CN113648262A

CN113648262A - Lily leaf and cherry fermented product for cosmetic and preparation method thereof

Info

Publication number: CN113648262A
Application number: CN202110990675.5A
Authority: CN
Inventors: 张佳婵; 王昌涛; 李萌; 赵丹; 王冬冬; 程文静
Original assignee: Beijing Technology and Business University
Current assignee: Beijing Technology and Business University
Priority date: 2021-08-26
Filing date: 2021-08-26
Publication date: 2021-11-16

Abstract

The invention provides a preparation method of a lily and acerola cherry fermented product for cosmetics, which comprises the following steps: mixing a proper amount of lily, acerola powder and water, sterilizing, and cooling to obtain a fermentation substrate, wherein the mass ratio of the lily to the water is 0.5-5: 100, and the mass ratio of the acerola powder to the water is 0.1-3: 100; the lily is dried lily, and the acerola cherry powder is an acerola cherry extract; and inoculating saccharomycetes and/or lactic acid bacteria to the fermentation substrate for fermentation treatment, and then sterilizing and separating to obtain the lily acerola cherry fermentation liquor. The method utilizes saccharomycetes and/or lactic acid bacteria to carry out liquid fermentation on the lily acerola, can extract active substances in lily, and realizes good fusion with the active substances of the acerola extract; especially, yeast is firstly used for fermentation, and then lactobacillus is used for secondary fermentation, so that the safety of the product is improved; can be used in cosmetics to improve oxidation resistance and antiaging property of cosmetics.

Description

Lily leaf and cherry fermented product for cosmetic and preparation method thereof

Technical Field

The disclosure belongs to the technical field of biological fermentation, and particularly relates to a lily and acerola fermented product for cosmetics and a preparation method thereof.

Background

Lily, also known as "Qiangshu", folium sennae, Shandan, Fanxian, Dahlian, Zhongting, Mo Rou, heavy box, Zhongmei, Lily garlic, Dashifu garlic, garlic sweet potato, evening primrose, etc., is a perennial herb bulbous plant of Lilium (the name: Lilium) in Liliaceae, originally produced in China, mainly distributed in northern hemisphere temperate regions such as east Asia, Europe, North America, etc., and has found at least 120 varieties globally, of which 55 varieties are produced in China. The bulb is rich in starch, and is edible and medicinal. The lily extract mainly contains steroidal saponins, lily polysaccharide and other components, and has the characteristics of reducing blood sugar, resisting oxidation, resisting tumors and fatigue, enhancing the immunity of the organism and improving the lymphocyte conversion rate; not only has wide clinical application, but also has great development prospect as the raw material for processing health products.

Acerola cherry (Acerola cherry) is a fruit of Acerola (Malpighiaceae) Acerola (Malpighia) Acerola (m.emarginata). The acerola cherry extract contains active components extracted from acerola cherry as raw materials, including protein, sugar, fruit acid, vitamins A, B1, B2, vitamin C, nicotinic acid, calcium, phosphorus, iron, etc.; has good anti-anemia, anti-fungus, anti-genetic toxicity and strong free radical scavenging ability, and can be used as natural antioxidant in food and cosmetic industry.

The natural active substance is extracted by water extraction, organic solvent extraction, ultrasonic extraction, microwave extraction, supercritical fluid extraction, microbial fermentation, etc., and the natural active substance is extracted from Bulbus Lilii by organic solvent extraction. How to screen a method more favorable for extracting the active substances is still a technical problem faced by developers in the field. Fermentation refers to a process in which people prepare microbial cells themselves, or direct metabolites or secondary metabolites, by virtue of the life activities of microorganisms under aerobic or anaerobic conditions, and is widely used in the food industry, the biological and chemical industries. The extraction of nutrients by fermentation technology has been reported, but the extraction effect is very different according to the fermentation substrate and the fermentation process conditions. The method depends on researchers to continuously explore more fermentation substrate combinations and fermentation methods to prepare more products with better effects so as to meet the requirements of consumers.

