CN115074285A

CN115074285A - Bifidobacterium enzymolysis lysate and preparation process and application thereof

Info

Publication number: CN115074285A
Application number: CN202210764938.5A
Authority: CN
Inventors: 陈江; 孔英俊; 柳媛媛
Original assignee: Zhongke Huaqi Beijing Biotechnology Research Institute Co ltd
Current assignee: Zhongke Huaqi Beijing Biotechnology Research Institute Co ltd
Priority date: 2022-07-01
Filing date: 2022-07-01
Publication date: 2022-09-20

Abstract

The invention discloses a bifidobacterium enzymolysis lysate and a preparation process and application thereof, wherein an oat culture medium is prepared by a solid state fermentation method; screening a plurality of bifidobacterium strains with antioxidant activity, respectively inoculating the bifidobacterium strains in the oat culture medium for fermentation to obtain a seed solution of each bifidobacterium strain; preparing a culture medium, inoculating one or two of the obtained seed liquids of the bifidobacterium strains in the culture medium, and performing primary fermentation to obtain a primary fermentation product; adding nutrient substances and seed liquid which is the same as that used for the primary fermentation into the primary fermentation product, and then performing secondary fermentation to obtain a secondary fermentation product; centrifuging the secondary fermentation product, filtering, homogenizing and cracking under high pressure to obtain bifidobacterium cytolytic solution and cell wall fragments; carrying out enzymolysis treatment on the obtained cell wall fragments, centrifuging and filtering to obtain cell fragment enzymolysis liquid; and compounding the bifidobacterium lysate and the cell debris enzymolysis liquid to obtain the bifidobacterium enzymolysis lysate.

Description

Bifidobacterium enzymolysis lysate and preparation process and application thereof

Technical Field

The invention relates to the technical field of probiotic fermentation, in particular to a bifidobacterium enzymolysis lysate and a preparation process and application thereof.

Background

The bifidobacterium is a physiological bacterium existing in a human body, is one of beneficial bacteria of the human body which is most valued and researched, and is inseparable from the health of the human body. It can be said that the nature bestows on the health law treasure of the human.

As with the human intestinal tract, hundreds of millions of microorganisms live on the skin. These various microorganisms interact with each other and with the skin of a person, and together constitute the skin's microecosystem. At present, the product development idea of the skin microecology concept mainly provides nutrient conditions for establishing a probiotic environment for skin by adding a probiotic fermentation product, so that the skin microecology maintenance effect is realized to a certain extent.

In the prior art, in the preparation process of bifidobacterium fermentation lysate, the treatment mode of cells after the fermentation process is usually centrifugation and other modes to separate the residual components of the cells and the culture medium, and the method causes the loss of active components in the cells; or cell wall is cracked to form cell wall fragment and cell lysate coexisting product, and this results in relatively great cell wall fragment, precipitate in the lower layer of cell lysate and is used in cosmetics.

Disclosure of Invention

One object of the present application is to provide a bifidobacterium enzymatic lysate with a soluble polysaccharide content 18.32 times higher than that of the product of the conventional fermentation process, and an increase in the content of hydrolyzed amino acids of 19.04%.

Another object of the present application is to provide a process for the preparation of a bifidobacterium enzymatic lysate and use thereof.

To achieve the above purpose, the scheme is as follows:

preparing an oat culture medium by adopting a solid state fermentation method;

screening a plurality of bifidobacterium strains with antioxidant activity, respectively inoculating the bifidobacterium strains in the oat culture medium for fermentation to obtain a seed solution of each bifidobacterium strain;

preparing a culture medium, inoculating one or two of the obtained seed solutions of multiple bifidobacteria strains in the culture medium, and performing primary fermentation to obtain a primary fermentation product;

adding nutrient substances and seed liquid which is the same as that used for the primary fermentation into the primary fermentation product, and then performing secondary fermentation to obtain a secondary fermentation product;

centrifuging the secondary fermentation product, filtering, homogenizing and cracking under high pressure to obtain bifidobacterium cytolytic solution and cell wall fragments;

carrying out enzymolysis treatment on the obtained cell wall fragments, centrifuging and filtering to obtain cell fragment enzymolysis liquid;

and compounding the bifidobacterium lysate and the cell debris enzymolysis liquid to obtain the bifidobacterium enzymolysis lysate.

