Diol type ginsenoside nano-emulsion with anti aging effect function and preparation method thereof
Technical field
The present invention relates to a kind of nanoemulsions and its preparation method and application, and in particular to anti aging effect function
Diol type ginsenoside nano-emulsion and preparation method thereof.
Background technique
Ginseng is Araliaceae herbaceos perennial (Panax ginseng, C.A.Mey.), containing amino acid, vitamin,
It is polysaccharide, ginseng essential oil, inorganic elements, alkaloid, peptide matters, fats, saponin(e, organic acid, flavonoids, sterol, wooden
The various chemical components such as element, enzyme, wherein ginsenoside is the main active constituent of ginseng.According to the difference of ginsengenin,
Ginsenoside is divided into oleanolic acid saponins, protopanaxatriol saponins and protopanoxadiol saponins.Diol type ginseng soap
Glycosides curative effect with highly significant in terms of anticancer, anti-aging, tonifying primordial Qi, enhancing Abwehrkraft des Koepers, improvement.In recent years
Come, since the effect of diol type ginsenoside is in terms of anti-aging is significant, is applied to drug and cosmetic field to reach
The research of anti aging effect becomes a popular research.But diol type ginsenoside polarity is big, fat-soluble difference, is not easy through skin
Skin, so bringing a very hang-up to dermatologic.
Summary of the invention
Deficiency in view of panoxadiol type ginsenoside extract described above as product, it is an object of the invention to
There is provided a kind of diol type ginsenoside nano-emulsion using diol type ginsenoside as raw material, with anti aging effect function and its
Preparation method.
A kind of diol type ginsenoside nano-emulsion, is grouped as by following group: by diol type ginsenoside extract and water group
At water phase, oil mutually and blended emulsifier;Wherein, the mass ratio of the water phase and oily phase is 5~8:45, the oil phase and mixing
The mass ratio of emulsifier is 7.5~8:2~2.5, and mass concentration of the diol type ginsenoside extract in water phase is 15
~25%, the diol type ginsenoside Rb that the diol type ginsenoside extract contains1、Rb2, Rc and Rd the sum of content no
Lower than 38%.
In the above-mentioned technical solutions, the oil is mutually isopropyl palmitate or isopropyl myristate, the mixing cream
Agent is obtained by sorbester p17 and 1,2- propylene glycol 3:1 in mass ratio~3 configuration.
In the above-mentioned technical solutions, the preparation method of the diol type ginsenoside extract includes the following steps:
(1) Ca (OH) for being 11~13 by ginseng coarse powder and pH2Aqueous solution in solid-liquid ratio g:mL=1:10~15 ratio
Mixing is 20 DEG C~35 DEG C in Extracting temperature, extracts under conditions of ultrasonic power 120w~160w, extracting solution filtering obtains
Ginsenoside extracting solution;
(2) the ginsenoside extracting solution for obtaining step (1) is first eluted with water, is then used by large pore resin absorption column
The ethanol gradient elution that concentration expressed in percentage by volume is 35%~85%, collects eluent, and concentration, freeze-drying obtain diol type ginseng
Saponin extract.
In the above-mentioned technical solutions, the preparation method of the diol type ginsenoside extract includes the following steps:
(1) Ca (OH) for being 13 by ginseng coarse powder and pH2Aqueous solution is mixed in the ratio of solid-liquid ratio g:mL=1:10, is being mentioned
Taking temperature is 20 DEG C, is extracted under conditions of ultrasonic extraction power 160w, and extracting solution filtering obtains ginsenoside extracting solution;
(2) the ginsenoside extracting solution for obtaining step (1) is by large pore resin absorption column, with the water of 3~6 times of column volumes
Elution, then successively with 35% ethyl alcohol of 3~6 times of column volumes, 85% ethanol elution of 3~6 times of column volumes, 85% ethyl alcohol of collection
Eluent, concentration, freeze-drying, obtains diol type ginsenoside extract.