In the prior art, both CN202010657142.0 (a traditional Chinese medicine fermented health food for improving sleep and a preparation method thereof) and CN202011315902.6 (a beauty and beauty probiotic fermented powder and a production method thereof) disclose schemes for preparing health foods by fermenting a plurality of medical and edible dual-purpose raw materials including lily with a plurality of probiotics. However, the raw material components in the scheme are complex, the preparation steps are multiple, and the obtained product is food, and the performance of the product for cosmetics is unknown. In addition, researchers prepare edible plant enzymes (sandiskei, and the like) by fermenting fresh lily and fermented sugar liquor at room temperature in a dark place according to the mass ratio of 3:5 [ J ] of organic acid and in-vitro antioxidant activity of the lily enzymes in the natural fermentation process, food and fermentation industry, 2019,45(22):39-45), explore the change rule of in-vitro antioxidant activity, organic acid composition and content of the lily enzymes in the natural fermentation process, and provide scientific basis for development and accurate preparation of lily enzyme products, but the scheme is also used for developing food and has unknown performance of cosmetics.

At present, no report of the application of lily and acerola cherry extract compound fermentation extract in the field of cosmetics exists. The applicant obtains a lily acerola cherry fermentation product for cosmetics and a preparation method thereof through research and exploration.

Disclosure of Invention

The following presents a simplified summary of the disclosure in order to provide a basic understanding of some aspects of the disclosure. It should be understood that this summary is not an exhaustive overview of the disclosure. It is not intended to identify key or critical elements of the disclosure or to delineate the scope of the disclosure. Its sole purpose is to present some concepts in a simplified form as a prelude to the more detailed description that is discussed later.

In view of the above defects of the prior art, the present disclosure aims to provide a lily and acerola cherry fermentation product for cosmetics and a preparation method thereof, wherein the preparation method is simple and mild, and the fermentation product has good safety, a certain anti-aging effect and good stability.

According to a first aspect of the present disclosure, there is provided a method for preparing a lily acerola cherry fermentate for cosmetics, comprising:

mixing a proper amount of dried lily, acerola powder (acerola cherry extract) and water, sterilizing, and cooling to obtain a fermentation substrate, wherein the mass ratio of the lily to the water is 0.5-5: 100 (such as 1:100, 1.5:100, 2:100, 2.5:100, 3:100, 3.5:100, 4:100, 4.5:100 and the like), and the mass ratio of the acerola powder to the water is 0.1-3: 100 (such as 0.3:100, 0.5:100, 1:100, 1.5:100, 2:100, 2.5:100, 2.8:100 and the like);

and inoculating saccharomycetes and/or lactic acid bacteria to the fermentation substrate for fermentation treatment, and then sterilizing and separating to obtain the lily acerola cherry fermentation liquor.

In the above preparation method, as a preferred embodiment, the acerola powder is an acerola extract obtained by a conventional hot water extraction method.

In the above preparation method, as a preferred embodiment, the dried lily is crushed to a particle size of 100 mesh.

In the above production method, as a preferred embodiment, the fermentation treatment includes: firstly, inoculating saccharomycetes to the fermentation substrate to carry out primary fermentation culture; and inoculating lactic acid bacteria for secondary fermentation culture after sterilization treatment.

In the above preparation method, as a preferred embodiment, the yeast is Saccharomyces cerevisiae (Saccharomyces cerevisiae) YWY-1, the preservation unit is china general microbiological culture collection center (CGMCC), the address is No. 3 of west road No.1 of north chen of the sunny region in beijing, the postal code is 100101, the preservation date is 2019, 03 and 27 days, and the preservation number is CGMCC No. 17452.

In the above preparation method, as a preferred embodiment, the lactic acid bacteria is at least one of the following species: lactobacillus delbrueckii subsp. bulgaricus, Lactobacillus buchneri, Bifidobacterium bifidum (Bifidobacterium bifidum). In practice, lactic acid bacteria may employ at least one of: lactobacillus delbrueckii subsp. bulgaricus (Lactobacillus delbruuchii subsp. bulgaricus) with the collection number of CGMCC 1.16075, available from CGMCC; lactobacillus buchneri (Lactobacillus buchneri) with preservation number of CGMCC 1.15607, available from CGMCC; bifidobacterium bifidum (CGMCC 1.5029) with preservation number can be purchased from CGMCC. More preferably, the lactic acid bacterium is Lactobacillus delbrueckii subsp.

