Application of ackermania kwangsiensis in preparation of products for skin whitening and / or skin aging resistance

Abstract

The invention provides application of ackermania kwangsiensis in preparation of products for skin whitening and / or skin aging resistance, and belongs to the technical field of microorganisms. A metabolite of the ackermansiella kwangsiensis has the effects of inhibiting tyrosinase activity and inhibiting generation of melanin in cells, and meanwhile, the ackermansiella kwangsiensis has the effects of maintaining the number and form of collagen fibers of skin cells and promoting increase of the number of sebaceous glands and hair follicles; and the composition has good body intestinal colonization ability and no toxic or side effect of cells. Therefore, the invention provides the application of the ackermania kwangsiensis, the ackermania kwangsiensis probiotic agent or the metabolite of the ackermania kwangsiensis in preparation of products for skin whitening and / or skin aging resistance.

Description

Technical Field

[0001] The invention belongs to the technical field of microorganisms, and particularly relates to application of Akkermansia guangxiensis in preparing skin whitening and / or anti-skin aging products. Background Art

[0002] When the human body is exposed to ultraviolet rays, human skin cells, especially keratinocytes in the first layer of the epidermis, will secrete melanocyte-stimulating hormone (MSH), which will further activate the melanin production of melanocytes in the basal layer of the epidermis, while causing the cells to lose some water, resulting in sagging skin and wrinkles, which in turn leads to natural skin aging. Since melanin is the main determinant of skin color, excessive melanin production will induce pigmentation-promoting skin diseases such as chloasma, freckles, and age spots. Melanin formation is closely related to the catalytic action of tyrosinase (TYR), tyrosinase-related protein 1 (TRP-1), and tyrosinase-related protein 2 (TRP-2). Inhibiting tyrosinase activity is the most effective way to develop melanin production inhibition products. At the same time, paying attention to skin sun protection is also an important measure to prevent skin aging. Currently known whitening and anti-aging products have the characteristics of inhibiting tyrosinase activity and moisturizing, but have side effects such as low stability and dermatitis. Summary of the invention

[0003] In view of this, the object of the present invention is to provide the use of Akkermansia guangxiensis in the preparation of skin whitening and / or anti-skin aging products.

[0004] The present invention provides application of Guangxi Akkermansia, Guangxi Akkermansia probiotics or metabolites of Guangxi Akkermansia in preparing skin whitening and / or anti-skin aging products.

[0005] Preferably, the Guangxi Akkermansia includes Guangxi Akkermansia strain N21116 and / or Guangxi Akkermansia strain N21169;

[0006] The deposit number of the Guangxi Akkermansia strain N21116 is GDMCC No: 62888;

[0007] The deposit number of the Guangxi Akkermansia strain N21169 is GDMCC No: 62889.

[0008] Preferably, the skin whitening comprises reducing the activity of tyrosinase in skin cells and inhibiting the formation of melanin.

[0009] Preferably, the anti-skin aging comprises maintaining the morphology of collagen fibers or promoting an increase in the number of hair follicles and / or sebaceous glands.

[0010] Preferably, the product includes any one of the following: medicines, health products, health foods and cosmetics.

[0011] Preferably, the Guangxi Akkermansia probiotic comprises at least one of the following dosage forms: powder, tablet, drop, capsule and aqueous solution.

[0012] Preferably, the number of viable bacteria in the Guangxi Akkermansia probiotic is not less than 1×10 8 CFU / mL or 1×10 8 CFU / g.

[0013] Preferably, the metabolites of Akkermansia Guangxi include at least one of the following forms: culture fluid of Akkermansia Guangxi, culture supernatant of Akkermansia Guangxi and purified culture supernatant of Akkermansia Guangxi.

