylpyrrolidone, to be dispersed in 50% deionized water for stirring even, then add 0.01%Melittin to continue to stir; 3% carbomer is layered on liquid level uniformly, and standing swelling 12-24 hour, stir standby, 1% polyvinyl alcohol put into to the deionized water heating for dissolving of surplus, cooling after, with the former, mix homogeneously, obtain B liquid.
By pre-configured A liquid, under the rotating speed stirring of 15-60rpm, add in B liquid and mix, form gel preparation, regulating pH value with triethanolamine or NaOH is 6.5, stirring and evenly mixing, can obtain gel preparation.
In embodiment 6～8, carrier used or excipient can at random be substituted by the carrier described in the present invention or excipient, thereby for the preparation of external preparation of the present invention.
The impact that embodiment 9.Melittin forms Mus tail epithelium granular layer
This experiment method therefor (as HE dyeing etc.) is the laboratory conventional method.
Experimental model: the too fast and parakeratosis of epidermal cell proliferation is psoriatic basic pathology characteristics, the epidermis of mouse tail scale lacks the formation of granular layer, similar to people's psoriasis epidermis pathological characters, be usually used in simulating psoriasis epidermis cell differentiation obstacle, the parakeratosis pharmacodynamic study with the curing psoriasis medicine of being correlated with.Medicine or preparation are if promotion Mus tail granular layer forms, and just prompting has function of resisting psoriasis.
Experimental technique:
Choose 30 experimental model mices (purchased from Chinese Academy of Sciences's Shanghai Experimental Animal Center) and be divided at random following 3 groups, 10 every group:
1. treatment group (Melittin): the Melittin external skin preparation made with embodiment 6 is processed;
2. matched group (W/O): with not processing containing Melittin external skin preparation, described not containing the method for making of Melittin external skin preparation with embodiment 6, difference is to lack Melittin;
3. blank group (NC), be left intact.
Respectively above three groups are processed: be applied in respectively the Mus tail bit of experimental model mice with each group preparation, (5mg/ time) once a day, coating is 14 days continuously.After the last coating, put to death animal next day, in distance Mus root of the tail section, 1.5 centimeters are cut off the Mus tail, avoid damage, cut Mus tail skin, peel off cartilage, getting length is about on 1 centimetre of left and right rectangle skin graft attaching and filter paper, drop into immediately in 10% formalin and fix, conventional dehydration, paraffin embedding, section, HE dyes and observe one by one the tail scale of every mice under light microscopic.
The granular layer cell person that every scale has connection to embark on journey, be called the scale that granular layer is arranged.Calculate the scale number that granular layer is arranged in every 100 scales, by treatment group and W/O and the contrast of blank group, and carry out statistical analysis, SAS 8.2 softwares commonly used carry out statistical disposition, there is the scale number of granular layer to compare each group, belong to single factor and organize the mean relative method more.
Experimental result:
As table 5 and Fig. 5 demonstration, Melittin external skin preparation containing certain concentration is formed with significant facilitation to mouse tail scale granular layer of epidermis, with W/O-, containing Melittin external skin preparation group and blank group difference, significance (p<0.05) is not arranged.The described external preparation of prompting this patent can obviously promote the formation of granular layer of epidermis, is very beneficial for regulating the differentiation of epidermis and maintaining of Selfstabilizing state, the function of resisting psoriasis excellence.
Table 5 respectively organize scale number that granular layer is arranged in 100 scales of Mus tail epithelium (mean ± standard deviation, n=10)
| Group | The scale number that granular layer is arranged in 100 scales |
| The NC group | 4.9±0.56 |
| The W/O group | 4.1±1.72 |
| The Melittin group | 11.7±0.94* |
Annotate: * P<0.05
The impact that embodiment 10. Venenum apis polypeptide form Mus tail epithelium granular layer
The Mus tail epithelium granular layer that repeats embodiment 9 forms the impact test, different, in treatment group (Venenum apis polypeptide), adopts the external skin preparation containing Melittin containing 6 preparations of Venenum apis polypeptide external skin preparation alternate embodiment of embodiment 7 preparations.
