CN111588677A

CN111588677A - Barley tremella fermentation raw pulp and preparation method and application thereof

Info

Publication number: CN111588677A
Application number: CN202010503554.9A
Authority: CN
Inventors: 方祥铭; 方晓薇
Original assignee: Shanghai Biotruly Biotechnology Co ltd
Current assignee: Shanghai Biotruly Biotechnology Co ltd
Priority date: 2020-06-05
Filing date: 2020-06-05
Publication date: 2020-08-28
Anticipated expiration: 2040-06-05
Also published as: CN111588677B

Abstract

The invention provides barley tremella fermentation raw pulp and a preparation method and application thereof, wherein the preparation method comprises the following steps: mixing the mixed powder of barley malt and tremella, water and zymocyte liquid to obtain an initial system, fermenting to obtain fermentation liquor, sterilizing and centrifuging the fermentation liquor to obtain supernatant, namely the barley tremella fermentation raw stock. The invention adopts the saccharomycetes to ferment the barley malt and the tremella, retains all functional components and activity of the barley malt and the tremella, and avoids loss of active components caused by an extraction method.

Description

Barley tremella fermentation raw pulp and preparation method and application thereof

Technical Field

The invention relates to a fermentation product, in particular to a fermentation product with a skin care function.

Background

The barley malt is spindle-shaped, 8-12 mm long and 3-4 mm in diameter. The surface is light yellow, and the back surface is surrounded by the lemma and has 5 veins; the ventral surface is the palea envelope. After removing the inner and outer seeds, the ventral surface has 1 longitudinal furrow; the radicle of the base part grows a bud and a fibrous root, and the bud is in a needle-shaped strip shape and is about 5mm long. The fiber has a plurality of strands, and is slender and bent. Hard, white cross section and powdery. Light smell, slightly sweet taste. Tremella is fruiting body of Basidiomycota fungus Tremella, and comprises more than 10 thin and multi-fold flat leaves. The white fungus fruiting body is pure white to milk white, generally in chrysanthemum or cockscomb shape, soft, white, semitransparent and elastic.

So far, in the skin care product manufacturing industry, most skin care products are not fermented skin care products, are mainly prepared by the traditional process and are most common by cooking. For example, after the medicinal materials are cooked, the medicinal materials are fished out and discarded, and ancient people drink soup, and when people use the soup on cosmetics, the soup can be filtered and added with preservative for direct use. However, the traditional process is limited by ancient thinking and is difficult to break through, and even the traditional process is used for extracting plant extracts in a plurality of skin care products, so that not only the medicinal materials are seriously consumed and the sustainable development is not realized, but also the obtained substances have only limited types and quantity and often have unknown irritability, and therefore, new technology should be considered. At present, many fermentation allegedly skin care products are actually prepared by adding only a little fermentation source components into the formula of the skin care products, so that the content of the fermentation products in the formula is only 1-2%. Many products are mainly prepared from traditional chemical substances, and various types of added plant extracts are very small in content, so that poor effects are obtained, and the appearance state is transparent and clear, but has no significance to consumers. Chinese patent (CN109330929A) discloses a preparation method of a tremella fermentation extract and application of the tremella fermentation extract in cosmetics, and particularly discloses a tremella extract refining solution prepared by adopting a mixture of tremella and water through the steps of high-temperature fermentation, ultrahigh-speed shearing, low-temperature fermentation and the like, and a tremella extract mask prepared by the method. The tremella polysaccharide molecular weight is degraded through the cooperation of two-step fermentation and high-speed shearing process, and absorption, water replenishing and moisture keeping on the surface of human skin are facilitated. The process is complex, the most important of fermentation products is the unified and pure environment, new bacteria are inevitably introduced by frequently replacing equipment after fermentation in the patent, the whole process is difficult to control, the process can only stay in a laboratory stage, and the real industrial production is difficult.

The statements in the background section are merely prior art as they are known to the inventors and do not, of course, represent prior art in the field.