[0014] Preferably, the method for preparing the metabolites of Akkermansia Guangxi comprises inoculating activated Akkermansia Guangxi into a fermentation medium, and anaerobic culturing at 36-38° C. for 70-74 hours to obtain a culture solution of Akkermansia Guangxi;

[0015] Alternatively, the culture solution of Akkermansia guangxi is subjected to bacterial liquid separation, and the liquid phase is collected to obtain the culture supernatant of Akkermansia guangxi;

[0016] Alternatively, the culture supernatant of Akkermansia Guangxi is free of impurities to obtain a purified product of the culture supernatant of Akkermansia Guangxi.

[0017] Preferably, the fermentation medium is an aqueous solution comprising the following ingredients: 7.2 g / L glucose, 9.0 g / L yeast extract, 6.0 g / L soy peptone, 0.5 g / L L-cysteine, 0.425 g / L sodium nitrate, 2.0 g / L N-acetylglucosamine, 4.5 g / L sodium chloride, 5.0 g / L disodium hydrogen phosphate and 8.0 g / L threonine.

[0018] The present invention provides the use of Guangxi Akkermansia, Guangxi Akkermansia probiotics or metabolites of Guangxi Akkermansia in the preparation of skin whitening and / or anti-skin aging products. The metabolites of Guangxi Akkermansia have the characteristics of inhibiting tyrosinase activity at the cellular level, and the inhibitory effect is stronger than kojic acid. Tyrosinase is a key enzyme in the formation of melanin, so the Guangxi Akkermansia or its metabolites have the ability to inhibit melanin production. At the same time, Guangxi Akkermansia has the characteristics of maintaining the normal morphology of skin tissue collagen fibers in a mouse aging model caused by D-galactose and promoting the increase in the number of sebaceous glands and hair follicles. At the same time, the Guangxi Akkermansia has no toxicity to cells, has a certain tolerance to bile salts and gastrointestinal fluid, and ensures its good colonization in the human intestine. Therefore, the present invention provides a new use of Guangxi Akkermansia or its probiotics or metabolites, which provides a new way for skin whitening and anti-skin aging. BRIEF DESCRIPTION OF THE DRAWINGS

[0019] Figure 1 The results of the tolerance test of Akkermansia guangxiensis to simulated artificial gastrointestinal fluid are shown in Figure 1. (a) is the result of the survival rate of the simulated artificial gastric fluid tolerance test, and (b) is the result of the survival of the simulated artificial intestinal fluid tolerance test.

[0020] Figure 2 This is the result of the bile salt tolerance experiment of Akkermansia guangxiensis;

[0021] Figure 3 This is the result of the effect of Akkermansia guangxiensis on the proliferation of B16 melanoma cells;

[0022] Figure 4 The results of the effects of Guangxi Akkermansia on the morphology of B16 melanoma cells; (a) is the N21116 group, (b) is the N21169 group, (c) is the ATCC BAA-835 group, (d) is the LGG group, (e) is the α-MSH stimulation group, and (f) is the blank group;

[0023] Figure 5 The results of the experiment on the relative activity inhibition of Akkermansia guangxiensis on tyrosinase in vitro;

[0024] Figure 6 The results of skin tissue repair of Akkermansia guangxiensis in D-galactose-induced mouse aging model; (a) is the blank group, (b) is the model group, (c) is the N21169 intervention group, and (d) is the N21116 intervention group.

[0025] Biomaterial Deposit Information

[0026] The Akkermansia sp. strain N21116 from Guangxi is deposited in the Guangdong Provincial Microbiological Culture Collection Center, abbreviated as GDMCC, located at 5th Floor, Building 59, No. 100 Xianlie Middle Road, Guangzhou, Guangdong Institute of Microbiology. The deposit date is December 2, 2022, and the deposit number is GDMCC No: 62888.

[0027] The Akkermansia sp. strain N21169 from Guangxi is deposited in the Guangdong Provincial Microbiological Culture Collection Center, abbreviated as GDMCC, with the address at 5th Floor, Building 59, No. 100 Xianlie Middle Road, Guangzhou, Guangdong Institute of Microbiology. The deposit date is December 2, 2022, and the deposit number is GDMCC No: 62889. DETAILED DESCRIPTION

[0028] The present invention provides application of Guangxi Akkermansia, Guangxi Akkermansia probiotics or metabolites of Guangxi Akkermansia in preparing skin whitening and / or anti-skin aging products.