In the above-mentioned technical solutions, in step (2), first with the water elution of 3~6 times of column volumes, then successively with 3~6
35% ethyl alcohol of times column volume, 60% ethyl alcohol of 3~6 times of column volumes, 3~6 times of column volumes 85% ethanol elution, collect 85%
Ethanol eluate, it is concentrated, be freeze-dried diol type ginsenoside extract dry product.
In the present invention, ginsenoside extracting solution that step (1) is obtained is adsorbed by large pore resin absorption column, stand 2h with
On, so that the nonpolarity such as ginsenoside and low pole ingredient is sufficiently adsorbed in macroporous absorbent resin, later, is first eluted with water, with
Remove unadsorbed most of polar impurity;Then miscellaneous to remove three alcohol type ginsenosides and small part with 35% ethanol elution
Matter;Other type ginsenosides and impurity are further removed with 60% ethanol elution again later, to improve the diol type in extract
The purity of ginsenoside, the present invention in the step can omit;Finally with 85% ethanol elution, to collect diol type ginsenoside,
Mainly contain Rb1、Rb2, Rc and tetra- kinds of Rd, the sum of content is not less than the 38% of total extract amount.
In the above-mentioned technical solutions, in step (1), in the extraction step, extraction time is 1~3 time, is mentioned every time
Taking the time is 20~60min, preferably 20min.
In the above-mentioned technical solutions, in step (2), by the ginsenoside extracting solution, filtering, filtrate is concentrated into leaching
Cream is added the dissolution of 80% ethyl alcohol, is filtered to obtain ginsenoside filtrate;The concentration of ginsenoside filtrate removes ethyl alcohol, and distilled water is added
Dissolution obtains ginsenoside aqueous solution, and aqueous solution is with the rate of 0.2~0.3mL/s of flow velocity by being washed with water in advance to no ethanol flavor
Large pore resin absorption column, 2h or more can be placed after upper prop, so that the ingredients such as ginsenoside in filtrate sufficiently adsorb
In on macroporous absorbent resin.The macroporous absorbent resin preferably uses macroporous absorbent resin D101.
The present invention also provides the preparation methods of above-mentioned diol type ginsenoside nano-emulsion, include the following steps:
(a) blended emulsifier being made of sorbester p17 and 1,2-PD is added in oily phase according to the proportion, at 40 DEG C
Heating condition under be stirred with the revolving speed of 650~1300r/min, obtain the mixed solution of clear state;
(b) diol type ginseng ginseng saponin extract is dissolved in water and obtains water phase, water phase is added dropwise to what step (a) obtained
In mixed solution, stir evenly.
In the above-mentioned technical solutions, in step (b), by water phase under the revolving speed stirring of 40 DEG C, 650~1300r/min
It is added dropwise in the mixed solution that step (a) obtains.
The present invention also provides a kind of cosmetic composition for anti aging effect, the composition includes above-mentioned glycol
Type ginsenoside nano-emulsion.The composition can also include the acceptable auxiliary material of cosmetics, type for the auxiliary material and
Usage amount can use the auxiliary material and usage amount of this field routine as needed without being particularly limited to, and can be prepared
The cosmetic composition with anti aging effect containing above-mentioned diol type ginsenoside nano-emulsion.The cosmetic composition can
Think liquid-type, emulsion-type, cream type cosmetics and facial mask class cosmetology skin care item.
Beneficial effects of the present invention:
W/o type nano-emulsion is made in diol type ginsenoside by the present invention, and diol type ginsenoside soluble easily in water is wrapped up
In inner aqueous phase, surface is oily phase, and this considerably increases the compatibilities of emulsion and skin, and since emulsion partial size is in Nano grade,
The performance for passing it through skin dramatically increases, and has certain protective effect and slow releasing function to diol type ginsenoside,
Significantly improve its anti aging effect function.It extracts to obtain the diol type ginseng as active constituent using buck ultrasonic extraction
Saponin extract, wherein diol type ginsenoside Rb1, Rb2, Rc, the sum of content of Rd is not less than 38%.The present invention uses constant temperature
Low-energy emulsification method prepares nano-emulsion, and preparation process is easy, and quality is easily controllable.
