CN107519033A - A kind of anti-aging skin care product composition and application thereof - Google Patents

Abstract

The present invention relates to a kind of anti-aging skin care product composition and application thereof, specifically, it is related to a kind of anti-aging skin care product containing small molecule inhereditary material, especially, further relate to a kind of screening technique of anti-aging small molecule inhereditary material mark and the purposes of anti-aging small molecule inhereditary material, anti-aging skin care product composition provided by the invention is strong to the penetration of epidermis, expression effect is strong, and the natural safety of raw material, no dependence, have no toxic side effect.

Description

A kind of anti-aging skin care product composition and application thereof

Technical field

The present invention relates to a kind of anti-aging skin care product composition, a kind of particularly anti-aging skin care product composition and anti- The purposes of aging small-molecule substance.

Background technology

Epidermis is made up of multilayer epithelial cell, and protective barrier effect is played to environment and stimulation.It is mainly formed by cutin Cell forms, then these cells start to break up, and move to surface from the deep layer of epidermis in the bottommost layer hyperplasia of epidermis.Table Skin be both base layer interior occur hyperplasia place, or intermediate layer cell differentiation place, ultimately form one The protection structure that kind is broken up completely, i.e. cuticula.Completing the circulation probably needs 20 days, and the process corresponds to a variety of specific genes Succeedingly it is activated with the mode of coordination in a certain order, then repressed expression.

With the development of biology, particularly Protocols in Molecular Biology, brand-new hair is brought to cosmetics industry Opportunity is opened up, skin care item develop from fine chemistry industry beauty, plant beauty to biological beauty, gene beauty.Biological skin care item Main component biologically active polypeptide is largely Porcine HGF, and content is atomic in vivo for they, but bioactivity is high, right Various kinds of cell physiological function and metabolic activity play biological regulation and acted on, growth that is direct, affecting indirectly polytype cell, Division, break up, breed and migrate, in beauty and skin care, plastic surgery, burn ulcer and various dermopathic wound reparations with being cured Played an important role in conjunction.

The research and development of most domestic anti-aging beauty-care skin care item fail to set about from cell ageing mechanism, but are carried with some plants Thing or Chinese medicinal compound extract are taken, as Xu Zhijun has found that bee pollen form cole carries for (2013, granted patent number CN102688178 B) Take thing effective in terms of antisenility skin care；As Wang Xinglin has found that one kind is multiple for (2012, the B of granted patent number CN 102258442) Square Chinese medical extract is effective in terms of antisenility skin care.Due to plant extracts or Chinese medicinal compound extract complicated, its work( Effect is easily queried.

The present invention screens the difference in epidermal cell aging course from cell ageing mechanism using microarray chip technology Different small molecule inhereditary material simultaneously carries out functional analysis, finds first and provides the small molecule heredity related to epidermal cell aging Material, and provide the screening technique of the small molecule inhereditary material.

The content of the invention

It is an object of the invention to provide a kind of anti-aging skin care product composition.

To achieve the above object, the present invention provides a kind of composite skin care product, and it is living that the composite skin care product includes biology Property material.

Further, the bioactive substance is one or more small molecule inhereditary materials.

Further, the small molecule inhereditary material is miRNA.

Preferably, the miRNA includes miR-127-3p, miR-3198, miR-146b-5p, miR-1973, miR-424, miR-487b,miR-1274a,miR-1274b,miR-377,miR-3651,miR-181b,miR-4291,miR-154,miR- 181a,miR-455-3p,miR-381,miR-1260b,miR-1260,miR-29a,miR-29c,miR-34b,miR-548, miR-1207-5p,miR-1280,miR-4286,miR-720,miR-1183,miR-3917,miR-505,miR-3667-5p, miR-3937,miR-557,miR-3127,miR-3610,miR-601,miR-29c,miR-3907,miR-3188,miR- 1273, miR-3194, miR-623, miR-29b, miR-34a, miR-1226, miR-572, miR-134, the sieve of the miRNA The step is selected to be：MiRNA screenings are carried out with microarray chip technology, wash dry instrument with chip carries out cleaning drying to chip, uses image Analysis software is extracted to data, and selecting for differential gene is carried out with software, the miRNA filtered out and cell ageing process phase Close.

Further, the miRNA includes but is not limited to miRNA-134, miRNA-1207, miRNA-720, described MiRNA is related to cell ageing process, and the composite skin care product also includes matrix, and matrix is oil phase A and aqueous phase B, by oil phase Part is heated to 85 DEG C, and aqueous portion is heated to 95 DEG C, autoclaving, is cooled to 85 DEG C, and oil phase is added into aqueous phase and stirred simultaneously Mix to emulsification, obtain matrix, oil phase A includes glycerin monostearate 68-83%, hydrogenated lanolin 10-30%, olive oil 5- 10%th, stearic acid 2%, aqueous phase B include glycerine 4%, hyaluronic acid 0.2%, depleted water 94.8%.