[0029] In the present invention, the Guangxi Akkermansia preferably includes Guangxi Akkermansia strain N21116 and / or Guangxi Akkermansia strain N21169. The deposit number of the Guangxi Akkermansia strain N21116 is GDMCC No: 62888. The deposit number of the Guangxi Akkermansia strain N21169 is GDMCC No: 62889. The Guangxi Akkermansia strain N21116 and Guangxi Akkermansia strain N21169 are both isolated from the feces of centenarians.

[0030] In the present invention, the Guangxi Akkermansia has the property of good colonization in the intestinal tract of the body and shows good probiotic potential. In one embodiment of the present invention, artificial simulated gastric juice and intestinal juice tolerance experiments were carried out, and the results showed that the Guangxi Akkermansia has a certain ability to tolerate gastrointestinal juice; the bile salt tolerance experiment showed that the Akkermansia has a certain bile salt tolerance and has probiotic potential. The artificial simulated gastric juice was purchased from Beijing Regen Biotechnology Co., Ltd. with the item number CZ0213. The artificial small intestinal fluid was purchased from Beijing Regen Biotechnology Co., Ltd. with the item number CZ0201. The bile salt was purchased from Shanghai Aladdin Biochemical Technology Co., Ltd. with the item number B485289.

[0031] In the present invention, the Guangxi Akkermansia has good cell safety and no toxic side effects. In one embodiment of the present invention, the toxic effect detection of Guangxi Akkermansia on B16 melanoma cells includes growth inhibition and morphological damage. The experimental results show that the Guangxi Akkermansia has no inhibitory effect on the growth of B16 melanoma cells and no toxic damage to the morphology of the cells. The B16 melanoma cells were purchased from the mall Beina Chuanglian Biotechnology Co., Ltd., with the number BNCC337628.

[0032] In the present invention, the Guangxi Akkermansia probiotics preferably include at least one of the following dosage forms: powder, tablet, drop, capsule and liquid. The number of viable bacteria in the Guangxi Akkermansia probiotics is preferably not less than 1×10 8 CFU / mL or 1×10 8 CFU / g, can be 2×10 8 CFU / mL or 2×10 8 CFU / g, 4×10 8 CFU / mL or 4×10 8 CFU / g, 5×10 8 CFU / mL or 5×10 8 CFU / g, 7×10 8 CFU / mL or 7×10 8 CFU / g, 10×10 8 CFU / mL or 10×10 8 CFU / g, 20×10 8 CFU / mL or 20×10 8 CFU / g. The present invention has no special restrictions on the preparation method of the Guangxi Akkermansia probiotic agent, and the preparation method of the probiotic agent known in the art can be used.