Result is as shown in table 6 and Fig. 6, external skin preparation containing certain concentration Venenum apis polypeptide is formed with to mouse tail scale granular layer of epidermis the facilitation shown, and with W/O-, containing Venenum apis external skin preparation group and blank group difference, significance (p<0.05) is not arranged.The Venenum apis external preparation that the prompting this patent is mentioned can be regulated the differentiation of epidermis and maintaining of Selfstabilizing state.
Table 6 respectively organize scale number that granular layer is arranged in 100 scales of Mus tail epithelium (mean ± standard deviation, n=10)
| Group | The scale number that granular layer is arranged in 100 scales |
| The NC group | 4.8±0.63 |
| The W/O group | 5.2±1.03 |
| Venenum apis polypeptide group | 9.8±1.31* |
Annotate: * P<0.05
Embodiment 11. is containing the experiment of Venenum apis polypeptide external skin preparation reduce wrinkle
The external skin preparation for treating containing the Venenum apis polypeptide that the present embodiment adopts embodiment 7 to prepare is wrinkle near the eyes.
Experimental subject:
Enter altogether 20 female volunteers of group, age 27-36 year.Enter group person's face, wrinkle is in various degree particularly arranged near the eyes; Enter group person and do not use any external, oral or injection reduce wrinkle or anti-aging product in nearest 30 days; Enter group person healthy, without any infectious disease.
Experimental technique:
Adopt face to smear mode, will evenly be coated with and put on the skin in face containing the external skin preparation of Venenum apis polypeptide, every day is respectively once (5mg/ time) sooner or later, continuous 14 days.Can use other the daily skin-protection products except declaring the products such as antidotal facial film, cream during entering group.
Entering to organize the first day doctor uses Hongkong JoinFree company skin analysis all-in-one (JF2000) to carry out skin near the eyes to take pictures, after finishing the 14th day course for the treatment of under same background, light, position, again take pictures, and utilize all-in-one to carry out the measuring and calculating of local wrinkle.
1. in conjunction with main, objective satisfaction, comprehensive assessment wrinkle improvement effect.
Carry out the classification of satisfaction according to local wrinkle improvement degree: the unchanged person of local wrinkle is for very dissatisfied; Local wrinkle improves 0%～25% for dissatisfied; It is general improving 25%～50%; Improve 50%～75% for satisfied; Improve 75% for very satisfied.
Continuous administration contained the external skin preparation of Venenum apis polypeptide after 14 days, and volunteer's subjective assessment satisfaction is 90%, only had two volunteer's satisfactions for general, not dissatisfied.It is 100% that the enforcement doctor assesses satisfaction.
2. skin analysis all-in-one results of measuring is as shown in table 7, and test value has represented the order of severity of test section wrinkle, and the larger expression wrinkle of numerical value problem is more serious.Result show into the group volunteer near the eyes wrinkle have clear improvement.
Table 7 skin analysis machine wrinkle test result
| Experimental subject | First day | The 14th day | Experimental subject | First day | The |
| 1 | 4.12 | 3.71 | 11 | 4.18 | 3.46 |
| 2 | 4.29 | 2.45 | 12 | 4.66 | 3.72 |
| 3 | 3.26 | 3.12 | 13 | 4.22 | 3.73 |
| 4 | 3.15 | 3.56 | 14 | 3.43 | 3.12 |
| 5 | 2.98 | 2.18 | 15 | 4.51 | 3.69 |
| 6 | 3.74 | 2.56 | 16 | 3.87 | 2.83 |
| 7 | 3.16 | 2.96 | 17 | 3.55 | 3.41 |
| 8 | 4.68 | 3.89 | 18 | 3.63 | 2.97 |
| 9 | 4.69 | 4.12 | 19 | 3.45 | 2.66 |
| 10 | 4.25 | 3.95 | 20 | 4.46 | 3.81 |
Embodiment 12. is containing the experiment of Venenum apis polypeptide external skin preparation pigmentation inhibitory action
Cavia porcellus pigmentation inhibition test is to estimate the classical way of external preparation white-skinned face function.