Further, the composite skin care product component content is preferably matrix 95-99%, miRNA-134 0.1%th, miRNA-1207 0.1%, miRNA-720 0.1%, nicotinamide mononucleotide 0.7~4.7%.

To achieve the above object, the present invention also provides small molecule inhereditary material miRNA and is preparing anti-aging skin care product combination Purposes in thing.

Preferably, the miRNA includes but is not limited to miRNA-134, miRNA-1207, miRNA-720, described MicroRNA is related to cell ageing process.

" microRNA " or " miRNA " of the present invention refer to small molecule RNA (microRNA, being abbreviated as miRNA) Micro ribonucleic acid is translated again, is a kind of ribonucleic acid (RNA) for being about 21 to 23 nucleotides point being widely present in eucaryote Son, it can adjust the expression of other genes.MiRNA transcribes from some from DNA, but can not further be translated into protein RNA (belongs to non-coding RNA).MiRNA suppresses the gene after transcription by being combined with target messenger RNA (mRNA) Expression, plays an important role in controlling gene expression, cell cycle, organismal development sequential etc..In animal, one micro- RNA can generally regulate and control dozens of gene.

" transcription " of the present invention refers to the process of that hereditary information is transformed into RNA by DNA.As Protein synthesis The first step, transcription is the synthesis step of mRNA and non-coding RNA (tRNA, rRNA etc.).In transcription, one section of gene can be read Take, copy as mRNA；That is using specific DNA fragmentation as template, (RNA polymerize the ribonucleic acid polymerase relied on DNA Enzyme or RNA synzyme) process of Pre-mRNA is synthesized as catalyst.When mRNA is transcribed, DNA molecular double-strand is opened, in RNA In the presence of polymerase, free 4 kinds of ribonucleotides according to base pair complementarity principle be attached to DNA it is single-stranded on, and in RNA Single mRNA molecule is formed in the presence of polymerase.So far, transcription is completed.Transcription is typically more multidirectional duplications of starting point, during transcription What is transcribed is only the fragment (DNA) for having hereditary effect on DNA, not including introne.

Term " bioactive substance " of the present invention refers to influential on biological phenomena specific practice in organism Micro or a small amount of material.Such as：Some foods contain a variety of compounds with bioactivity, can cause after being acted on body each Kind biological effect, referred to as bioactive substance.Their species are various, there is carbohydrate, lipid, protein and peptide class, gene, sterol Class, alkaloid, glucoside, volatile oil etc..They are primarily present in plant food, and what is had to people is favourable, and some is harmful. Paracelsus thinks that plant is the source of human foods and medicine, and the source of poisonous substance, all foods are all poisonous substances, are not had There is avirulent food, how much left and right for only measuring size of their toxicity.

Term " genetic chip " (genechip) (also known as DNA chip, biochip) of the present invention refers to utilize sequencing by hybridization Method, the i.e. method by carrying out determining nucleic acid sequence with the nucleic acid probe hybridization of one group of known array, in one piece of substrate surface Secure the probe of target nucleotide known to sequence.When in solution carry fluorescence labeling nucleotide sequence TATGCAATCTAG, with When the nucleic acid probe of correspondence position produces complementary matching on genetic chip, the probe location most strong by determining fluorescence intensity, obtain The probe sequence of one group of sequence complete complementary is obtained, the technology of the sequence of target nucleic acid can be recombinated out accordingly.

The beneficial effects of the invention are as follows：Anti-aging skin care product composition provided by the invention is strong to the penetration of epidermis, table It is strong up to effect, and raw material are naturally safe, no dependence, have no toxic side effect

Brief description of the drawings

Fig. 1 micro-array chips screening aging correlation microRNA.

Embodiment

To make the object, technical solutions and advantages of the present invention clearer, the present invention is made into one below in conjunction with accompanying drawing It is described in detail on step ground.

Embodiment 1：

1st, epidermal cell microRNA acquisition

The epidermal cell of 200 volunteers's (age distribution is in 18-50 year) is collected, aspiration is obtained in the collection of all samples Person or its family members agree to, and sign informed consent form.

Extraction microRNA is comprised the following steps that：Sample is separately added into 750 μ l TRI REAGENT BD (Molecular Research Center companies) and 20 μ l 5N acetic acid, the Cel-miR-39 that 5 μ l concentration are 50pmol/L, shake are added after mixing Mixing is swung, stands 5 minutes at room temperature, adds 200 μ l chloroforms, concussion mixes, and stands 5 minutes at room temperature, 4 DEG C, 12000g centrifugations 15min, supernatant is extracted, add 4 μ l DNA mate (TAKARA companies), the rear addition isopropanol isometric with supernatant mixes It is even, -20 DEG C of standing 20min, 4 DEG C, 12000g centrifugation 15min, supernatant is removed, it is heavy that the cleaning of the ethanol of 1ml 75% is added into precipitation Form sediment, 4 DEG C, 7500g centrifuge washings precipitation 5min.Supernatant is removed, the water that 40 μ l RNase-free are added after precipitation room temperature is dried fills Divide dissolving, measure gained RNA concentration.