Disclosure of Invention

The invention aims to provide a fermentation raw stock prepared from barley malt and tremella aiming at one or more problems in the prior art;

the invention also aims to provide a preparation method of the fermentation raw stock prepared by the method;

the invention also aims to provide application of the fermentation raw pulp.

The purpose of the invention is realized by the following technical scheme:

a barley tremella fermented raw stock is prepared by fermenting substances including barley malt and tremella.

According to one aspect of the invention, the barley tremella fermentation raw pulp contains 1-10mg/ml of protein, 10-40mg/ml of crude polysaccharide, 0.2-1.0mg/ml of total flavone and 0.1-0.8mg/ml of total phenol

According to one aspect of the invention, the mass ratio of the barley malt to the tremella is (2-4): (2-4); preferably, the mass ratio of the barley malt to the tremella is 1: 1.

according to one aspect of the invention, the fermentation is carried out using yellow wine yeast.

A skin care product comprises the barley tremella fermentation raw stock.

The barley white fungus fermentation raw pulp is applied as a skin care product.

A preparation method of barley tremella fermented raw stock comprises mixing barley malt, tremella mixed powder, water and zymocyte liquid to obtain an initial system, fermenting to obtain fermentation liquid, sterilizing the fermentation liquid, and centrifuging to obtain supernatant, i.e. barley tremella fermented raw stock.

according to one aspect of the present invention, the mesh number of the barley malt and tremella mixed powder is 20-50 mesh, preferably 50 mesh.

According to one aspect of the invention, the mixed powder of barley malt and tremella: the proportion of water is (5-20) g (100-300) mL.

According to one aspect of the invention, the mixed powder of barley malt and tremella: the mass ratio of water is 10g:200 mL.

According to one aspect of the invention, the concentration of the zymogen liquid is 10⁵-10⁸CFU/ml。

According to one aspect of the invention, the ratio of the zymophyte liquid to the mixed powder of the barley malt and the tremella is (10-40) ml: (5-20) g.

According to one aspect of the invention, the ratio of the zymophyte liquid to the mixed powder of barley malt and tremella is 20 ml: 10 g.

The invention also provides a preparation method of the barley tremella fermentation raw stock, and the fermentation temperature of the fermentation is 30-45 ℃, and is preferably 37 ℃.

According to one aspect of the invention, the fermentation time of the fermentation is 10-50h, preferably 36 h.

According to one aspect of the invention, the centrifugation is carried out at 4000-12000r/min for 15-30 min;

according to one aspect of the invention, the centrifugation is performed at 4500r/min for 20 min.

According to one aspect of the invention, the centrifugation radius of the centrifugation is 9-12 cm.

According to one aspect of the invention, the fermentation bacteria are yellow wine yeast.

According to one aspect of the invention, the zymophyte is pretreated in advance, and the pretreatment process comprises the following steps:

activating strains: putting the bacterial colony of the zymocyte into a liquid culture medium, and then putting the culture medium into an incubator to activate the strain;

and (3) strain purification: the activated strains are subjected to gradient dilution and plating so as to obtain single colonies; and

expanding culture of strains: inoculating the strain to be used into corresponding liquid culture medium, and culturing in 30-45 deg.C incubatorWhen the OD value is 0.5-1.0, the strain is in logarithmic phase, i.e. the proper inoculation concentration is 10⁵-10⁸CFU/ml; preferably, the medium is a yeast medium.

The barley tremella fermentation raw pulp provided by the invention can be used as an effective component in a skin care product, can also be independently used as a skin care product, and has an antioxidant function and a whitening function.

The preparation method of the fermented raw juice by using the barley malt and the tremella has the following advantages:

(1) the invention adopts the saccharomycetes to ferment the barley malt and the tremella, retains all functional components and activity of the barley malt and the tremella, and avoids loss of active components caused by an extraction method.

(2) The fermentation method adopted by the invention does not add any organic reagent in the process of extracting the effective components of barley malt and tremella, has mild fermentation temperature and fermentation pH, does not damage the structure of the active components of the plants, and keeps the natural activity of the plants.