In the above preparation method, as a preferred embodiment, the concentration of the yeast liquid used for inoculation is 10⁴-10⁹CFU/mL (e.g., 10)⁵CFU/mL、10⁶CFU/mL、10⁷CFU/mL、10⁸CFU/mL, etc.), the volume ratio of the yeast liquid to the fermentation substrate is 5% -20% (such as 8%, 10%, 12%, 15%, 18%, etc.).

In the above-mentioned preparation method, as a preferred embodiment, the concentration of the lactic acid bacteria liquid used for inoculation is 10⁴-10⁹CFU/mL (e.g., 10)⁵CFU/mL、10⁶CFU/mL、10⁷CFU/mL、10⁸CFU/mL, etc.), the ratio of the volume of the lactic acid bacteria liquid to the volume of the fermentation substrate is 5% to 20% (e.g., 8%, 10%, 12%, 15%, 18%, etc.).

In the above preparation method, as a preferred embodiment, in the fermentation treatment, if yeast is used, the temperature of the fermentation culture is 20 ℃ to 28 ℃ (for example, 22 ℃, 24 ℃, 26 ℃ and the like), the time is 30 to 50h (for example, 32h, 35h, 40h, 45h, 48h and the like), and the rotation speed is 100-; if lactic acid bacteria are used, the fermentation culture temperature is 37-45 deg.C (such as 38 deg.C, 40 deg.C, 42 deg.C, 44 deg.C, etc.), the fermentation culture time is 6-16h (such as 7h, 8h, 10h, 12h, 14h, 15h, etc.), and standing culture.

In the above preparation method, as a preferred embodiment, the temperature of the sterilization treatment is 100-.

In the above production method, as a preferred embodiment, the separation treatment is performed by a centrifugation method; more preferably, the centrifugation speed is 4500r/min-6000r/min (such as 4800r/min, 5000r/min, 5200r/min, 5500r/min, 5800r/min, etc.), and the centrifugation time is 30min-60min (such as 35min, 40min, 45min, 50min, 55min, etc.).

In the above preparation method, as a preferred embodiment, the method further comprises: performing preservative treatment on the lily acerola cherry fermentation liquor; more preferably, the preservation treatment is to add 2.3% by mass of ethylene glycol and 0.5% by mass of pentanediol to the fermentation broth of the lily acerola after the fermentation broth of the lily acerola is cooled to 45 ℃ or lower, that is, to add 2.3% by mass of ethylene glycol and 0.5% by mass of pentanediol to the fermentation broth of the lily acerola.

In the above preparation method, as a preferred embodiment, the method further comprises: drying the lily acerola cherry fermentation liquor to finally obtain lily acerola cherry fermentation dry powder; more preferably, the drying treatment may be spray drying, vacuum freeze drying, or the like.

According to a second aspect of the present disclosure, there is also provided a fermentation product prepared by the above method, including fermentation broth, fermentation dry powder, and the like.

According to a third aspect of the present disclosure, there is also provided the use of the above-mentioned fermented product in the preparation of a cosmetic; preferably, the cosmetic may be a mask, essence, toner, emulsion, etc.

The lily and acerola cherry fermentation product prepared by the method disclosed by the invention contains active substances such as polysaccharide, polypeptide and the like, has good antioxidation and certain anti-aging effect, and can be added into the following components in parts by weight: facial mask, essence, toner, and lotion. The inventor finds that the acerola cherry powder extract is added into the lily fermentation liquor directly to generate precipitation and instability, the two raw materials are fermented together to form new substances or intermolecular physical action under the action of microorganisms, and the final products such as the fermentation liquor and the like are more stable.

The method utilizes saccharomycetes and/or lactic acid bacteria to carry out liquid fermentation on the lily acerola, can extract active substances in lily, and realizes good fusion with the active substances of the acerola extract; firstly, yeast is used for fermentation, and then lactobacillus is used for secondary fermentation, so that the safety of the product is improved. The lily and acerola cherry fermentate prepared according to the present disclosure can be applied to cosmetics, and can improve the oxidation resistance and the anti-aging performance of the cosmetics.

