CN105147586A - Rose fermentation puree as well as preparation method and application thereof - Google Patents
Rose fermentation puree as well as preparation method and application thereof Download PDFInfo
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Abstract
The invention discloses rose fermentation puree as well as a preparation method and an application thereof. The preparation method of the rose fermentation puree comprises the following steps: dry rose powder, water and yeast are mixed to obtain an initial system, the initial system is subjected to fermentation cultivation, the rose fermentation puree is obtained, wherein, the yeast is Pasteur yeast. A rose fermentation puree cosmetic doesn't contain any chemical component, foreign matters such as enzyme are not required to be added, not only is production cost saved, but also the production steps are simplified maximally, the fermentation technology can realize mass production and industrial production, and the product quality stability can be fully guaranteed. At the same time, the rose fermentation puree can be directly used as a finished production of a facial mask, essence or toner and is more natural compared with other products existing in the market, and no negative effect is caused to the skin; low-molecular-weight peptide can be obtained, the skin can absorb the rose fermentation puree thoroughly, and the rose fermentation puree can be widely applied in preparing whitening and/or anti-aging products.
Description
Technical field
The invention belongs to biological technical field, be specifically related to a kind of Flos Rosae Rugosae fermentation oleo stock and preparation method thereof and application.
Background technology
Flos Rosae Rugosae, Rosaceae Rosa fallen leaves bush.Flos Rosae Rugosae has very high economic worth, is generally used for refining quintessence oil and pigment.Flos Rosae Rugosae complicated component, containing the number of chemical such as Polyphenols, flavonoid composition.These chemical compositions have physiologically actives such as reducing and eliminate free radical, antioxidant activity, antithrombotic, anticancer, antiinflammatory, antibacterial, immunoregulation effect, blood fat reducing and prevention of cardiac.China's Rosa Rugosa Resources enriches, for Application and Development Flos Rosae Rugosae provides advantage.Develop rapidly along with Flos Rosae Rugosae cultivation industry and range of application is widened, to Flos Rosae Rugosae nutritional labeling and in medical care effect, research is also progressively deeply.Also constantly perfect to the extraction process of rose active material, but extracting method is mainly based on homogenate extraction method, water extraction.
What existing rose active thing extraction the most often adopted is water extraction: Flos Rosae Rugosae dry product adds deionized water after pulverizing, and stirring and leaching a period of time after tune pH, get supernatant concentration after centrifugal, finally use alcohol settling, crushed after being dried obtains product.But the method introduces Organic substance in leaching process, not only welding, and potential potential safety hazard is there is in the process of application.
Summary of the invention
An object of the present invention is to provide a kind of Flos Rosae Rugosae fermentation oleo stock and preparation method thereof.
The preparation method of Flos Rosae Rugosae fermentation oleo stock provided by the invention, comprises the steps: Flos Rosae Rugosae dry powder, water and zymocyte to be mixed to get initial system, and carries out fermentation culture to initial system, obtain Flos Rosae Rugosae proferment pulp cosmetic.
In above-mentioned preparation method, the proportioning of described zymocyte, described Flos Rosae Rugosae dry powder and water is (15-25) ml:(7-15) g:350g, specifically can be 15ml:12g:350g, described zymogenic bacterium bacterial concentration is 10
5-10
8cFU/ml.
The temperature of described fermentation culture is 25-38 DEG C, and specifically can be 35 DEG C, the time is 24-32h, specifically can be 24h.
Described zymocyte can be yeast, and specifically can be saccharomyces pastorianus bacterium Saccharomycespastorianus, ATCC is numbered 28827.
Described zymocyte specifically can the form of its culture fluid or suspension exist, and the pH value of described zymogenic culture fluid or suspension is 6.5-7.5.
Rose Blume Getrocknete alabastrum taken from by described Flos Rosae Rugosae dry powder, and the order number of described Flos Rosae Rugosae dry powder is 20-50 order, specifically can be 30 orders, can by Rose Blume Getrocknete alabastrum through pulverizer pulverize, sieve after obtain.