(3) After the fermentation is finished in the method, chemical components such as essence and the like are not added into the fermentation raw stock, so that the safety of the product to human bodies is ensured.

(4) The barley tremella fermented raw pulp provided by the invention is rich in active substances which have tyrosinase activity inhibition and melanin synthesis inhibition, so that the barley tremella fermented raw pulp has a strong skin whitening function and has a synergistic effect with components in a fermentation filtrate.

Drawings

FIG. 1 is a graph showing the scavenging effect of barley white fungus fermentation raw juice on hydroxyl radicals in an embodiment of the present invention;

FIG. 2 is a bar graph of the inhibition rate of barley white fungus fermentation raw pulp on tyrosinase activity and arbutin on tyrosinase activity in the embodiment of the invention;

FIG. 3 is a graph of long-term skin moisture data of barley white fungus fermentation broth in an embodiment of the present invention;

FIG. 4 is data of long-term transdermal water evaporation of primary fermented barley tremella pulp in an embodiment of the present invention.

Detailed Description

In the following, only certain exemplary embodiments are briefly described. As those skilled in the art will recognize, the described embodiments may be modified in various different ways, all without departing from the spirit or scope of the present invention. Accordingly, the drawings and description are to be regarded as illustrative in nature, and not as restrictive.

The following disclosure provides many different embodiments or examples for implementing different features of the invention. To simplify the disclosure of the present invention, the components and arrangements of specific examples are described below. Of course, they are merely examples and are not intended to limit the present invention. Furthermore, the present invention may repeat reference numerals and/or letters in the various examples, such repetition is for the purpose of simplicity and clarity and does not in itself dictate a relationship between the various embodiments and/or configurations discussed. In addition, the present invention provides examples of various specific processes and materials, but one of ordinary skill in the art may recognize applications of other processes and/or uses of other materials.

The following description of the preferred embodiments of the present invention is provided for the purpose of illustration and description, and is in no way intended to limit the invention.

In the following examples, the fermentation tubes used were pretreated in advance, and the pretreatment process included:

expanding culture of strains: inoculating the strain to corresponding liquid culture medium, culturing at 30-45 deg.C in culture box until OD value is 0.5-1.0, and the strain is in log phase, i.e. proper inoculation concentration is 10⁵-10⁸CFU/ml; the culture medium is a yeast culture medium.

The specific process of strain activation and purification is the conventional technology.

Example 1:

the concentration of zymophyte in the culture medium is 10⁵-10⁸20mL of CFU/mL zymocyte liquid is inoculated into 50 meshes of 10g of barley malt and tremella mixed powder (the mass ratio of barley malt to tremella is 1: 1) and 200g of water, after the mixture is cultured for 36 hours in a 37 ℃ incubator, the obtained zymotic fluid is sterilized under high pressure at 121 ℃, sterilized under high pressure for 15min to inactivate the bacteria, the sterilized zymotic fluid is centrifuged for 20min at 4500r/min and the centrifugation radius is 9cm, and the supernatant is collected, namely the barley tremella zymogen primary pulp cosmetic provided by the invention.

The barley and tremella fermented puree cosmetic prepared in the embodiment is viscous liquid in appearance and is colorless, transparent and brown in color. The pH value is 4.5-6.8, the viscosity is 100-800cP, the content of soluble solid is 1.0-5.0%, the total number of colonies is less than 50CFU/ml, and no pathogenic bacteria are detected. Meets the quality requirement of cosmetics.

Performing component analysis on the barley and tremella fermented raw pulp cosmetic: the barley tremella fermentation protoplasm cosmetic prepared by the invention contains 1-10mg/ml of protein, 10-40mg/ml of crude polysaccharide, 0.2-1.0mg/ml of total flavone and 0.1-0.8mg/ml of total phenol.

Analyzing the antioxidant effect of the barley and tremella fermentation raw pulp:

referring to table 1, using 10mL, test tubes were set up with 3 parallel tubes for each concentration tested sample tube (T) for each sample, sample background (T0), blank tube (C), control tube (C0).