MiRNA screenings are carried out using microarray chip technology, chip uses Agilent human microRNAs Microarray (offer of Agilent companies), the strict by specification of operating process are carried out, usedSlide The chips of WasherTM 8 wash dry instrument and carry out cleaning drying to chip, LuxScanTM 10K/A twin-channel laser scanners Chip is scanned, the image analysis softwares of Lux Scan 3.0 are extracted to data, and SAM softwares carry out choosing for differential gene Choosing.Screening conditions are that False discovery rate (false discovery rate, FDR) is controlled within 5%, fold differences (fold Change, FC) it is 1.5.Filter out the following microRNAs related to human epidermal cell aging:miR-127-3p,miR-3198, miR-146b-5p,miR-1973,miR-424,miR-487b,miR-1274a,miR-1274b,miR-377,miR-3651, miR-181b,miR-4291,miR-154,miR-181a,miR-455-3p,miR-381,miR-1260b,miR-1260,miR- 29a,miR-29c,miR-34b,miR-548,miR-1207-5p,miR-1280,miR-4286,miR-720,miR-1183, miR-3917,miR-505,miR-3667-5p,miR-3937,miR-557,miR-3127,miR-3610,miR-601,miR- 29c,miR-3907,miR-3188,miR-1273,miR-3194,miR-623,miR-29b,miR-34a,miR-1226,miR- 572,miR-134。

Embodiment 2：

Genechip detection result filters out as shown in figure 1, according to genechip detection result from embodiment 1 The miRNA-134 that expression is significantly lowered in senile cell is chosen in miRNA, miRNA-1207, miRNA-720 carry out fixed in real time RT-PCR detections are measured, are concretely comprised the following steps：Use the Taqman microRNA Reverse Transcriptions of Applied Biosystems companies Box carries out reverse transcription reaction to the RNA of extraction.

Reaction system is following (20 μ l)：

Carry out 16 DEG C 30 minutes using Applied Biosystems companies thermal cyclers, 42 DEG C 30 minutes, 85 DEG C 5 minutes Reverse transcription.Reverse transcription product is carried out after suitably diluting, templates of the 5 μ l as real-time quantitative PCR is taken, uses Applied The TaqMan Universal PCR Master Mix kits of Biosystems companies carry out real-time quantitative PCR identification.PCR Reaction is in Applied Biosystems 7900HT sequence detection systems (P/N：4329002, Applied Biosystems) in Carry out.PCR reaction systems are 20 μ l, including 4 μ l RT products, 11 μ l 10 × Universal PCR MasterMix (P/N： 4324018, Applied Biosystems), 1 μ l probes (0.2mM), 4 μ l RNase-free water.By all reaction systems First carry out 95 DEG C of preheatings of 10 minutes, then often circulated 95 DEG C 15 seconds, 60 DEG C 1 minute, totally 50 circulation PCR reaction. All reactions in triplicate, take fixed threshold value every time, and minimum Ct values are as sample Ct values in determining three times.Primer sequence Row are shown in Table 1.

Table 1microRNA primer sequences

According to above-mentioned experiment, measurement result shows miRNA-134, and miRNA-1207, miRNA-720 show in senile cell Write lower, be aging correlation microRNA, by epidermal cell importing miRNA-134, miRNA-1207, miRNA-720 or its Analog, the purpose for delaying epidermal cell aging can be reached.

Embodiment 3：

Nicotinamide mononucleotide (β-Nicotinamide mononucleotide, be abbreviated as NMN) is biological cell internal memory A kind of biochemical substances, be the fit important as precursors of intracellular NADH (also referred to as DPN).Niacinamide Adenine-dinucleotide is a kind of basic redox coenzyme, whether in respiration or photosynthesis, is all risen Core pivotal role.At present, nicotinamide mononucleotide (NMN) can pass through saccharomycetes to make fermentation, chemical synthesis or external enzymatic Prepare.And NMN prepared by enzymatic in vitro is remaining without solvent, also in the absence of chiral problem, produce and homotype in body β-NMN.And there is cost advantage.The present invention uses NMN prepared by external enzymatic and small molecule inhereditary material microRNA Connected applications are on anti-aging beauty-care composite skin care product.