In above-mentioned preparation method, also comprise and sterilizing, centrifugal, the step of getting supernatant are carried out successively to gained fermentation liquid after described fermentation culture, namely obtain Flos Rosae Rugosae fermentation oleo stock.
Wherein, the condition of described sterilizing is as follows: sterilising temp is 115-121 DEG C (specifically can be 115 DEG C), and sterilization time is 15-25min (specifically can be 20min).
Described centrifugal condition is as follows: centrifugal rotational speed is 3800-4200r/min (specifically can be 4000r/min), centrifugation time is 15-30min (specifically can be 20min), and centrifugal radius is 9cm.
The present invention's preparation-obtained Flos Rosae Rugosae fermentation oleo stock also belongs to protection scope of the present invention.
The present invention preparation-obtained Flos Rosae Rugosae fermentation oleo stock prepare the application that has in the cosmetics of following at least one function or directly as the application had in the cosmetics of following at least one function:
1) whitening; 2) defying age; 3) DPPH free radical is removed; 4) restraint of tyrosinase.
In above-mentioned application, described cosmetics specifically can be any one in facial film, essence or cosmetic water.
Prove by experiment: Flos Rosae Rugosae proferment pulp cosmetic prepared by the present invention is not containing any chemical composition, can directly use as the finished product of facial film or essence or cosmetic water, more natural than existing other products on the market, any negative interaction can not be caused to skin; Compared with common extracting method, the present invention obtains small component polypeptide, is easy to allow skin fully absorb.
Compared with prior art, the present invention has following beneficial effect:
(1) the present invention adopts saccharomyces pastorianus to ferment to Flos Rosae Rugosae, remains whole functional component and the activity thereof of plant, avoids the active component loss that extracting method causes;
(2) fermentation method of the present invention, to in Flos Rosae Rugosae extracts active ingredients process, do not add any organic reagent, fermentation temperature and fermentation pH gentleness, active components of plants structure is not destroyed, keep the natural activity of plant,, save production cost meanwhile, and maximizedly simplify production stage, enable this fermentation technique realize a large amount of production, suitability for industrialized production, and fully can ensure the stability of product quality;
(3) do not add essence etc. in Flos Rosae Rugosae proferment pulp cosmetic provided by the invention to chemical composition, guarantee the safety of product to human body.
(4) material such as protein, polysaccharide, flavone be rich in Flos Rosae Rugosae fermentation oleo stock provided by the invention, has synergism with the probiotics fermention product component in ferment filtrate, thus has antioxidation, defying age and whitening function significantly.
Accompanying drawing explanation
Fig. 1 is the active polypeptide molecular weight distribution HPLC collection of illustrative plates of Flos Rosae Rugosae fermentation oleo stock in embodiment 2.
Fig. 2 is Flos Rosae Rugosae fermentation oleo stock (solvent is deionized water) of variable concentrations in embodiment 2 and the relation curve of DPPH free radical scavenging activity.
Fig. 3 be in embodiment 2 10% Flos Rosae Rugosae fermentation oleo stock (solvent is deionized water) and 1% arbutin to the comparison diagram of the suppression ratio of tryrosinase.
Detailed description of the invention
The experimental technique used in following embodiment if no special instructions, is conventional method.
Material used in following embodiment, reagent etc., if no special instructions, all can obtain from commercial channels.
Fluid medium in following embodiment is by solute and solvent composition, and solvent is water, and the concentration in solute and liquid medium within thereof is: glucose 20g/L, peptone 20g/L, yeast extract 10g/L, pH value is 7.0.
YPD culture medium in following embodiment is by solute and solvent composition, solvent is water, concentration in solute and liquid medium within thereof is: yeast extract 1% (mass fraction), tryptone 2% (mass fraction), glucose 2% (mass fraction), pH value is 7.0.
Flos Rosae Rugosae dry flower in following embodiment is sold from market.