The lily and acerola cherry fermentation product prepared by the method has excellent safety, can be directly used as a facial mask solution, an essence, a toner and other finished cosmetics, and has no side effect on skin.

Detailed Description

The technical solutions of the present disclosure will be described below with reference to exemplary embodiments.

The experimental procedures used in the following examples are all conventional procedures unless otherwise specified.

Materials, reagents and the like used in the following examples are commercially available unless otherwise specified.

The following examples of lily and acerola powder were from commercial sources, with dried lily purchased from kyoto, the brand name of dried lily being shop, and acerola powder purchased from seoul, biotechnology limited as 10:1 hot water extract.

The yeast in the following examples is Saccharomyces cerevisiae YWY-1, which is owned by the applicant's laboratory and has been submitted to China general microbiological culture Collection center (CGMCC for short) for preservation with the preservation date of 2019, 03 and 27 days and the preservation number of CGMCC No. 17452.

The lactic acid bacterium in the following examples was Lactobacillus delbrueckii subsp.Bulgaricus (Lactobacillus delbruuchii) with a collection number of CGMCC 1.16075 and purchased from CGMCC.

The seed culture medium of yeast and lactobacillus is yeast extract peptone glucose culture medium (YPD culture medium) and lactobacillus culture medium (MRS culture medium), respectively. Culturing yeast in a constant-temperature shaking incubator at 28 deg.C for 48 hr at 150r/min to obtain yeast liquid with concentration of about 10⁸CFU/mL. Standing and culturing lactobacillus in a constant temperature shaking incubator at 45 deg.C for 24 hr to obtain lactobacillus bacterial liquid with concentration of about 10 for inoculation¹⁰CFU/mL。

EXAMPLE 1 Yeast isolated fermentation

1) Mixing lily and acerola cherry powder with deionized water according to a material-liquid ratio (weight ratio) of 3:0.5:100, placing the mixture into a triangular flask, shaking the mixture evenly, placing the mixture into a high-pressure steam sterilization pot for sterilization at 121 ℃ for 20min, and cooling the mixture to obtain a lily acerola cherry culture medium (namely a fermentation substrate);

2) inoculating yeast liquid into a lily acerola cherry culture medium according to the volume ratio of 8%, and fermenting for 48 hours in a constant-temperature shaking culture box at the temperature of 28 ℃ and the rotating speed of 150 r/min;

3) after fermentation, centrifuging, filtering to obtain supernatant, centrifuging at 5500rpm for 45min, and sterilizing the supernatant in a high-pressure steam sterilizer at 110 deg.C for 30 min;

4) after sterilization, the supernatant was left at room temperature, cooled to below 45 ℃, and preservative system, i.e. 2.3 wt.% ethylene glycol and 0.5 wt.% pentanediol, was added.

EXAMPLE 2 lactic acid bacteria fermentation alone

2) inoculating lactobacillus liquid into a lily acerola cherry culture medium according to the volume ratio of 10%, and standing and fermenting for 8 hours in a constant-temperature shaking incubator at the temperature of 45 ℃;

3) after sterilization, the mixture is left at room temperature, cooled to below 45 ℃, and added with a preservative system, namely 2.3 wt.% of ethylene glycol and 0.5 wt.% of pentanediol.

Example 3 sequential fermentation of Yeast and lactic acid bacteria

1) Mixing lily and acerola cherry powder with deionized water according to a material-liquid ratio of 3:0.5:100, placing the mixture in a triangular flask, shaking uniformly, placing the mixture in a high-pressure steam sterilization pot for sterilization at 121 ℃ for 20min, and cooling to obtain a lily acerola cherry culture medium (namely a fermentation substrate);

3) after fermentation, placing into a high-pressure steam sterilization pot for sterilization, wherein the sterilization temperature is 110 ℃, and the sterilization time is 30 min;

4) cooling to below 45 ℃ after sterilization is finished, inoculating lactobacillus liquid according to the volume proportion of 10%, and standing and fermenting for 8 hours in a constant-temperature shaking incubator at the temperature of 45 ℃;

5) after fermentation, centrifuging, filtering to obtain supernatant, centrifuging at 5500rpm for 45min, and sterilizing the supernatant in a high-pressure steam sterilizer at 110 deg.C for 30 min;

6) after sterilization, the mixture is left at room temperature, cooled to below 45 ℃, and added with a preservative system, namely 2.3 wt.% of ethylene glycol and 0.5 wt.% of pentanediol.