Preferably, the step of preparing the oat culture medium by the solid state fermentation method comprises the following steps:

soaking and boiling oat peel in deionized water at 95-100 deg.C, taking out, cooling, adding yeast, mixing, spreading white sugar, and performing solid fermentation to obtain solid fermentation product;

pulverizing the solid fermentation product, adding deionized water, maintaining the temperature, stirring, sieving to obtain filtrate, adding glucose and agar into the filtrate, and stirring to obtain oat culture medium.

Preferably, the cell debris enzymatic hydrolysate obtaining step is as follows:

adding enzyme into cell wall fragments obtained by high-pressure homogenization and cracking for enzymolysis, adjusting the pH value to 8-8.5, controlling the water bath temperature to 52-58 ℃, and heating for 3-4 hours;

then placing the reaction in a water bath at the temperature of 90-95 ℃, and heating for 15-20min for enzyme inactivation;

after inactivation, placing in a refrigerator for refrigeration, centrifuging, and separating out supernate which is cell debris enzymolysis liquid;

preferably, the enzyme used for the enzymolysis treatment of the obtained cell wall fragments is glycoside hydrolase or one of the following enzymes:

alcalase alkaline protease, animal proteases, plant proteases and microbial proteases.

Preferably, the bifidobacterium species comprises one or two of bifidobacterium adolescentis, bifidobacterium animalis, bifidobacterium breve, bifidobacterium longum and bifidobacterium thermophilum.

Preferably, the culture medium for inoculating one or two of the obtained seed solutions of the plurality of bifidobacterium species in the culture medium comprises the following components in percentage by mass based on the total weight of the culture medium: 40-80 wt% of oat culture medium, 0.1-0.2 wt% of vitamin complex, 0.1-0.2 wt% of urea and the balance of seed liquid.

Preferably, the nutrient substances added into the primary fermentation product comprise glucose and stachyose, and the mass percentage of the glucose added into the primary fermentation product is 0.5-2 wt% based on the total weight of the primary fermentation product; the weight percentage of the added stachyose is 0.05-0.1 wt%.

Preferably, the total sugar content in the primary fermentation product is below 0.5 wt%, and the pH value is below 4.5; the pH value of the secondary fermentation product is above 4.5.

Preferably, the step of obtaining the bifidobacterium enzymolysis lysate by compounding the bifidobacterium lysate and the cell debris enzymolysis solution comprises the following steps:

adding p-hydroxyacetophenone and one or more of hexanediol, pentanediol and methyl propylene glycol into the mixed liquid of the bacillus bifidus cytolytic solution and the cell debris enzymolysis solution.

The bifidobacterium enzymolysis lysate prepared by the steps can be applied to cosmetics.

The invention has the following beneficial effects:

the bifidobacterium enzymolysis lysate is prepared in a mode of combining a solid oat fermentation medium and a liquid fermentation process, and compared with a single liquid fermentation lysate, the oxidation resistance is improved by 20%; according to the method, the culture medium is prepared from the oat fermentation filtrate, contains abundant sugar sources, nitrogen sources and inorganic substances, and can reduce the introduction of manganese sulfate, beef jelly and other components compared with the traditional culture medium, so that the safety of bifidobacterium fermentation products is greatly improved, and the generation of unpleasant odor is reduced; the method is characterized in that an enzymolysis process is added, namely cell fragments after fermentation are subjected to enzymolysis treatment to obtain soluble polysaccharide and micromolecular peptide; the polysaccharide content in the bifidobacterium enzymolysis lysate prepared by the enzymolysis process is increased by 18.32 times of the polysaccharide content in the lysate prepared by the traditional fermentation process, and the hydrolyzed amino acid content is improved by 19.04 percent. The cell wall fragments form water-soluble components through enzymolysis, no precipitate exists in the product, the use in various formulations of cosmetics is facilitated, and the influence of the cell wall fragments on the appearance of the product is not required to be considered.