Precisely transferring 3mL of ferrous sulfate solution into a test tube, sequentially adding 3mL of hydrogen peroxide solution and 3mL of salicylic acid solution, uniformly mixing, and heating in a water bath at 37 ℃ for 15 min; respectively adding 1mL of sample solutions with different concentrations as T, and continuously heating for 15 min; the method is the same as above, the blank tube (C) and the control tube (C0) replace the sample with the same volume of distilled water, and the sample background (T0) and the control tube (C0) replace the hydrogen peroxide reaction system with the same volume of distilled water. The absorbance was measured at 510 nm.

TABLE 1 sample addition requirement

A curve of action of barley tremella fermentation raw stock on scavenging hydroxyl free radical is shown in figure 1.

As can be seen from figure 1, the barley tremella protoplasm has obvious effect of removing hydroxyl radicals, and the capacity of removing the hydroxyl radicals is gradually enhanced along with the increase of the concentration of the barley tremella protoplasm. The 30% barley tremella original pulp can remove about 38.67% hydroxyl free radical, and IC50 is 38.074%.

And (3) analyzing the whitening efficacy of the barley and tremella fermented raw juice:

tyrosinase is a key enzyme in melanogenesis, which controls the process of melanogenesis, and its degree of activity plays a major role in pigment deposition. Many whitening and freckle-removing products sold in the market at present achieve the whitening effect by inhibiting tyrosinase, so the strength of the tyrosinase inhibition effect is a main index for evaluating whitening cosmetics.

The whitening function of the sample is evaluated by measuring the influence of the sample on tyrosinase, and the specific method comprises the following steps:

prepare the solution as in table 2:

TABLE 2 solution preparation List

Unit (mL)	C₁	C₂	T₁	T₂
					L-tyrosine	2	2	2	2
Sample (I)	0	0	2	2
					PBS	4	5	2	3
Tyrosinase enzyme	1	0	1	0
					Total volume	7	7	7	7

Note: c₁And T₁Adding 1mL of tyrosinase, and enabling the enzyme activity to be 100U/mL.

(1) After the C2 pipe is well prepared and shaken up, the mixture is heated in a water bath kettle at 37 ℃ for 10min, and the zero setting is carried out under the wavelength of 475 nm.

(2) Mixing the solution in a C1 tube, shaking, performing water bath at 37 ℃ for 10min, adding 1ml of tyrosinase, continuing the water bath for 10min, and determining the absorbance value of C1.

(3) The absorbance value of T1 was measured by zeroing with T2 in the same manner as in (1) and (2).

(4) The inhibition rate T (%) of the sample on tyrosinase activity was calculated. T (%) - (C1-T1)/C1X 100%

The 1% arbutin is a positive control of a tyrosinase activity inhibition experiment, and the inhibition rate of the arbutin on the tyrosinase activity is 75%. 30% of barley tremella protoplasm can inhibit 25.56% of tyrosinase activity, so the barley tremella aromatic protoplasm has the effect of inhibiting the tyrosinase activity and has a certain whitening effect.

Moisture retention efficacy analysis of barley white fungus fermentation raw pulp:

the moisture retention capacity is an important index for judging the performance of the product. Skin moisture content can be increased after use of the product and can remain higher than unapplied for extended periods of time, which is recognized as a good moisturizing capability. Can moisten and soften skin.

The experimental method comprises the following steps: first, the measurement area of the left arm and the right arm of the subject (3 people) is made, the area of the area is 3cmX3cm, and the sample is measured and is applied in a single smearing mode according to the dosage of (2.0 +/-1.0) mg/cm 2. The initial values for each test area were determined (before sample application, then after a set time to determine the moisture content and water loss of the test and control areas. volunteers selected and tested according to the methods QL-SOP-YF-3-01 and QL-SOP-YF-3-02.

As shown in fig. 3: compared with the uncoated blank control area, the barley white fungus fermentation raw juice has obvious advantage in long-term moisture retention; the skin moisture content of the test subject is always higher than that of the uncoated blank control area after 1 day, 7 days and 14 days of barley tremella fermentation primary pulp fermentation.