Comparative example 1

Compared with the embodiment 1, the difference of the comparative example is that the fermentation substrate does not contain acerola cherry powder, other steps and process conditions are the same, and finally the lily fermentation liquor is prepared.

Comparative example 2

Compared with the embodiment 2, the difference of the comparative example is that the fermentation substrate does not contain acerola cherry powder, other steps and process conditions are the same, and finally the lily fermentation liquor is prepared.

Comparative example 3

Compared with the embodiment 3, the difference of the comparative example is that the fermentation substrate does not contain acerola cherry powder, other steps and process conditions are the same, and finally the lily fermentation liquor is prepared.

Performance test

Analysis of physicochemical Properties of fermentation broth

The physical and chemical properties of the fermentation broth of lily acerola cherry prepared in example 1 were analyzed. The appearance of the bacterial colony is light yellow transparent liquid, the pH value is 4.7, the solid content is 6.4 percent, the total number of the bacterial colonies is less than 50CFU/mL, and no pathogenic bacteria are detected.

The physical and chemical properties of the fermentation broth of lily acerola cherry prepared in example 2 were analyzed. The appearance of the bacterial colony is light yellow transparent liquid, the pH value is 4.3, the solid content is 4.7 percent, the total number of the bacterial colonies is less than 50CFU/mL, and no pathogenic bacteria are detected.

The physical and chemical properties of the fermentation broth of lily acerola cherry prepared in example 3 were analyzed. The appearance of the bacterial colony is light yellow liquid, the pH value is 4.7, the solid content is 4.3 percent, the total number of the bacterial colonies is less than 50CFU/mL, and no pathogenic bacteria are detected. According to the cosmetic hygiene standard GB7916-87, the total number of bacteria in the cosmetics is not higher than 1000CFU/ml, so that the lily acerola cherry fermentation liquid meets the requirement of cosmetic quality.

For comparison, the pH values of the lily fermentation liquids prepared in comparative examples 1 to 3 were 4.7, 4.3, and 4.7, respectively, and the solid contents were 6.4%, 4.7%, and 4.7%, respectively.

The components of the lily acerola cherry fermentation broths prepared in examples 1 to 3 were analyzed, and the results are shown in table 1 below, and the components of the lily fermentation broths prepared in comparative examples 1 to 3 are also shown in table 1 below:

TABLE 1 comparison of physicochemical Properties of fermentation broths obtained in examples 1 to 3 and comparative examples 1 to 3

	Example 1	Example 2	Example 3	Pre-fermentation blank control
					Polypeptide content mg/ml	2.54	2.41	3.06	2.50
Standard deviation of	0.05	0.12	0.007	0.004
					Polysaccharide content mg/mL	13.62	14.98	19.14	18.09
Standard deviation of	1.21	0.42	0.37	0.46
						Comparative example 1	Comparative example 2	Comparative example 3	Comparative example Pre-fermentation blank control
Polypeptide content mg/ml	2.40	2.30	2.97	2.28
					Standard deviation of	0.05	0.16	0.21	0.024
Polysaccharide content mg/mL	18.83	17.30	19.90	17.55
					Standard deviation of	0.33	0.20	0.08	1.25

As can be seen from table 1: the polypeptide content of the examples 1 and 3 is higher than that before fermentation, and the polypeptide content after double-bacterium fermentation of the example 3 is the highest, which shows that the microbial action is helpful for the dissolution of the polypeptide; example 3 polysaccharide content increased after fermentation, indicating that the microorganism produced exopolysaccharides; in the comparative example without acerola powder, it can be seen that the polypeptide content after fermentation is higher than that of the blank control before fermentation, which indicates that the microbial action is favorable for the dissolution of the polypeptide, and the polysaccharide content after fermentation is higher than that before fermentation in the comparative examples 1 and 3, which indicates that the microorganism is propagated to produce extracellular polysaccharide.