Drawings

In order to illustrate the implementation of the solution more clearly, the drawings that are needed in the description of the embodiments will be briefly described below, it being obvious that the drawings in the following description are only some embodiments of the solution, and that other drawings may be derived from these drawings by a person skilled in the art without inventive effort.

FIG. 1 is a flow chart of a preparation process of a solid state fermentation oat culture medium;

FIG. 2 is a flow diagram of a process for the preparation of a bifidobacterium enzymatic lysate;

FIG. 3 is a diagram of a bifidobacterium enzymatic lysate of the present application (right) versus a conventional bifidobacterium lysate (left) as prepared;

fig. 4 is a graph of a bifidobacterium enzymatic lysate of the present application (right) versus a conventional bifidobacterium lysate (left) resting for three hours.

Detailed Description

The embodiments of the present application are described in further detail below, and it is apparent that the described examples are only a part of the examples of the present application, and are not exhaustive of all the examples. It should be noted that the embodiments and features of the embodiments in the present application may be combined with each other without conflict.

The terms "first," "second," and the like in the description and in the claims, and in the drawings described above, if any, are used for distinguishing between similar elements and not necessarily for describing a particular sequential or chronological order. It will be appreciated that the data so used may be interchanged under appropriate circumstances such that the embodiments described herein may be practiced otherwise than as specifically illustrated or described herein. Furthermore, the terms "comprises," "comprising," and "having," and any variations thereof, are intended to cover a non-exclusive inclusion, such that a process, method, system, article, or apparatus that comprises a list of steps or elements is not necessarily limited to those steps or elements expressly listed, but may include other steps or elements not expressly listed or inherent to such process, method, article, or apparatus.

The bifidobacterium fermentation lysate is a physiological lysate obtained by processing after bifidobacterium fermentation, and the main components of the lysate are cell metabolites, cytoplasmic fragments, polysaccharides, proteins, polypeptides and the like. A large number of in vitro experiments prove that various amino acids, proteins and various molecular media in the bifidobacterium fermentation lysate have the effects of regulating and balancing skin and regulating immune function, are also nutritional molecules of human skin cells, have certain anti-wrinkle, compact, repairing, sun-screening, anti-oxidation and anti-aging effects, can prevent damage caused by external stimulation such as ultraviolet rays and the like, and promote repair of damaged DNA. However, the existing preparation process of bifidobacterium fermentation lysate used as cosmetic raw materials has the problems of low content of functional active ingredients such as proteins and small molecular polypeptides easy to be absorbed by skin and the like, and the quality of the fermentation lysate is influenced.

The method prepares the bifidobacterium enzymolysis lysate by combining a solid fermentation oat culture medium and a liquid fermentation process, and the antioxidant performance of the prepared bifidobacterium lysate is improved by 20 percent compared with that of a single liquid fermentation lysate; the method is characterized in that an enzymolysis process is added, namely cell fragments after fermentation are subjected to enzymolysis treatment to obtain soluble polysaccharide and micromolecular peptide; the polysaccharide content in the bifidobacterium enzymolysis lysate prepared by the enzymolysis process is increased by 18.32 times of the polysaccharide content in the lysate prepared by the traditional fermentation process, the hydrolyzed amino acid content is increased by 19.04 percent, and the enzyme used in the enzymolysis process is not limited to Alcalase alkaline protease, and can also be animal protease, plant protease, microbial protease and the like.