The ATCC of the saccharomyces pastorianus Saccharomycespastorianus in following embodiment is numbered 28827, can obtain from ATCC.
Flos Rosae Rugosae fermentation oleo stock used in following embodiment is for carrying out sterilizing, centrifugal rear obtained supernatant successively to gained fermentation liquid after fermentation culture.
Embodiment 1, a kind of Flos Rosae Rugosae fermentation oleo stock and preparation method thereof:
1, the activation of strain:
Picking saccharomyces pastorianus (Saccharomycespastorianus) bacterium colony one ring, in fluid medium, puts into shaking table by actication of culture, obtains activating rear saccharomyces pastorianus bacterium liquid.
2, the purification of strain:
Bacterium liquid gradient dilution bed board after activation step 1 obtained, to obtain single bacterium colony, obtains saccharomyces pastorianus after purification.
3, the amplification culture of strain:
Strain inoculation after purification step 2 obtained, in YPD culture medium (pH value is 7.0), is cultivated in the shaking table of 25 DEG C, and as OD value=0.5-1.0, (strain is in logarithmic (log) phase, and concentration is 10 to obtain zymocyte bacterium liquid
5-10
8cFU/ml).
4, the acquisition of Flos Rosae Rugosae fermentation oleo stock:
Zymocyte bacterium liquid 15ml step 3 obtained is inoculated in the Flos Rosae Rugosae dry powder of 12g and the water of 350g, obtains fermentation system (in fermentation system, the proportioning of strain total amount and Flos Rosae Rugosae, water is 15ml:12g:350g); Ferment fermentation system in 35 DEG C of shaking tables 24h, obtains tunning; By tunning sterilizing 20min at 115 DEG C, make bacterium inactivation, obtain the tunning after sterilizing; By the centrifugal 20min under 4000r/min, centrifugal radius are the condition of 9cm of the tunning after sterilizing, abandon precipitation, collect supernatant, be Flos Rosae Rugosae fermentation oleo stock.
Embodiment 2, Flos Rosae Rugosae fermentation oleo stock are as the application in cosmetics:
One, the character of Flos Rosae Rugosae proferment pulp cosmetic:
Being fermented by preparation-obtained for embodiment 1 Flos Rosae Rugosae, oleo stock outward appearance is thick liquid, color is faint yellow to brown color.PH value 5.2-6.8, viscosity 200-500cP, solubility is admittedly containing thing content 1.5-5.0%, and total plate count is less than 50CFU/ml, detects without pathogenic bacterium.According to hygienic standards for cosmetics GB7916-87, cosmetics total number of bacteria is not higher than 1000CFU/ml, so this fermented product extract meets cosmetics quality requirement.
Carry out component analysis to this Flos Rosae Rugosae fermentation oleo stock, protein detection method is with reference to GB5009.5-2010; Crude polysaccharides detection method is with reference to GB/T5009.8-2008; Flavone detection method is with reference to GB/T5009.124-2003; Total phenols detection method is with reference to GB/T8313-2008, and acquired results is as follows:
The Flos Rosae Rugosae fermentation oleo stock that the present invention obtains is containing protein 7.91g/kg, crude polysaccharides 12.97g/kg, total flavones 0.25g/kg (in rutin), total phenols 0.85g/kg.
Two, the safety of Flos Rosae Rugosae fermentation oleo stock detects:
Human body patch test is mainly used for the zest detecting cosmetics finished product or raw material.The present invention carries out human closed formula patch test to the Flos Rosae Rugosae fermentation oleo stock that embodiment 1 obtains, and is intended to assess its potential skin irritation.
1, subjects:
Select suitable volunteer 30 people, the range of age is at 18-60 year Stochastic choice.
2, test method:
If solid sample or semisolid sample, then take 0.020g-0.025g be placed in speckle examination device for subsequent use; If fluid sample, then measure 0.025mL-0.2mL and drip on filter paper, then filter paper is placed in speckle examination device.The present invention adopts the latter, and each sample all arranges blank, and in contrast speckle examination device hole, add the sample solvent with sample equivalent, as distilled water or olive oil, the present invention adopts distilled water.