Detailed description of the invention
Fig. 1 is diol type ginsenoside extract dry product HPLC chromatogram.
Fig. 2 is the particle diameter measurements of diol type ginsenoside nano-emulsion.
Fig. 3 is that the skin of diol type ginsenoside nano-emulsion adds up transit dose result.
Fig. 4 is the skin hold-up result of diol type ginsenoside nano-emulsion.
Fig. 5 is the hydroxyproline content measurement result of diol type ginsenoside nano-emulsion each group.
Fig. 6 is the type i collagen assay result of diol type ginsenoside nano-emulsion each group.
Fig. 7 is the type III Contents of collagen result of diol type ginsenoside nano-emulsion each group.
Fig. 8 is the TIMP-1 assay result of diol type ginsenoside nano-emulsion each group.
Fig. 9 is the TIMP-2 assay result of diol type ginsenoside nano-emulsion each group.
Figure 10 is the MMP-1 assay result of diol type ginsenoside nano-emulsion each group.
Specific embodiment
Present invention will be described in further detail below with reference to the accompanying drawings and specific embodiments, but the present invention does not limit to
In these embodiments.In following embodiments, unless otherwise specified, used experimental method is conventional method, material used
Material, reagent etc. can be bought from biological or chemical company.
Ginseng coarse powder: 5 years raw dry ginsengs crush, powder crosses No. four sieves, about 250 μm of partial size of coarse powder obtained, for mentioning
It takes.
Embodiment 1
Diol type ginsenoside extract is prepared as follows to obtain:
(1) by the Ca (OH) of ginseng coarse powder and pH=132Aqueous solution is mixed in the ratio of solid-liquid ratio=1:10 (g:mL),
Extracting temperature is 20 DEG C, and ultrasonic extraction power is 160w, is extracted under conditions of extraction time 20min, will after the completion of extracting
Drug residue filter obtains ginsenoside extracting solution;Extracting solution filtering, filtrate are concentrated into medicinal extract, and the dissolution of 80% ethyl alcohol, filtering is added
Ginsenoside filtrate is obtained, the concentration of ginsenoside filtrate removes ethyl alcohol, and distilled water dissolution is added, obtains ginsenoside aqueous solution;
(2) macroporous absorbent resin (D101, Sinopharm Chemical Reagent Co., Ltd.) is fitted into splitter, and pillar is washed with water
To macroporous absorbent resin without alcohol taste, ginsenoside aqueous solution wet process upper prop, flow velocity is 0.2mL per second, with 4 times after standing 2 hours
Column volume washes column, to remove most of impurity, then with 3 times of 35% ethanol elutions of column volume, removes three alcohol type ginsenosides
With small part impurity, then with 3 times of 85% ethanol elutions of column volume, 85% ethanol eluate is collected;
(3) by 85% ethanol eluate, ethyl alcohol is recycled, concentrate is freeze-dried 72h at -20 DEG C, and diol type people is made
Join saponin extract dry product.
Dry product 500mg is taken, 50mL methanol is added, ultrasonic dissolution obtains 10mg/mL solution.It is detected with HPLC method, detection side
Method: chromatographic column Hypersil Gold C18 (250mm × 4.6 μm, 5 μm), Detection wavelength 203nm, sample volume are 10 μ L, column
Temperature is 40 DEG C, flow velocity 1.000mL/min, gradient elution such as the following table 1:
1 gradient elution table of table
Testing result such as Fig. 1 can be obtained by the calculating of peak value and be contained diol type ginsenoside Rb in dry product1, Rb2, Rc,
Rd, the sum of content are about 42%.