Experimental subject: select 10 of healthy male brown color Cavia porcelluss, body weight 320-390g, average 346.5g.
In the selected 4.0cm of guinea pig back * 4.0cm zone, after depilation, with SS-03 ultraviolet (UVB) therapeutic instrument (Xigema High Technology Co., Ltd., Shanghai), irradiate.Dosage is that 16.5mJ/d irradiates 10 days, and the irradiation total amount is 165mJ.
Experimental technique: after irradiating end, skin area by guinea pig back through irradiating is divided into 4 mutual zones that isolate, and is coated with 3% Melanex (positive control) outside difference, contains Melittin external skin preparation group (embodiment 6), contains Venenum apis polypeptide external skin preparation group (embodiment 7), does not contain Venenum apis polypeptide/Melittin external skin preparation matched group (W/O).Every day 2 times (each 5mg), amount to 3 weeks, biopsy after 3 weeks, and and do not irradiate between normal skin, each group mutually relatively to be estimated.
Histological observation: adopt DOPA dyeing.The permanent cold cut sheet of flesh tissue low temperature in 0.1% DOPA dye liquor 37 ℃ hatch 2h, understain haematoxylin, eosin dye liquor after washing.Active melanocyte cell dyeing is brownish black.
DOPA positive cell counting: under high power microscope, (Japan, Nikon company, * 100) choose 10 visuals field at random, counting epidermis dopa-positive melanocytes number.Adopt the SPSS statistical software to do date processing and analysis (Fig. 7).
Result shows: the pigmentation that the external skin preparation prepared according to this patent embodiment 6 and embodiment 7 methods causes UVB has remarkable inhibitory action (with W/O group ratio, P<0.05).The Pigmented inhibition caused with classic treatment preparation 3% hydroquinone comparison UVB does not have significant difference (P > 0.05).
Show that the external preparation containing Melittin or Venenum apis polypeptide of the present invention can suppress the pigmentation that UVB causes, and points out this external preparation to have white-skinned face function.
Above-mentioned having experimental results show that:
1. the external preparation containing the Venenum apis polypeptide of the present invention all can suppress epidermal cell proliferation effectively, induce the apoptosis of hyper-proliferative epidermis cell, can also obviously promote granular layer of epidermis to form, effectively participate in epidermis cell Equilibrium mechanism, thereby, for regulation and control or treatment epidermis propagation and/or the relevant dermatopathy of disdifferentiation, especially there is excellent function of resisting psoriasis.
2. the external preparation containing the Venenum apis polypeptide of the present invention has the effect that reduces wrinkle.
3. the external preparation containing the Venenum apis polypeptide of the present invention has the pigmentation that inhibition UVB causes, and the external skin preparation that prompting the present invention mentions has white-skinned face function.
All documents of mentioning in the present invention are all quoted as a reference in this application, just as each piece of document quoted separately as a reference.Should be understood that in addition those skilled in the art can make various changes or modifications the present invention after having read above-mentioned teachings of the present invention, these equivalent form of values fall within the application's appended claims limited range equally.
Claims (10)
1. the purposes of a Venenum apis polypeptide, is characterized in that, regulates and controls the compositions of the formation of granular layer of epidermis for the preparation of (a); (b) compositions of fibroblast and/or melanocyte hypertrophy in regulation and control skin; And/or (c) compositions of reduce wrinkle or whitening.
2. purposes as claimed in claim 1, is characterized in that, described compositions comprises pharmaceutical composition and cosmetics.
3. purposes as claimed in claim 1, is characterized in that, described compositions is to be used for the treatment of psoriatic pharmaceutical composition.
4. purposes as claimed in claim 1, is characterized in that, described compositions also for: suppress epidermis cell propagation, promote apoptosis in epidermal cell or the abnormal relevant dermatopathy for the treatment of epidermal hyperplasia.