Examples

1. Preparation of oat culture medium by solid state fermentation method

FIG. 1 is a process flow of preparing oat culture medium by solid state fermentation in this example;

peeling 100 parts by mass of oat, then placing the oat in deionized water at 95-100 ℃ for immersion and boiling for 4 hours, fishing out the oat until the oat is soft and rotten, and cooling to room temperature;

uniformly scattering 0.4 part by mass of saccharomycetes on the cooled oat, stirring to uniformly mix the saccharomycetes and the cooled oat, uniformly scattering white sugar on the surface until the surface is completely covered by the white sugar, finally covering a cover with an exhaust function, standing for 7 days at the temperature of 20-30 ℃, and performing solid state fermentation;

and (3) carrying out wall breaking and crushing on the oat subjected to solid state fermentation, and then mixing the oat with the water according to a mass ratio of 1: adding 50 of the filtrate into deionized water, keeping the temperature at 85-90 ℃, stirring for 2 hours, sieving by a 300-mesh double-layer sieve, and collecting the filtrate for later use;

mixing 0.5 wt% of glucose, 2.0 wt% of agar and 97.5 wt% of filtrate according to the mass percentage of all substances in the total mass of the culture medium, and uniformly stirring to obtain the oat culture medium.

2. Liquid fermentation method for preparing bifidobacterium enzymolysis lysate

FIG. 2 is a process flow of preparing a lysate of Bifidobacterium enzymolysis by liquid fermentation in steps 2.1-2.7 in this embodiment.

2.1 obtaining Bifidobacterium seed liquid

Different species of bifidobacteria obtained based on 16S rDNA sequencing technology, including bifidobacterium adolescentis, bifidobacterium horn, bifidobacterium animalis, bifidobacterium asteroides, bifidobacterium bifidum, bifidobacterium bovis, bifidobacterium breve, bifidobacterium odonta, bifidobacterium infantis, namely bifidobacterium longum subspecies infantis, bifidobacterium lactis, namely bifidobacterium animalis subspecies cremoris, bifidobacterium longum, bifidobacterium pseudocatenulatum, bifidobacterium thermophilum and bifidobacterium acidophilum, are subjected to safety screening after fermentation to select strains with high safety: bifidobacterium adolescentis, Bifidobacterium animalis, Bifidobacterium breve, and Bifidobacterium longum and Bifidobacterium thermophilum;

inoculating one or two of the selected strains with high safety according to 3-10 wt% of oat culture medium, fermenting at 36-38 deg.C for 2 days (about 48 hr), and performing microscopic examination to ensure the activation of Bifidobacterium and obtain Bifidobacterium seed solution.

2.2 obtaining a primary fermentation product

According to the mass percentage of all the substances in the total mass of the culture medium, 40-80 wt% of an oat culture medium, 0.1-0.2 wt% of a vitamin complex, 0.1-0.2 wt% of urea and the balance of bifidobacterium seed liquid are supplemented, and anaerobic fermentation is carried out after the materials are uniformly mixed and stirred;

and (3) carrying out anaerobic fermentation for 20-24 hours at the temperature of 36-38 ℃, and obtaining a primary fermentation product when the total sugar content is reduced to be below 0.5% and the pH is below 4.5.

2.3 obtaining the Secondary fermentation product

Adding 0.5-2 wt% of glucose, 0.05-0.1 wt% of stachyose and 2-20 wt% of bifidobacterium seed liquid into the obtained primary fermentation product according to the mass percentage of all the substances in the total mass of the culture medium for secondary fermentation;

and (3) obtaining a secondary fermentation product under the conditions of 36-38 ℃ and 6-8 h, wherein the pH value is above 4.5.

2.4 centrifugal separation

And (3) centrifuging the obtained secondary fermentation product at a high speed under the centrifugation condition of 2000-3000 rmp/min for 15-20min, then filtering, and separating supernatant from lower-layer cells.