Embodiment 2
With the raw material for the diol type ginsenoside extract that embodiment 1 is prepared, diol type ginsenoside nanometer is prepared
Cream, the preparation method is as follows:
(1) sorbester p17 and 1,2-PD 2:1 in mass ratio mix to obtain blended emulsifier, (i.e. by isopropyl palmitate
Oily phase) it is mixed with blended emulsifier according to mass ratio 7.5:2.5, under 40 DEG C of heating condition, with magnetic stirring apparatus with revolving speed
The rate of 650r/min is stirred, and stirring to mixed solution is in clear state, obtains mixed solution;
(2) diol type ginsenoside extract is dissolved in water and obtains water phase, under 40 DEG C of heating condition, with magnetic stirring apparatus with
Under conditions of the rate stirring of revolving speed 650r/min, water phase is added dropwise to oil mutually and in the mixed solution of blended emulsifier,
Mixed system is stopped being added dropwise, is obtained diol type ginsenoside nano-emulsion by clarifying the critical point become cloudy.
Wherein, mass concentration of the diol type ginsenoside extract in water phase is 200mg/mL;The water phase and oily phase
Mass ratio be 8:45.
Embodiment 3
With the raw material for the diol type ginsenoside extract that embodiment 1 is prepared, diol type ginsenoside nanometer is prepared
Cream, the preparation method is as follows:
(1) sorbester p17 and 1,2-PD 2:1 in mass ratio mix to obtain blended emulsifier, by isopropyl myristate
(i.e. oily phase) is mixed with blended emulsifier according to mass ratio 7.5:2.5, under 40 DEG C of heating condition, with magnetic stirring apparatus to turn
The rate of fast 650r/min is stirred, and stirring to mixed solution is in clear state, obtains mixed solution;
(2) diol type ginsenoside extract is dissolved in water and obtains water phase, under 40 DEG C of heating condition, with magnetic stirring apparatus with
Under conditions of the rate stirring of revolving speed 650r/min, water phase is added dropwise to oil mutually and in the mixed solution of blended emulsifier,
Mixed system is stopped being added dropwise, is obtained diol type ginsenoside nano-emulsion by clarifying the critical point become cloudy.
Wherein, mass concentration of the diol type ginsenoside extract in water phase is 200mg/mL;The water phase and oily phase
Mass ratio be 8:45.
Embodiment 4
With the raw material for the diol type ginsenoside extract that embodiment 1 is prepared, diol type ginsenoside nanometer is prepared
Cream, the preparation method is as follows:
(1) sorbester p17 and PEG400 2:1 in mass ratio mix to obtain blended emulsifier, by isopropyl palmitate (i.e. oily phase)
It is mixed with blended emulsifier according to mass ratio 7.5:2.5, under 40 DEG C of heating condition, with magnetic stirring apparatus with revolving speed 650r/
The rate of min is stirred, and stirring to mixed solution is in clear state, obtains mixed solution;
(2) diol type ginsenoside extract is dissolved in water and obtains water phase, under 40 DEG C of heating condition, with magnetic stirring apparatus with
Under conditions of the rate stirring of revolving speed 650r/min, water phase is added dropwise to oil mutually and in the mixed solution of blended emulsifier,
Mixed system is stopped being added dropwise, is obtained diol type ginsenoside nano-emulsion by clarifying the critical point become cloudy.
Wherein, mass concentration of the diol type ginsenoside extract in water phase is 200mg/mL;The water phase and oily phase
Mass ratio be 8:45.