5. purposes as claimed in claim 1, is characterized in that, the external preparation that described compositions is percutaneous or transdermal.
6. an external preparation, is characterized in that, comprises:
(a) Venenum apis polypeptide; And
(b) pharmaceutically acceptable carrier or excipient.
7. external preparation as claimed in claim 6, is characterized in that, described external preparation is cosmetic formulations or pharmaceutical preparation; Or
Described external preparation is unguentum, cream, patch, varnish, spray or micropin agent.
8. external preparation as claimed in claim 6, is characterized in that, described pharmaceutically acceptable carrier or excipient are to be selected from one or more of lower group: water, oils and fats, high molecular polymer and moisturizing isostearyl glyceryl pentaerythrityl ether; And/or
Described oils and fats is to be selected from one or more of lower group: Vegetable oil lipoprotein, animal oil and wax, mineral grease and wax and synthetic ester oil and wax; And/or
Described high molecular polymer is natural polymers or artificial-synthetic copolymer.
9. external preparation as claimed in claim 6, it is characterized in that, described moisturizing isostearyl glyceryl pentaerythrityl ether is selected from lower group: fatty acid ester, polyol fatty acid ester, triglyceride, fatty acid, fatty alcohol and fatty alcohol-polyoxyethylene ether, silicone oil, wax ester, hydrocarbon ils and chloroflo, phospholipid or its combination.
10. the method for making of an external preparation as claimed in claim 6, is characterized in that, comprises step: mix (a) Venenum apis polypeptide and (b) pharmaceutically acceptable carrier or excipient, thereby make external preparation claimed in claim 6.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN2012101722311A CN103446030A (en) | 2012-05-29 | 2012-05-29 | External skin preparation and applications thereof |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN2012101722311A CN103446030A (en) | 2012-05-29 | 2012-05-29 | External skin preparation and applications thereof |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| CN103446030A true CN103446030A (en) | 2013-12-18 |
Family
ID=49729152
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN2012101722311A Pending CN103446030A (en) | 2012-05-29 | 2012-05-29 | External skin preparation and applications thereof |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN103446030A (en) |
Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN109692327A (en) * | 2017-10-23 | 2019-04-30 | 华中科技大学 | A kind of application of the nano particle of the application and delivery melittin of melittin |
Citations (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1088215A (en) * | 1992-12-14 | 1994-06-22 | 刘尽晖 | The method of sharp separation honeybee peptide from bee venom |
| US20040081702A1 (en) * | 2000-04-14 | 2004-04-29 | Kim Christopher M. | Bee venom treatment without the sting |
| KR20100118629A (en) * | 2009-04-29 | 2010-11-08 | (주)아모레퍼시픽 | Cosmetic composition containing bee venom or extracts of the same |
| KR20110082335A (en) * | 2010-01-11 | 2011-07-19 | 대한민국(관리부서:농촌진흥청장) | Composition for whitenings and treating skin damage or disease comprising bee venom |
-
2012
- 2012-05-29 CN CN2012101722311A patent/CN103446030A/en active Pending
Patent Citations (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1088215A (en) * | 1992-12-14 | 1994-06-22 | 刘尽晖 | The method of sharp separation honeybee peptide from bee venom |
| US20040081702A1 (en) * | 2000-04-14 | 2004-04-29 | Kim Christopher M. | Bee venom treatment without the sting |
| KR20100118629A (en) * | 2009-04-29 | 2010-11-08 | (주)아모레퍼시픽 | Cosmetic composition containing bee venom or extracts of the same |
| KR20110082335A (en) * | 2010-01-11 | 2011-07-19 | 대한민국(관리부서:농촌진흥청장) | Composition for whitenings and treating skin damage or disease comprising bee venom |