Secondly, safety detection of fermentation liquor

The human body patch test is mainly used for detecting the irritation of the final cosmetic product or raw materials. The invention carries out a human body closed patch test on the lily acerola cherry fermentation liquor obtained in the examples 1 to 3, and aims to evaluate the potential skin irritation of the lily acerola cherry fermentation liquor.

1. Test object

Suitable volunteers were selected for 30 persons, and were randomly selected in the age range of 18-60 years.

2. Test method

0.02mL to 0.025mL of the liquid sample (i.e., the lily acerola cherry fermentation broth) is dripped on a filter paper sheet, and then the filter paper sheet is placed in a spot tester. A blank control is set for each sample, and an equal amount of sample solvent, such as distilled water or olive oil (distilled water is used in this example), 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 24 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 24h, 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 24h and 48h after the patch test.

3. Test results

The patch test results are shown in table 2, and each level in table 2 indicates the following: grade 0 is a negative reaction. Grade 1 ═ suspicious response; only faint erythema. Grade 2 ═ weakly positive reaction (erythema reaction); erythema, infiltration, edema, and possibly pimples. Grade 3 ═ strong positive response (herpes response); erythema, infiltration, edema, pimples, herpes; the reaction may be beyond the test area. Grade 4 ═ very strong positive response (fusogenic herpes response); obvious erythema, severe infiltration, edema, and fusional herpes; the reaction goes beyond the test area.

TABLE 2 Patch test results of fermentation liquors obtained in examples 1 to 3 and comparative examples 1 to 3

As can be seen from table 2: the fact that the yeast obtained in the example 1 singly ferments the fermentation broth of the lily acerola cherry to generate slight suspicious reaction, while the fermentation broth of the lily acerola cherry obtained in the example 2 by two-step fermentation of the yeast lactic acid bacteria and the fermentation broth of the lily acerola cherry obtained in the example 3 singly ferments the lactic acid bacteria do not generate suspicious reaction, indicates that the fermentation broth of the lily acerola cherry provided by the invention has higher safety and does not bring adverse reaction to human bodies. The substrate which has not been fermented in the comparative example may have slight suspicious reaction, and the lily fermentation liquids prepared in comparative examples 1 to 3 have no suspicious reaction, and the safety is high.

Thirdly, detecting the oxidation resistance of the fermentation liquor

DPPH is an early synthesized organic radical, commonly used to evaluate the hydrogen donating ability of antioxidants, is very stable in organic solvents, is purple in color, and has a characteristic absorption peak at 517nm, when encountering a radical scavenger, the lone pair of DPPH is paired to discolor it, i.e., the absorbance at the maximum absorption wavelength becomes small. Therefore, the effect of the sample on DPPH radical scavenging can be evaluated by measuring the change in absorbance.

The lily acerola cherry fermentation liquid prepared in the examples 1 to 3 is used as a liquid to be tested, and vitamin C is used as a positive control. The specific experimental steps of the DPPH free radical scavenging experiment are as follows:

(1) taking the same volume (3mL) of the solution to be detected and 2X 10^-4mixing (A) with a solution of DPPH in mol/L₁A tube);

(2) taking equal volume (3mL) of absolute ethanol (solvent of the test substance) and 2X 10^-4mixing (A) with a solution of DPPH in mol/L₂A tube);

(3) mixing the same volume (3mL) of anhydrous ethanol with the solution to be detected (A)₃A tube);

(4) after 30min of reaction, A was measured at 517nm₁Pipe, A₂Pipe, A₃Tube absorbance values.

The clearance calculation formula is: clearance (%) - (A)₂+A₃)-A₁]/A₂. See table 3 for specific results.