Embodiment 5
With the raw material for the diol type ginsenoside extract that embodiment 1 is prepared, diol type ginsenoside nanometer is prepared
Cream, the preparation method is as follows:
(1) sorbester p17 and 1,2-PD 3:1 in mass ratio mix to obtain blended emulsifier, (i.e. by isopropyl palmitate
Oily phase) it is mixed with blended emulsifier according to mass ratio 7.5:2.5, under 40 DEG C of heating condition, with magnetic stirring apparatus with revolving speed
The rate of 650r/min is stirred, and stirring to mixed solution is in clear state, obtains mixed solution;
(2) diol type ginsenoside extract is dissolved in water and obtains water phase, under 40 DEG C of heating condition, with magnetic stirring apparatus with
Under conditions of the rate stirring of revolving speed 650r/min, water phase is added dropwise to oil mutually and in the mixed solution of blended emulsifier,
Mixed system is stopped being added dropwise, is obtained diol type ginsenoside nano-emulsion by clarifying the critical point become cloudy.
Wherein, mass concentration of the diol type ginsenoside extract in water phase is 200mg/mL;The water phase and oily phase
Mass ratio be 8:45.
Embodiment 6
With the raw material for the diol type ginsenoside extract that embodiment 1 is prepared, diol type ginsenoside nanometer is prepared
Cream, the preparation method is as follows:
(1) sorbester p17 and 1,2-PD 1:1 in mass ratio mix to obtain blended emulsifier, (i.e. by isopropyl palmitate
Oily phase) it is mixed with blended emulsifier according to mass ratio 7.5:2.5, under 40 DEG C of heating condition, with magnetic stirring apparatus with revolving speed
The rate of 650r/min is stirred, and stirring to mixed solution is in clear state, obtains mixed solution;
(2) diol type ginsenoside extract is dissolved in water and obtains water phase, under 40 DEG C of heating condition, with magnetic stirring apparatus with
Under conditions of the rate stirring of revolving speed 650r/min, water phase is added dropwise to oil mutually and in the mixed solution of blended emulsifier,
Mixed system is stopped being added dropwise, is obtained diol type ginsenoside nano-emulsion by clarifying the critical point become cloudy.
Wherein, mass concentration of the diol type ginsenoside extract in water phase is 200mg/mL;The water phase and oily phase
Mass ratio be 8:45.
Embodiment 7
With the raw material for the diol type ginsenoside extract that embodiment 1 is prepared, diol type ginsenoside nanometer is prepared
Cream, the preparation method is as follows:
(1) sorbester p17 mixes to obtain blended emulsifier with 1,2-PD 2:1 in mass ratio, and isopropyl palmitate is (i.e. oily
Phase) it is mixed with blended emulsifier according to mass ratio 8:2, under 40 DEG C of heating condition, with magnetic stirring apparatus with revolving speed 650r/
The rate of min is stirred, and stirring to mixed solution is in clear state, obtains mixed solution;
(2) diol type ginsenoside extract is dissolved in water and obtains water phase, under 40 DEG C of heating condition, with magnetic stirring apparatus with
Under conditions of the rate stirring of revolving speed 650r/min, water phase is added dropwise to oil mutually and in the mixed solution of blended emulsifier,
Mixed system is stopped being added dropwise, is obtained diol type ginsenoside nano-emulsion by clarifying the critical point become cloudy.
Wherein, mass concentration of the diol type ginsenoside extract in water phase is 200mg/mL;The water phase and oily phase
Mass ratio be 1:6.
Embodiment 8
With the raw material for the diol type ginsenoside extract that embodiment 1 is prepared, diol type ginsenoside nanometer is prepared
Cream, the preparation method is as follows:
(1) sorbester p17 mixes to obtain blended emulsifier with 1,2-PD 2:1 in mass ratio, and isopropyl palmitate is (i.e. oily
Phase) it is mixed with blended emulsifier according to mass ratio 7.5:2.5, under 30 DEG C of heating condition, with magnetic stirring apparatus with revolving speed
The rate of 650r/min is stirred, and stirring to mixed solution is in clear state, obtains mixed solution;
(2) diol type ginsenoside extract is dissolved in water and obtains water phase, under 30 DEG C of heating condition, with magnetic stirring apparatus with
Under conditions of the rate stirring of revolving speed 650r/min, water phase is added dropwise to oil mutually and in the mixed solution of blended emulsifier,
Mixed system is stopped being added dropwise, is obtained diol type ginsenoside nano-emulsion by clarifying the critical point become cloudy.