Human body back is elected at test position as, utilizes non-irritating adhesive tape speckle to be tried device and fixedly sticks in experimenter back.Test period continues 24h.In order to the accurate, credible of result of the test and science, test period volunteer as requested, speckle examination device can not be removed, recipient site also can not be made to contact water.Remove speckle examination device after 24h, after leaving standstill 30min, wait for that impression disappears, observe the reaction of skin.If result of the test is negative, then 24h and 48h after patch test is needed to observe once respectively again.
3, result of the test:
Patch test results is as shown in table 1:
Wherein, * "-"=negative reaction;
One-level untoward reaction: " ± "=suspicious reaction: only have faint erythema;
Secondary untoward reaction: "+"=weak positive reaction (erythematous response): erythema, infiltration, edema, can pimple be had;
Three grades of untoward reaction: " ++ "=strong positive reaction (herpes reaction): erythema, infiltration, edema, pimple, herpes, reaction can exceed tested district;
Level Four untoward reaction: " +++ "=extremely strong positive reaction (reaction of amalgamation herpes): obviously erythema, severe infiltration, edema, amalgamation herpes, reaction exceeds tested district.
According to cosmetics health specification 2007, human body patch test criterion is: occur in 30 routine experimenters that the number of 1 grade of skin adverse reaction is more than 5 examples, or the number of secondary skin adverse reaction is more than 2 examples, or when there is any 1 example more than three grades or three grades skin adverse reactions, then judge that tested material has untoward reaction to human body, otherwise, be then considered as having no adverse reaction to human body.
As can be seen from Table 1: the Flos Rosae Rugosae fermentation oleo stock that embodiment 1 obtains produces 1 routine suspicious reaction, and after 24h, this routine suspicious loss for reaction, illustrates that Flos Rosae Rugosae fermented product provided by the invention all has safety, can not bring untoward reaction to human body.
The patch test results of the Flos Rosae Rugosae fermentation oleo stock that table 1, embodiment 1 obtain
Three, the molecular size range of Flos Rosae Rugosae fermentation oleo stock detects:
Molecular size range checks the key factor whether can carrying out Cutaneous permeation.The present invention measures the molecular size range of Flos Rosae Rugosae fermentation oleo stock prepared by embodiment 1 by high performance liquid chromatography (HPLC).Concrete steps are as follows:
1, the reagent of test and the preparation of solution:
(1) mobile phase: acetonitrile: water: trifluoroacetic acid=30:70:0.1 (v/v/v).
(2) sample preparation: the sample take mobile phase as solvent compound concentration being 5mg/ml, then for sample introduction after filtering with microporous membrane (0.45 μm).
(3) preparation of standard sample: by bovine serum albumin (Mw67000), vitamin B
12(Mw1335), oxidized form of glutathione (Mw614) is made into mixed mark, and the concentration of often kind of material is 5mg/ml.
2, test method:
Standard substance by three kinds of known molecular amounts: bovine serum albumin (Mw67000), vitamin B
12(Mw1335) and oxidized form of glutathione (Mw614) by high performance liquid chromatograph (Waters company) reference literature " Wang Chengzhong etc. the research of polypeptide molecular weight distribution in protease hydrolyzed wheat germ protein and enzymolysis solution. grain processing; 2012:37 (2) " in method carry out HPLC analysis, draw the standard curve of molecular weight logarithm and elution time.According to gel column permeation chromatography principle, the material that polypeptide molecular weight is large is first by eluting out, and the logarithm of elution time and molecular weight is linear.After the same method the Flos Rosae Rugosae of preparation in embodiment 1 fermentation oleo stock is carried out HPLC analysis again, obtain the elution time of respective peaks, elution time is substituted into standard curve, the molecular size range of Flos Rosae Rugosae fermentation oleo stock prepared by embodiment 1 can be obtained.