TABLE 3 DPPH radical scavenging test results for broths prepared in examples 1 to 3 and comparative examples 1 to 3

Example 1	Example 2	Example 3	Pre-fermentation blank control
				DPPH radical clearance rate	53.58％	69.61％	82.43％	32％
Comparative example 1	Comparative example 2	Comparative example 3	Comparative example Pre-fermentation blank control
				DPPH radical clearance rate	38.4％	40.25％	47.2％	36％

As can be seen from Table 3, the DPPH free radical clearance after fermentation is higher than that of the blank control before fermentation, the fermentation broth is beneficial to clearing DPPH free radicals, and the free radical clearance of the dual-strain fermentation in example 3 is higher than that of examples 1 and 2; the radical scavenging ability after fermentation in comparative example was higher than that of the pre-fermentation blank, with the radical scavenging ability being highest in comparative example 3; the DPPH free radical of the fermentation broth obtained in the example is clearly more effective than the comparative example.

Fourth, detecting cytotoxicity of fermentation liquor

Human Skin Fibroblasts (HSF) were cultured in a medium containing 10% fetal bovine serum and 1% double antibody (1X 10)⁵U/L penicillin, 100mg/L streptomycin). Cells were grown at 37 ℃ with 5% CO₂In the incubator with saturated humidity, when the cell fusion reaches more than 85%, the cells are digested and passaged with 0.05% pancreatin.Human dermal fibroblasts in good logarithmic growth phase were seeded in a cell culture plate for experiment.

The MTT method is used for detecting the influence of fermentation liquor on the survival rate of human skin fibroblasts. The method comprises the following specific steps: cells in the logarithmic growth phase were digested and the digestion was stopped with DMEM containing serum. Counting with cell counting plate, adjusting cell suspension concentration to 5 × 10⁴one/mL, the cell suspension was inoculated into a 96-well plate at a rate of 100. mu.L/well, at 37 ℃ with 5% CO₂Incubate under conditions overnight. Removing culture solution, adding filtered and sterilized fermentation liquid of lily acerola cherry, making 6 multiple holes for each sample, and culturing for 24 h. After the completion of the culture, 100. mu.L of a mixed solution of MTT solution (5mg/mL) and DMEM (v/v, 1: 5) was added to each well and incubated for 4 hours. The medium was removed, 150. mu.L of DMSO was added to each well, and the absorbance (denoted A) of the experimental group was read at 490nm after incubation at 37 ℃ for 10 min. The cell-free treated group (cell suspension was replaced with serum-free DMEM, and the rest was the same) was set as a blank control group and designated as B. The cell control group (sample solution was replaced with serum-free DMEM, and the rest of the procedure was the same) was designated as C.

The calculation formula of the cell viability is as follows: cell viability (%) - (a-B)/(C-B) × 100;

see table 4 for specific results.

TABLE 4 results of MTT cell assay of fermentation broths prepared in examples 1 to 3 and comparative examples 1 to 3

	Pre-fermentation blank control	Example 1	Example 2	Example 3
					Cell viability%	50.53	98.20	86.30	103.46
Standard deviation std	5.079406	8.73269	4.516238	6.131151
						Comparative example Pre-fermentation blank control	Comparative example 1	Comparative example 2	Comparative example 3
Cell viability%	57.07	100.08	108.24	124.62
					Standard deviation std	5.079406	3.818974	12.20607	22.32536

As can be seen from table 4, the survival rate of cells treated with the substrate that has not yet been fermented is low, and the yeast-fermented single lily acerola cherry fermentation broth obtained in example 1, the lactic acid bacteria-fermented single lily acerola cherry fermentation broth obtained in example 2, and the yeast-fermented lactic acid bacteria-fermented two-step lily acerola cherry fermentation broth obtained in example 3 all have a good proliferation effect on cells, and particularly the fermentation broth obtained in example 3 is more effective; in the comparative examples, the fermentation broth without acerola powder was also non-cytotoxic and higher than the examples.

Finally, it is also noted that, in the present disclosure, relational terms such as first and second, and the like are used solely to distinguish one entity or action from another entity or action without necessarily requiring or implying any actual such relationship or order between such entities or actions. Also, the terms "comprises," "comprising," or any other variation thereof, are intended to cover a non-exclusive inclusion, such that a process, method, article, or apparatus that comprises a list of elements does not include only those elements but may include other elements not expressly listed or inherent to such process, method, article, or apparatus. Without further limitation, an element defined by the phrase "comprising an … …" does not exclude the presence of other identical elements in a process, method, article, or apparatus that comprises the element.