2.5 cell lysis

Adding distilled water into the lower layer cells obtained after centrifugal separation according to the mass ratio of 1:10, stirring for 10-20 min, dissolving out intracellular matrixes of the lower layer cells, and performing high-speed centrifugation;

centrifuging for 15-20min at 2000-3000 rmp/min, filtering, separating supernatant from lower layer cells, and repeating the steps twice;

carrying out high-pressure homogenization treatment on the lower layer of cells obtained after filtration for 2-3 times, carrying out microscopic examination until the cell lysis degree reaches more than 95%, and separating the upper layer of cytolytic fluid from the lower layer of cell wall fragments by using a 300-mesh double-layer sieve;

then combining the lysate subjected to the high-pressure homogenization treatment with the supernatant subjected to centrifugal separation to obtain a bifidobacterium cytolytic solution;

and reserving the lower-layer cell wall fragments after high-pressure homogenization treatment for later use.

2.6 obtaining cell debris enzymatic hydrolysate

The mass ratio of each substance to the cell debris enzymolysis liquid is calculated. Mixing 9 wt% of the lower cell wall fragments subjected to high-pressure homogenization with deionized water, adding 5-6 wt% of Alcalase alkaline protease (enzyme activity is more than 60 million U/g, enzyme substrate ratio is more than 5000U/g), 10-12 wt% of glycoside hydrolase (enzyme activity is more than 30 million U/g, enzyme substrate ratio is more than 5000U/g), adding the above substances into a reaction container, adding the rest of deionized water, fully dispersing and mixing the substances uniformly by using a water bath oscillator, adjusting the pH value to 8-8.5, the water bath temperature to be 55 ℃, heating for 3-4 hours, putting the reaction container into a water bath at 90-95 ℃ for 15-20 minutes to inactivate the Alcalase alkaline protease and the glycoside hydrolase, putting the inactivated reaction container into a refrigerator for refrigeration at 0-5 ℃ overnight, centrifuging, the centrifugation condition is 6000rmp/min, 8min, centrifuging the supernatant, obtaining cell debris enzymatic hydrolysate;

in this step, the selected enzyme is not limited to Alcalase alkaline protease, but animal protease, plant protease, microbial protease and the like can be selected, and the corresponding pH value, temperature and reaction time are adjusted according to the type and characteristics of the used enzyme.

2.7 obtaining Bacillus bifidus enzymolysis lysate by compounding

Mixing the bacillus bifidus cytolytic solution obtained in the step 2.5 with the cell debris enzymolysis solution obtained in the step 2.6, and adding 1-3 of hexanediol, pentanediol and methyl propanediol into the mixed solution according to the proportion of 0.5-4.0 wt% of the total amount; 0.1-0.5 wt% of p-hydroxyacetophenone, and the product obtained after uniform mixing is the bifidobacterium enzymolysis lysate.

Effects of the embodiment

3. Composition detection

3.1 hydrolyzed amino acid content

The bifidobacterium enzymolysis lysate obtained in the embodiment of the application adopts an amino acid automatic analyzer (Japanese Kokai Higao New technology science, LA8080) to determine the hydrolyzed amino acid experimental method;

the hydrolyzed amino acid content of the bifidobacterium enzymatic lysate obtained in the examples of the present application (sample 1) was compared to the traditional bifidobacterium lysate (sample 2); the amino acid content of the bifidobacterium enzymatic lysate and bifidobacterium lysate is as follows in table 1:

TABLE 1

From the results comparison analysis in table 1, it follows: compared with the hydrolyzed amino acid content of the conventional bifidobacterium lysate (sample 2), the bifidobacterium enzymolysis lysate (sample 1) obtained in the embodiment of the application is improved by 19.04 percent, and an unexpected technical effect is achieved.