Non-Patent Citations (5)
| Title |
|---|
| 作者未知: "蜂毒祛皱系列化妆品即将面世 可增强肌肤弹性延缓衰老", 《新浪新闻中心（HTTP://NEWS.SINA.COM.CN/W/2011-11-28/090923536206.SHTML）》, 28 November 2011 (2011-11-28), pages 1 - 2 * |
| 吴松青等: "胶原蛋白对人皮肤成纤维细胞生长状态的影响", 《求医问药》, vol. 9, no. 12, 31 December 2011 (2011-12-31), pages 187 - 188 * |
| 李万瑶主编: "《蜂毒疗法》", 31 January 2002, article "蜂毒软膏外用 银屑病", pages: 57 - 196-198 * |
| 杜武俊等: "蜂毒抑制癌细胞与肿瘤之作用", 《海峡两岸第四届蜜蜂生物学研讨会论文集》, 30 November 2004 (2004-11-30), pages 119 * |
| 马百芳等: "蜂毒注射液治疗寻常型银屑病", 《中华皮肤科杂志》, vol. 29, no. 2, 30 April 1996 (1996-04-30), pages 125 * |
Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN109692327A (en) * | 2017-10-23 | 2019-04-30 | 华中科技大学 | A kind of application of the nano particle of the application and delivery melittin of melittin |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| CN105106028B (en) | A kind of peptide composition for hair growth | |
| KR101323487B1 (en) | Antiwrinkle agent | |
| BRPI0619967A2 (en) | skin care compositions | |
| CN108379110B (en) | Skin care composition, skin care cream and preparation method thereof | |
| CN109568169A (en) | A kind of active peptides of tool hair growth effect | |
| US20220331389A1 (en) | Skin composition | |
| AU2015261352B2 (en) | Extemporaneous cosmetic and/or dermatological preparations | |
| CN109310598A (en) | Educate a mao agent | |
| WO2023109289A1 (en) | Composition containing plant fruit oil and plant seed oil, and use thereof | |
| CN114224787A (en) | Composition containing vegetable seed oil and fat and application thereof | |
| CN110302091A (en) | A kind of anti aging effect composition of niacinamide-containing adenine-dinucleotide and its preparation method and application | |
| CN105640793B (en) | A kind of composition being used to prepare face cream, the face cream and preparation method thereof comprising the composition | |
| KR20150010691A (en) | Hair loss preventing, skin and scalp protecting herbal cosmetic composition containing extracts of saxifrage, coptis, houttuynia, ginseng, peony root, paeonia suffruticosa, yeast and pleuropterus multiflorus, and its manufacturing method | |
| CN114146027A (en) | Composition containing microbial oil and vegetable oil and application thereof | |
| CN106413681A (en) | Ceramide-formulated external agent composition | |
| CN107115220A (en) | A kind of activity of fighting against senium compositions and preparation method and application | |
| CN103446030A (en) | External skin preparation and applications thereof | |
| US20070286915A1 (en) | Nerve Growth Factor Production Inhibitor and External Preparation for the Skin, Cosmetic, Quasi Drug, Preventive and Remedy for Atopic Dermatitis Containing the Nerve Growth Factor Production Inhibitor | |
| KR100490212B1 (en) | Anti-aging cosmetics composition containing amino acid complex | |
| TWI740964B (en) | Use of promoting adipocyte differentiation and method for screening materials for promoting adipocyte differentiation | |
| JP2931083B2 (en) | Skin cosmetics | |
| JP2014214094A (en) | Cosmetic composition containing liposome including culture solution extract of stem cells derived from human | |
| US11744791B2 (en) | Cosmetic composition comprising amide-based compound | |
| RU2119790C1 (en) | Agent for biological skin rejuvenation | |
| KR101373714B1 (en) | Cosmetic composition promoting cornified envelope formation |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| C06 | Publication | ||
| PB01 | Publication | ||
| C10 | Entry into substantive examination | ||
| SE01 | Entry into force of request for substantive examination | ||
| RJ01 | Rejection of invention patent application after publication |
Application publication date: 20131218 |
|
| RJ01 | Rejection of invention patent application after publication |