While the disclosure has been disclosed above by the description of specific embodiments thereof, it should be understood that various modifications, improvements or equivalents of the disclosure may be devised by those skilled in the art within the spirit and scope of the appended claims. Such modifications, improvements and equivalents are intended to be included within the scope of the present disclosure as claimed.

Claims

1. A preparation method of a lily and acerola cherry fermented product for cosmetics is characterized by comprising the following steps:

mixing a proper amount of lily, acerola powder and water, sterilizing, and cooling to obtain a fermentation substrate, wherein the mass ratio of the lily to the water is 0.5-5: 100, and the mass ratio of the acerola powder to the water is 0.1-3: 100; the lily is dried lily, and the acerola cherry powder is an acerola cherry extract;

2. The method of claim 1, wherein the fermentation treatment comprises: firstly, inoculating saccharomycetes to the fermentation substrate to carry out primary fermentation culture; inoculating lactobacillus after sterilization treatment for secondary fermentation culture; preferably, the dried lily is crushed, and the particle size is 100 meshes.

3. The preparation method according to claim 1 or 2, wherein the yeast is Saccharomyces cerevisiae (Saccharomyces cerevisiae) YWY-1, the preservation unit is China general microbiological culture Collection center (CGMCC for short), the address is No. 3 of Xilu 1 of Beijing, Chaoyang, on the north of the morning, the postal code is 100101, the preservation date is 2019, 03 and 27 days, and the preservation number is CGMCC No. 17452;

preferably, the concentration of the yeast liquid for inoculation is 10⁴-10⁹CFU/mL, wherein the volume ratio of the yeast liquid to the fermentation substrate is 5-20%.

4. The method according to any one of claims 1 to 3, wherein the lactic acid bacteria are at least one of the following species: lactobacillus delbrueckii subsp. bulgaricus, Lactobacillus buchneri, Bifidobacterium bifidum (Bifidobacterium bifidum);

preferably, the lactic acid bacteria are at least one of the following species: lactobacillus delbrueckii subsp. bulgaricus with a collection number of CGMCC 1.16075; lactobacillus buchneri (Lactobacillus buchneri) with the preservation number of CGMCC 1.15607; bifidobacterium bifidum (CGMCC 1.5029) with preservation number;

more preferably, the lactic acid bacterium is Lactobacillus delbruuchii subsp. bulgaricus;

further, the Lactobacillus is Lactobacillus delbruuchii subsp.bulgaricus with the preservation number of CGMCC 1.16075;

further, the concentration of the lactobacillus bacterial liquid for inoculation is 10⁴-10⁹CFU/mL, the volume ratio of the lactobacillus liquid to the fermentation substrate is 5-20%.

5. The method according to any one of claims 1 to 4, wherein in the fermentation treatment, if yeast is used, the temperature of the fermentation culture is 20 ℃ to 28 ℃, the time is 30 to 50 hours, and the rotation speed is 100-; if lactic acid bacteria are adopted, the temperature of fermentation culture is 37-45 deg.C, the time is 6-16h, and standing for fermentation.

6. The method according to any one of claims 1 to 5, wherein the temperature of the sterilization treatment is 100 ℃ and 121 ℃ for 15 to 30 min.

7. The production method according to any one of claims 1 to 6, wherein the separation treatment is performed by a centrifugation method; more preferably, the centrifugal rotating speed is 4500r/min-6000r/min, and the centrifugal time is 30min-60 min.

8. The production method according to any one of claims 1 to 7, further comprising: performing preservative treatment on the lily acerola cherry fermentation liquor; more preferably, the preservation treatment is adding 2.3 wt.% of ethylene glycol and 0.5 wt.% of pentanediol after the lily acerola fermentation liquor is cooled to below 45 ℃.

9. The method of any one of claims 1-8, further comprising: drying the lily acerola cherry fermentation liquor to finally obtain lily acerola cherry fermentation dry powder; more preferably, the drying process is spray drying or vacuum freeze drying.

10. A fermentation product produced by the method of any one of claims 1-9.