As shown in fig. 4: compared with the uncoated blank control area, the barley tremella protoplasm can reduce the water loss of the skin through the epidermis and keep moisture for a long time. After the test subject uses the barley tremella primary pulp for 14 days, the water locking capacity is increased by 12.5%. The skin's ability to prevent water evaporation was consistently greater than the uncoated placebo area, i.e. the amount of water evaporation was significantly lower, indicating a sustained increase in barrier capacity.

And (3) evaluating the safety of the barley tremella fermentation primary pulp:

the patch test is mainly used for detecting the irritation of the final cosmetic product or raw material. The invention carries out a human body closed patch test on the barley and tremella fermentation protoplasm cosmetic and evaluates the potential skin irritation of the barley and tremella fermentation protoplasm cosmetic.

Suitable volunteers were selected for 30 persons, and were randomly selected in the age range of 18-60 years. 0.020g to 0.025g of solid or semi-solid sample is weighed into a plaque test device for use. The liquid sample, 0.2mL to 0.025mL, was dropped onto the filter paper sheet, which was then placed in the plaque tester. A blank control is set for each sample and an equal amount of sample solvent, such as distilled water or olive oil, is added to the control chamber.

The test part is selected as the back of a human body, and the spot tester is fixedly attached to the back of the testee by using a non-irritant adhesive tape. The test period lasted 36 h. In order to ensure the accuracy, credibility and scientificity of test results, the volunteers cannot remove the spot tester or make the tested part contact water according to the requirements during the test.

And removing the spot tester after 36h, standing for 30min, waiting for the indentation to disappear, and observing the reaction of the skin. If the test result is negative, the test needs to be observed once more at 36h and 48h after the patch test.

The patch test results are shown in table 3:

TABLE 3 Patch test results

"-" ═ negative reactions;

"±" ═ suspicious reaction: only faint erythema;

"+" ═ weak positive reaction (erythema reaction): erythema, infiltration, edema, and possibly pimples;

"+ +", strong positive reaction (herpes response); erythema, infiltration, edema, pimples, herpes; the reaction may be beyond the test area;

"+ + + +" -very strong positive reaction (fusogenic herpes response); obvious erythema, severe infiltration, edema, and fusional herpes; the reaction goes beyond the test area.

As can be seen from Table 3, the barley white fungus fermentation broth provided in example 1 produced no more than 1 of grade 2 reactions in the experiment. The barley tremella fermentation raw stock is judged to be safe and does not bring adverse reaction to human bodies.

Example 2:

the concentration of zymophyte in the culture medium is 10⁵-10⁸Inoculating 40mL of CFU/mL zymocyte liquid into 50 meshes of 5g of barley malt and tremella mixed powder (the mass ratio of barley malt to tremella is 1: 2) and 300g of water, culturing for 50 hours in a 30 ℃ incubator, then carrying out high-pressure sterilization at 121 ℃ on the obtained zymotic fluid, carrying out high-pressure sterilization for 15min to inactivate bacteria, centrifuging the sterilized zymotic fluid for 15min under the conditions of 12000r/min and 12cm of centrifugal radius, and collecting supernatant fluid, namely the barley tremella zymocyte raw stock cosmetic provided by the invention.

Example 3:

the concentration of zymophyte in the culture medium is 10⁵-10⁸Inoculating 10mL of CFU/mL zymocyte liquid into 50 meshes of 20g of barley malt and tremella mixed powder (the mass ratio of barley malt to tremella is 2: 1) and 100g of water, culturing for 10 hours in a 45 ℃ incubator, then carrying out high-pressure sterilization at 121 ℃ on the obtained zymotic fluid, carrying out high-pressure sterilization for 15min to inactivate bacteria, centrifuging the sterilized zymotic fluid for 30min under the conditions of 4000r/min and 10cm of centrifugal radius, and collecting supernatant fluid, namely the barley tremella fermented protoplasm cosmetic provided by the invention.