3.2 polysaccharide content

The polysaccharide content of the bifidobacterium enzymatic lysate obtained in the examples of the present application (sample 1) was compared to the polysaccharide content of a conventional bifidobacterium lysate (sample 2); the polysaccharide content in the bifidobacterium enzymatic lysate and bifidobacterium lysate is as follows in table 2:

TABLE 2

Test sample	Sample 1	Sample 2
			Polysaccharide content mg/ml	11.4	0.59

From the comparative analysis of the results in table 2, it follows: the bifidobacterium enzymolysis lysate (sample 1) obtained in the examples of the present application is 19.32 times the polysaccharide content of the conventional bifidobacterium lysate (sample 2), and unexpected technical effects are achieved.

3.3 Small molecule peptide content

The bifidobacterium enzymolysis lysate obtained in the examples of the present application (sample 1) is compared to the conventional bifidobacterium lysate (sample 2) for its content of small molecule peptides below 300D; the following table 3 shows the contents of 300D-less small molecule peptides in the bifidobacterium enzymatic lysate and bifidobacterium lysate:

TABLE 3

Test sample	Sample 1	Sample 2
			Peptides below 300D (g/100mL)	0.13	0.06

From a comparative analysis of the results in table 2: the bifidobacterium enzymolysis lysate (sample 1) obtained in the embodiment of the application is 2.2 times of the content of the small molecular peptide below 300D of the traditional bifidobacterium lysate (sample 2), and the unexpected technical effect is achieved.

3.4 evaluation of Oxidation resistance

The experiment adopts a stable and representative free radical-DPPH free radical as an experimental object, DPPH is a very stable free radical taking nitrogen as a center, if a test object can remove the free radical, the test object has the effect of removing the free radical, and the method is widely used for research on free radical removal, and the principle of the method is as follows: the DPPH free radical with single electron is dark purple in ethanol solution, has strong absorption at 517nm wavelength, when the DPPH solution is added with the free radical scavenger, the solution becomes light in color due to the pairing with the single electron, the absorption disappears gradually, the fading degree is in linear relation with the number of electrons received, and the measurement can be carried out by a spectroscopic method.

The antioxidant properties of the bifidobacterium enzymatic lysate obtained in the examples of the present application (sample 1) were compared with those of the conventional bifidobacterium lysate (sample 2) and of the commercially available samples 3 and 4 of the same type, the antioxidant properties of the four samples being shown in table 4 below:

TABLE 4

Test sample	IC50 value mg/ml
		Sample 1	0.0032
Sample 2	0.0041
		Competition 3	0.0102
Competition 4	0.0078

From the comparative analysis of the results in table 4: compared with the traditional bifidobacterium lysate (sample 2), the bifidobacterium enzymolysis lysate (sample 1) obtained in the embodiment of the application has the advantage that the antioxidant property is improved by 20 percent;

compared with the samples 3 and 4 of the same type which are sold in the market, the bifidobacterium enzymolysis lysate (sample 1) obtained in the embodiment of the application has the antioxidant performance which is improved by 58.9 to 68.6 percent.

3.5 evaluation of stability

Figures 3-4 show the bifidobacterium enzymatic lysate prepared in the present application in solution with a conventional bifidobacterium lysate. Neither the freshly prepared bifidobacterium enzymatic lysate in fig. 3 (sample 1 on the right of the figure) nor the conventional bifidobacterium lysate solution (sample 2 on the left of the figure) was precipitated; however, the bifidobacterium enzymolysis lysate (sample 1 on the right of the figure) is clear and transparent, and has lighter color, and the bifidobacterium lysate solution (sample 2 on the left of the figure) has darker color. Fig. 4 shows that the bifidobacterium enzymolysis lysate (sample 1 on the right of the figure) is left for three hours, and the conventional bifidobacterium lysate solution (sample 2 on the left of the figure), the bifidobacterium enzymolysis lysate (sample 1 on the right of the figure) has no obvious precipitation, and the bifidobacterium lysate solution (sample 2 on the left of the figure) has obvious precipitation.