3.1 prepare skin care item matrix

Following component is remembered by weight percentage

Oil phase A：Glycerin monostearate 68-83%, hydrogenated lanolin 10-30%, olive oil 5-10%, stearic acid 2%；

Aqueous phase B：Glycerine 4%, hyaluronic acid 0.2%, depleted water is supplemented to 100%；

Oil phase part is heated to 85 DEG C, aqueous portion is heated to 95 DEG C, autoclaving, is cooled to 85 DEG C, by oil phase plus Enter aqueous phase and while stirring extremely emulsification.

3.2 prepare anti-aging skin care product

Following component remembers that raw material and proportioning are by weight percentage：

Dosages of the NMN (nicotinamide mononucleotide) in anti-aging beauty-care composite skin care product is 0.7~4.7%；

Emulsification skin care item matrix is cooled to less than 35 DEG C addition microRNA, and stirring adds NMN to mixing, and stirs to mixed It is even, obtain skin care item mixture；

The skin care item as made from above-mentioned steps can be packed after check analysis is qualified.

Embodiment 4：

The product of trial-production is respectively supplied to old (one's mid-60s), middle aged (one's mid-40s), prime of life group (one's mid-30s), blue or green In year (20 years old or so), totally four groups (every group of 30 people), try out 4 months.Once in the morning and once at night, the about big small capital of soybean grain is once smeared Product, product is applied directly to exposed skin (face and the back of the hand), skin obvious delicate moist degree in position is smeared after one week and is increased Add, crystalistic sense enhancing.Wherein, in Younger group 5 subjects it is original go out variola shape substantially relax；7 application on human skin are thin in middle aged group Line significantly subtracts light, and skin is bright and new flexible；Old age group feedback skin water humidity improves and skin falls to consider symptom mitigation to be worth doing, wherein 6 people Color spot substantially subtracts light, and wrinkle and cutis laxa phenomenon obtain different degrees of improvement.

It should be noted last that above embodiment is merely illustrative of the technical solution of the present invention and unrestricted, Although the present invention is described in detail with reference to preferred embodiment, it will be understood by those within the art that, can be right Technical scheme is modified or equivalent substitution, and without departing from the spirit and scope of technical solution of the present invention, its is equal It should cover among scope of the presently claimed invention.

Claims (8)

1. a kind of composite skin care product, it is characterised in that the composite skin care product includes bioactive substance.

2. composite skin care product according to claim 1, it is characterised in that the bioactive substance is one or more small Molecular genetic material.

3. any composite skin care product according to claim 1 or claim 2, it is characterised in that the small molecule inhereditary material is miRNA。

4. composite skin care product according to claim 3, it is characterised in that including miR-127-3p, miR-3198, miR- 146b-5p,miR-1973,miR-424,miR-487b,miR-1274a,miR-1274b,miR-377,miR-3651,miR- 181b,miR-4291,miR-154,miR-181a,miR-455-3p,miR-381,miR-1260b,miR-1260,miR-29a, miR-29c,miR-34b,miR-548,miR-1207-5p,miR-1280,miR-4286,miR-720,miR-1183,miR- 3917,miR-505,miR-3667-5p,miR-3937,miR-557,miR-3127,miR-3610,miR-601,miR-29c, miR-3907,miR-3188,miR-1273,miR-3194,miR-623,miR-29b,miR-34a,miR-1226,miR-572, MiR-134, the miRNA screen according to following steps：MiRNA screenings are carried out with microarray chip technology, dry instrument is washed with chip Cleaning drying is carried out to chip, data extracted with image analysis software, selecting for differential gene is carried out with software, is screened The miRNA gone out is related to cell ageing process.

5. according to any composite skin care products of claim 1-4, it is characterised in that the miRNA includes but is not limited to MiRNA-134, miRNA-1207, miRNA-720, the miRNA is related to cell ageing process, and the composite skin care product is also Including matrix, matrix is oil phase A and aqueous phase B, and oil phase part is heated into 85 DEG C, and aqueous portion is heated to 95 DEG C, autoclaving, 85 DEG C are cooled to, oil phase is added into aqueous phase and stirred simultaneously to emulsifying, obtains matrix, oil phase A includes glycerin monostearate 68- 83%th, hydrogenated lanolin 10-30%, olive oil 5-10%, stearic acid 2%, aqueous phase B include glycerine 4%, hyaluronic acid 0.2%, Depleted water 94.8%.

6. according to any composite skin care product of claim 5, it is characterised in that the composite skin care product component content is Matrix 95-99%, miRNA-1340.1%, miRNA-12070.1%, miRNA-7200.1%, nicotinamide mononucleotide 0.7~ 4.7%.

7. use of the small molecule inhereditary material miRNA in anti-aging skin care product composition any in claim 1 to 6 is prepared On the way.

8. purposes according to claim 7, it is characterised in that the miRNA includes but is not limited to miRNA-134, miRNA- 1207, miRNA-720, the miRNA is related to cell ageing process.