Wherein, mass concentration of the diol type ginsenoside extract in water phase is 200mg/mL;The water phase and oily phase
Mass ratio be 8:45.
Embodiment 9
With the raw material for the diol type ginsenoside extract that embodiment 1 is prepared, diol type ginsenoside nanometer is prepared
Cream, the preparation method is as follows:
(1) sorbester p17 mixes to obtain blended emulsifier with 1,2-PD 2:1 in mass ratio, and isopropyl palmitate is (i.e. oily
Phase) it is mixed with blended emulsifier according to mass ratio 7.5:2.5, under 40 DEG C of heating condition, with magnetic stirring apparatus with revolving speed
The rate of 1300r/min is stirred, and stirring to mixed solution is in clear state, obtains mixed solution;
(2) diol type ginsenoside extract is dissolved in water and obtains water phase, under 40 DEG C of heating condition, with magnetic stirring apparatus with
Under conditions of the rate stirring of revolving speed 1300r/min, water phase is added dropwise to oil mutually and in the mixed solution of blended emulsifier,
Mixed system is stopped being added dropwise, is obtained diol type ginsenoside nano-emulsion by clarifying the critical point become cloudy.
Wherein, mass concentration of the diol type ginsenoside extract in water phase is 200mg/mL;The water phase and oily phase
Mass ratio be 8:45.
Embodiment 10
With the raw material for the diol type ginsenoside extract that embodiment 1 is prepared, diol type ginsenoside nanometer is prepared
Cream, the preparation method is as follows:
(1) sorbester p17 mixes to obtain blended emulsifier with 1,2-PD 2:1 in mass ratio, and isopropyl palmitate is (i.e. oily
Phase) it is mixed with blended emulsifier according to mass ratio 7.5:2.5, under 40 DEG C of heating condition, with magnetic stirring apparatus with revolving speed
The rate of 650r/min is stirred, and stirring to mixed solution is in clear state, obtains mixed solution;
(2) diol type ginsenoside extract is dissolved in water and obtains water phase, under 40 DEG C of heating condition, with magnetic stirring apparatus with
Under conditions of the rate stirring of revolving speed 650r/min, water phase is added dropwise to oil mutually and in the mixed solution of blended emulsifier,
Mixed system is stopped being added dropwise, is obtained diol type ginsenoside nano-emulsion by clarifying the critical point become cloudy.
Wherein, mass concentration of the diol type ginsenoside extract in water phase is 250mg/mL;The water phase and oily phase
Mass ratio be 8:45.
The measurement of 11 diol type ginsenoside nano-emulsion particle diameter of embodiment
The diol type ginsenoside nano-emulsion for taking appropriate embodiment 2 to be prepared is placed in testing graininess ware, uses Zeta
Current potential particle size analyzer (manoBrook 90plus zeta, Brookhaven) carries out particle size determination to it, measures 3 times.Measurement
As a result such as Fig. 2, Fig. 2 meets nano-emulsion granularity requirements as it can be seen that diol type ginsenoside nano-emulsion average grain diameter is 17.4nm.
The measurement of 12 diol type ginsenoside nano-emulsion vitro cumulative transit dose of embodiment
Six adult male mices are taken, are randomly divided into two groups, every group three, respectively diol type ginsenoside nano-emulsion
(embodiment 2 is prepared) group and diol type ginsenoside aqueous solution group.Mouse part skin is taken, Transdermal diffusion cell is fixed on,
It is 4.2mg/mL ginsenoside nano-emulsion and 4.2mg/mL ginsenoside aqueous solution that two groups are administered 3mL concentration respectively.Receiving liquid is
PBS phosphate buffer is sampled in 0.5h, 1h, 2h, 3h, 4h, 6h, 8h, 12h, amount of sampling 2mL, supplements 2mL after sampling every time
PBS phosphate buffer, receiving liquid of sampling every time carry out high performance liquid chromatography measurement, calculate two groups of diol type ginsenoside units
Skin area adds up transit dose.