The cell wall fragments form water-soluble components through enzymolysis, no precipitation exists in the bifidobacterium enzymolysis lysate, the bifidobacterium enzymolysis lysate is favorable for being used in various formulations of cosmetics, and the influence of the cell wall fragment precipitation on the appearance of the product is avoided.

The effect of the above embodiment can be obtained, the method adopts a unique mode of combining the solid fermentation oat culture medium and the liquid fermentation process, and the bifidobacterium enzymolysis lysate prepared by the enzymolysis process has the characteristics of better oxidation resistance, more soluble polysaccharides, more small molecular peptides, higher content of hydrolyzed amino acids and the like.

It should be understood that the above-mentioned embodiments of the present invention are only examples for clearly illustrating the present invention, and are not intended to limit the embodiments of the present invention, and it will be obvious to those skilled in the art that other variations or modifications may be made on the basis of the above description, and all embodiments may not be exhaustive, and all obvious variations or modifications may be included within the scope of the present invention.

Claims

1. A bifidobacterium enzymatic lysate prepared by the method comprising:

preparing an oat culture medium by adopting a solid state fermentation method;

preparing a culture medium, inoculating one or two of the obtained seed liquids of the bifidobacterium strains in the culture medium, and performing primary fermentation to obtain a primary fermentation product;

2. A bifidobacterium enzymatic lysate as claimed in claim 1 wherein the oat culture medium is prepared by a solid state fermentation process comprising the steps of:

3. A bifidobacterium enzymatic lysate as claimed in claim 1 in which the cell debris enzymatic lysate is obtained by the steps of:

adding enzyme into cell wall fragments obtained by high-pressure homogeneous cracking for enzymolysis, adjusting the pH value to 8-8.5, adjusting the water bath temperature to 52-58 ℃, and heating for 3-4 hours;

inactivating, refrigerating in refrigerator, centrifuging, and separating supernatant as cell debris enzymolysis solution.

4. A Bifidobacterium enzymatic lysate according to claim 1, wherein the enzyme used in the enzymatic treatment of the obtained cell wall fragments is a glycoside hydrolase and one of the following enzymes:

5. A bifidobacterium enzymatic lysate as claimed in claim 1 wherein the bifidobacterium species comprises one or both of bifidobacterium adolescentis, bifidobacterium animalis, bifidobacterium breve and bifidobacterium longum and bifidobacterium thermophilum.

6. The bifidobacterium enzymolysis lysate of claim 1, wherein the culture medium for inoculating the seed solution of one or two of the obtained bifidobacterium species in the culture medium comprises the following components in percentage by mass based on the total weight of the culture medium:

40-80 wt% of oat culture medium, 0.1-0.2 wt% of vitamin complex, 0.1-0.2 wt% of urea and the balance of seed liquid.

7. The bifidobacterium enzymolysis lysate of claim 1, wherein the nutrient substances added to the primary fermentation product comprise glucose and stachyose, and the mass percentage of the glucose added to the primary fermentation product is 0.5-2 wt% based on the total weight of the primary fermentation product; the weight percentage of the added stachyose is 0.05-0.1 wt%.

8. A bifidobacterium enzymatic lysate as claimed in claim 1 in which the total sugar content of the primary fermentation product is less than 0.5 wt% and the pH is less than 4.5; the pH value of the secondary fermentation product is above 4.5.

9. The bifidobacterium enzymatic lysate of claim 1, wherein the step of combining the bifidobacterium lysate and the cell debris enzymatic hydrolysate to obtain the bifidobacterium enzymatic lysate comprises: adding p-hydroxyacetophenone and one or more of hexanediol, pentanediol and methyl propylene glycol into the mixed liquid of the bacillus bifidus cytolytic solution and the cell debris enzymolysis solution.

10. Use of a bifidobacterium enzymatic lysate as claimed in any of claims 1 to 9 in the manufacture of a cosmetic product.