Chromatographic condition: chromatographic column: TsKgel2000SWXL300mm × 7.8mm; Determined wavelength: UV280nm; Flow velocity: 1ml/min; Column temperature: 30 DEG C.
3, result of the test:
Molecular weight and the elution time of standard substance are as shown in table 2 below, are figure and obtain standard curve: y=-2.4738x+17.656, R according to the logarithm of molecular weight and elution time
2=0.9998.Wherein y is elution time (min); X is molecular weight logarithm.
The molecular weight of table 2, standard substance and elution time
Experimental group 1 prepare Flos Rosae Rugosae fermentation oleo stock HPLC collection of illustrative plates as shown in Figure 1: as can be seen from Figure 1: Flos Rosae Rugosae fermentation oleo stock collection of illustrative plates in mainly comprise 1 component peaks, elution time is 11.69min, elution time is substituted into standard curve: y=-2.4738x+17.656, the molecular weight calculating peak is 259.23Da, and accounts for 96.85% according to calculated by peak area Flos Rosae Rugosae fermentation oleo stock.It is generally acknowledged that molecular weight is that the bioactive peptide composition of about 500Da is more easily absorbed by the skin, so there is absorbable active polypeptide in Flos Rosae Rugosae fermented product.
Four, the antioxygenic property of Flos Rosae Rugosae fermentation oleo stock detects:
DPPH is a kind of organic free radical of early stage synthesis, be commonly used to the hydrogen supply capacity assessing polyphenoils, it is highly stable in organic solvent, in purple, and have a characteristic absorption peak at 517nm place, when running into free radical scavenger, the lone pair electrons of DPPH are paired and make it fade, and namely diminish at the light absorption value of maximum absorption wave strong point.Therefore, the change by measuring light absorption value carrys out the elimination effect of assess sample to DPPH free radical.
Getting the Flos Rosae Rugosae fermentation oleo stock that not commensurability embodiment 1 prepares is dissolved in deionized water, the Flos Rosae Rugosae fermentation oleo stock liquid to be measured of obtained 0.63%, 1.25%, 2.5%, 5% and 10% volumn concentration.
The specific experiment step of DPPH free radical scavenging experiment is:
(1) liquid to be measured and 2 × 10 of equal-volume (being generally 3mL) is got
-4the DPPH solution mixing (A of mol/L
1pipe);
(2) isopyknic dehydrated alcohol (determinand solvent) and 2 × 10 is got
-4the DPPH solution mixing (A of mol/L
2pipe);
(3) isopyknic dehydrated alcohol is got and liquid to be measured mixes (A
3pipe);
(4), after reacting 30min, under 517nm, A is surveyed
1, A
2, A
3pipe absorbance.
Clearance rate computing formula is: clearance rate (%)=[(A
2+ A
3)-A
1]/A
2
With the volumn concentration of Flos Rosae Rugosae fermentation oleo stock liquid to be measured for abscissa, clearance rate is vertical coordinate, makes Flos Rosae Rugosae fermentation oleo stock liquid to be measured to DPPH free radical scavenging effect curves, sees Fig. 2.
As shown in Figure 2: gained Flos Rosae Rugosae fermentation oleo stock is to the IC50=0.56% of DPPH scavenging action, illustrating that Flos Rosae Rugosae fermentation oleo stock has stronger oxidation resistance can scavenging free radicals, promote cellular metabolism, strengthen cell viability, improve the 26S Proteasome Structure and Function of body, improve body vitality, thus delay cell senescence, play its antidotal effect.
Five, the white-skinned face function analysis of Flos Rosae Rugosae fermentation oleo stock:
Tryrosinase is melanogenic key enzyme, which control the forming process of melanocyte, and the deposition of its level of activity to pigment plays a major role.Many whitenings of selling in the market, speckle dispelling product are all reach whitening function with restraint of tyrosinase, therefore are evaluate the leading indicator of whitening cosmetic to the power of tyrosinase inhibitory action.