Transit dose mapping is added up to nano-emulsion group and aqueous solution group unit skin respectively, as a result such as Fig. 3, it is seen then that nano-emulsion
Group diol type ginsenoside transit dose is apparently higher than aqueous solution group, shows that diol type ginsenoside nano-emulsion can effectively penetrate skin
Skin has good percutaneous abilities.
The measurement of 13 diol type ginsenoside nano-emulsion skin hold-up of embodiment
In embodiment 11, mouse skin after every group of administration 12h is taken, blank receiving liquid rinse off skin remained on surface drug,
Fixed clip medicine-feeding part skin, shreds, and is homogenized with tissue homogenizer, adds 2mL PBS phosphate buffer ultrasonic extraction 20min,
8000r/min centrifugation 15min takes supernatant, carries out efficient liquid phase measurement, calculates and be detained diol type content of ginsenoside in skin.
As a result such as Fig. 4, it is seen then that the diol type ginsenoside skin hold-up of nano-emulsion group is significantly higher than aqueous solution group, this
Diol type ginsenoside can be detained in skin part after meaning nano-emulsion percutaneous drug delivery, there is certain slow releasing function, be extended
Drug treating time improves bioavilability.
14 diol type ginsenoside nano-emulsion anti aging effect zoopery of embodiment
It studies the influence of diol type ginsenoside nano-emulsion (preparation of embodiment 2) to collagen content in skin: taking into
Year male mice 30, back depilation are divided into six groups, are divided into ginsenoside nano-emulsion high dose group (1.54mg/mL), people
Join saponin(e nano-emulsion middle dose group (0.77mg/mL), ginsenoside nano-emulsion low dose group (0.38mg/mL), ginsenoside water
Solution group (0.77mg/mL), blank nano-emulsion group, positive controls (the five compact frost of precious moisturizing of skin makeup treasured).
Every group of mouse is smeared daily to be administered once, and every is administered 0.2mL every time, and successive administration 4 weeks, it is small to record each group weekly
Mouse changes of weight, the 4th week execution mouse, takes back medicine-feeding part skin, -40 DEG C freeze.
Every group of every mouse skin 0.5g is respectively weighed, is shredded, is added in homogenate tube, adds 9 times of amount 0.01mmol PBS
Buffer is homogenized with interior cut type tissue homogenizer, and 10% homogenate is made, and every pipe homogenate crushes 1min with cell pulverization instrument,
It is centrifuged 10min in 8000r/min centrifuge, takes supernatant, is frozen.
Hydroxyproline in supernatant, type i collagen, type III collagen, matrix gold are detected respectively by ELisa kit specification
Proteases -1 (MMP-1), matrix metalloproteinase inhibitors -1 (TIMP-1), Tissue inhibitor of metalloproteinase-2
(TIMP-2) content.
As a result such as Fig. 5~10, ginsenoside nano-emulsion group can be dramatically increased compared with other groups hydroxyproline, type i collagen,
Type III collagen content inhibits the matrix metalloproteinase inhibitors -1 (TIMP-1) and matrix metalloproteinase of decomposition of collagen
Inhibiting factor -2 (TIMP-2) significantly increases, and the Fibroblast collagenase (MMP-1) of decomposition of collagen is promoted to reduce.The above index
And show dose dependent.It can be seen that diol type ginsenoside nano-emulsion can be promoted by activating collagen regulatory factor
It is synthesized into skin collagen, collagenase activity is inhibited to achieve the purpose that anti aging effect.
The preferred embodiment of the present invention has been described in detail above, but the invention be not limited to it is described
Embodiment, those skilled in the art can also make various equivalent on the premise of not violating the inventive spirit of the present invention
Variation or replacement, these equivalent variation or replacement are all included in the scope defined by the claims of the present application.