The methods and results of analyzing the components of the products of examples 2 and 3 are the same as those of example 1, and therefore will not be described again.

The products of examples 2 and 3 were analyzed for antioxidant effect, whitening effect and moisturizing effect in the same manner as in example 1, and the results were substantially the same as in example 1. And therefore will not be reiterated.

The safety evaluation methods and results of the products of examples 2 and 3 are the same as those of example 1, and therefore will not be described again.

Finally, it should be noted that: although the present invention has been described in detail with reference to the foregoing embodiments, it will be apparent to those skilled in the art that changes may be made in the embodiments and/or equivalents thereof without departing from the spirit and scope of the invention. Any modification, equivalent replacement, or improvement made within the spirit and principle of the present invention should be included in the protection scope of the present invention.

Claims

1. A barley tremella fermentation raw stock is characterized by being prepared by fermenting substances including barley malt and tremella.

2. The barley tremella fermentation puree according to claim 1, wherein the barley tremella fermentation puree contains 1-10mg/ml protein, 10-40mg/ml crude polysaccharide, 0.2-1.0mg/ml total flavonoids, and 0.1-0.8mg/ml total phenols;

preferably, the pH value of the barley tremella fermentation protoplasm is 4.5-6.8.

3. The barley tremella fermentation puree according to claim 1, wherein the mass ratio of barley malt to tremella is (2-4): (2-4); preferably, the mass ratio of the barley malt to the tremella is 1: 1.

4. the barley tremella fermented puree according to claim 1, wherein the fermentation is performed using yellow wine yeast.

5. A skin care product comprising the barley tremella fermented puree of any one of claims 1 to 3.

6. The use of barley tremella fermented puree according to any one of claims 1 to 3 as a skin care product.

7. A preparation method of barley tremella fermentation protoplasm is characterized by comprising the following steps: mixing the mixed powder of barley and tremella, water and zymocyte liquid to obtain an initial system, fermenting to obtain fermentation liquor, sterilizing and centrifuging the obtained fermentation liquor to obtain supernatant, namely the barley tremella fermentation protoplasm.

8. The method for preparing barley tremella fermentation puree according to claim 7, wherein the method comprises the following steps: the preparation method is characterized in that the mass ratio of barley malt to tremella in the mixed powder is (2-4): (2-4); preferably, the mass ratio of the barley malt to the tremella is 1: 1;

preferably, the mesh number of the mixed powder of the barley malt and the tremella is 20-50 meshes, and more preferably is 50 meshes;

preferably, the mixed powder of barley malt and tremella: the proportion of water is (5-20) g (100-300) mL; further preferably, the mixed powder of barley malt and tremella: the mass ratio of water is 10g:200 mL;

preferably, the concentration of the zymogen liquid is 10⁵-10⁸CFU/ml；

Preferably, the proportion relation between the zymocyte liquid and the mixed powder of the barley malt and the tremella is (10-40) ml: (5-20) g; further preferably, the proportion relationship between the zymocyte liquid and the mixed powder of the barley malt and the tremella is 20 ml: 10 g.

9. The method for preparing barley tremella fermentation puree according to claim 7, wherein the method comprises the following steps: the fermentation temperature of the fermentation is 30-45 ℃, and preferably 37 ℃;

preferably, the fermentation time of the fermentation is 10-50h, preferably 36 h;

preferably, the centrifugation is carried out for 15-30min at 4000-12000 r/min;

preferably, the centrifugation is performed at 4500r/min for 20 min;

preferably, the centrifugation radius of the centrifugation is 9-12 cm.

10. The method for preparing barley tremella fermentation puree according to claim 7, wherein the method comprises the following steps: the fermentation bacteria are yellow wine yeast;

preferably, the zymophyte is pretreated in advance, and the pretreatment process comprises the following steps:

expanding culture of strains: inoculating the strain to corresponding liquid culture medium, culturing at 30-45 deg.C in culture box until OD value is 0.5-1.0, and the strain is in log phase, i.e. proper inoculation concentration is 10⁵-10⁸CFU/ml; preferably, the medium is a yeast medium.