The whitening function of assess sample is carried out by the impact of working sample on tryrosinase, for testing sample, its inhibition to tyrosinase activity is measured with 10% Flos Rosae Rugosae fermentation oleo stock (adopting deionized water to be diluted to volumn concentration for 10% embodiment 1 gained Flos Rosae Rugosae fermentation oleo stock).
Concrete grammar is as follows:
Solution is configured by table 3:
Table 3, solution preparation list
| Unit (mL) | C 1 | C 2 | T 1 | T 2 |
| TYR | 2 | 2 | 2 | 2 |
| Sample | 0 | 0 | 2 | 2 |
| PBS | 4 | 5 | 2 | 3 |
| Tryrosinase | 1 | 0 | 1 | 0 |
| Cumulative volume | 7 | 7 | 7 | 7 |
Note: C
1and T
1add 1mL tryrosinase, enzyme is lived as 100U/mL.
(1) C
2after pipe prepares and shakes up, heating in water bath 10min in 37 DEG C of water-baths, returns to zero under wavelength 475nm.
(2) C
1pipe solution prepares and shakes up, and after 37 DEG C of water-bath 10min, adds tryrosinase 1ml, continues water-bath 10min, measures C
1absorbance.
(3) by (1) (2) same method, with T
2zeroing measures T
1absorbance.
(4) calculation sample is to the maximum inhibition T (%) of tryrosinase.T(％)＝(C
1-T
1)/C
1×100％
Experiment proof 10% Flos Rosae Rugosae fermentation oleo stock is 65.42% to the suppression ratio of tyrosinase activity, the arbutin of 1% is to the suppression ratio 99.52% of tyrosinase activity, see Fig. 3, can learn from Fig. 3: the whitening effect of Flos Rosae Rugosae fermentation oleo stock is equivalent to the effect of the arbutin of 0.657%, has certain white-skinned face function.
Claims (10)
1. a preparation method for Flos Rosae Rugosae fermentation oleo stock, comprises the steps: Flos Rosae Rugosae dry powder, water and zymocyte to be mixed to get initial system, and carries out fermentation culture to initial system, obtains Flos Rosae Rugosae fermentation oleo stock.
2. preparation method as claimed in claim 1, is characterized in that: the proportioning of described zymocyte, described Flos Rosae Rugosae dry powder and water is (15-25) ml:(7-15) g:350g, described zymocyte concentration is 10
5-10
8cFU/ml.
3. preparation method as claimed in claim 1 or 2, is characterized in that: the temperature of described fermentation culture is 25-38 DEG C, and the time is 24-32h.
4. the preparation method according to any one of claim 1-3, is characterized in that: described zymocyte is yeast, is specially saccharomyces pastorianus bacterium Saccharomycespastorianus;
Rose Blume Getrocknete alabastrum taken from by described Flos Rosae Rugosae dry powder.
5. the preparation method according to any one of claim 1-4, is characterized in that: the order number of described Flos Rosae Rugosae dry powder is 20-50 order.
6. the preparation method according to any one of claim 1-5, is characterized in that: in above-mentioned preparation method, also comprises and carries out sterilizing, centrifugal, the step of getting supernatant successively to gained fermentation liquid after described fermentation culture.
7. preparation method as claimed in claim 6, it is characterized in that: the condition of described sterilizing is as follows: sterilising temp is 115-121 DEG C, sterilization time is 15-25min;
Described centrifugal condition is as follows: centrifugal rotational speed is 3800-4200r/min, centrifugation time is 15-30min, and centrifugal radius is 9cm.
8. the preparation method according to any one of claim 1-7 and obtain Flos Rosae Rugosae fermentation oleo stock.
9. Flos Rosae Rugosae according to claim 8 fermentation oleo stock prepare the application that has in the cosmetics of following at least one function or directly as the application had in the cosmetics of following at least one function:
1) whitening; 2) defying age; 3) DPPH free radical is removed; 4) restraint of tyrosinase.
10. application according to claim 9, is characterized in that: described cosmetics are facial film, essence or cosmetic water.
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