[0033] In one embodiment of the present invention, the preparation method of Guangxi Akkermansia powder preferably expands the activated Guangxi Akkermansia to obtain the Guangxi Akkermansia bacterial liquid for bacterial liquid separation, collects the bacterial body; mixes the bacterial body with the freeze-drying protective agent, freeze-dries, and obtains Guangxi Akkermansia powder. The activation culture method of Guangxi Akkermansia preferably inoculates Guangxi Akkermansia in the activation culture medium for activation culture. The activation culture medium is preferably a liquid composite culture medium replacing the mucin component. The liquid composite culture medium replacing the mucin component is an aqueous solution containing the following components: glucose 7.2g / L, yeast extract powder 9.0g / L, soy peptone 6.0g / L, L-cysteine ​​0.5g / L, sodium nitrate 0.425g / L, N-acetylglucosamine 2.0g / L, sodium chloride 4.5g / L, disodium hydrogen phosphate 5.0g / L and threonine 8.0g / L. The temperature of the activation culture is preferably 36-38°C, more preferably 37°C. The activation culture time is preferably 10 to 14 hours, and can be 12 hours. The activation culture is preferably carried out for 2 to 4 rounds, and can be 3 rounds. The method of the expansion culture is preferably inoculated in 4 to 6L of culture medium according to the inoculation amount of 8% to 12%. The inoculation amount is preferably 10%. The volume of the culture medium can be 5L. The activation culture or expansion culture is preferably anaerobic culture. The method of separating the bacterial liquid is preferably centrifugation. The speed of the centrifugation is preferably 8000 to 12000rpm, and can be 8000 to 10000rpm. The centrifugation time is preferably 5 to 15min, and can be 8min. The centrifugation temperature is preferably 3 to 5°C, and can be 4°C. Before the bacteria are mixed with the freeze-dried protective agent, the bacteria are preferably washed with physiological saline. The mass ratio of the bacteria and the freeze-dried protective agent is preferably 1 to 2: 1 to 2, and can be 1: 1. The formula of the freeze-dried protective agent is preferably 10% skim milk, 10% trehalose, 1% sorbitol, and 5% vitamin C. The freeze-drying procedure is preferably as follows: the pre-freezing temperature is 40-50°C for 2-4 hours, the primary drying temperature is -20--30°C for 18-25 hours, and the secondary drying temperature is 25-35°C for 10-16 hours.

[0034] In the present invention, the metabolites of Akkermansia in Guangxi preferably include at least one of the following forms: culture fluid of Akkermansia in Guangxi, culture supernatant of Akkermansia in Guangxi and purified culture supernatant of Akkermansia in Guangxi. The preparation method of the metabolites of Akkermansia in Guangxi is to inoculate activated Akkermansia in Guangxi into a fermentation medium, and then culture the culture fluid of Akkermansia in Guangxi at 36-38°C for 70-74h to obtain the culture fluid of Akkermansia in Guangxi; or separate the culture fluid of Akkermansia in Guangxi through bacterial fluid, collect the liquid phase, and obtain the culture supernatant of Akkermansia in Guangxi; or remove impurities from the culture supernatant of Akkermansia in Guangxi to obtain the purified culture supernatant of Akkermansia in Guangxi. The temperature of the anaerobic culture can be 37°C. The time of the anaerobic culture can be 72h. The method of separating the bacterial fluid is preferably centrifugation. The centrifugation method is the same as the definition of the above technical solution, and will not be repeated here. The method for removing impurities from the culture supernatant of Akkermansia guangxi preferably comprises membrane filtration to remove bacterial fragments. The pore size of the filter membrane of the membrane filtration is preferably 0.2 μm.

[0035] In the present invention, the skin whitening preferably includes reducing tyrosinase activity in skin cells and inhibiting melanin formation. Tyrosinase is a key enzyme involved in the synthesis of melanin. High tyrosinase activity promotes the production of melanin, and low tyrosinase activity inhibits the production of melanin. In one embodiment of the present invention, the influence of the culture supernatant of the Guangxi Akkermansia on tyrosinase activity at the cellular level was carried out. The results show that compared with the α-MSH positive stimulation group, the tyrosinase activity of the positive drug (kojic acid) group and the two Guangxi Akkermansia groups is reduced, and Guangxi Akkermansia all shows a certain ability to inhibit tyrosinase activity, and the inhibitory effect is better than the 2mM positive drug (kojic acid) group. Two Guangxi Akkermansia are compared, and the Guangxi Akkermansia strain N21169 has a stronger ability to inhibit tyrosinase activity.

[0036] In the present invention, the anti-skin aging preferably includes maintaining the morphology of collagen fibers or promoting the increase in the number of hair follicles and / or sebaceous glands. In one embodiment of the present invention, the aging mouse model induced by D-galactose was used as the experimental subject, and the effect of Guangxi Akkermansia on the skin of aging mice was carried out. The mouse skin was stained with a Masson staining kit and the slices were observed. The results showed that the collagen fibers of the skin of mice in the N21169 intervention group and the N21116 intervention group were relatively complete and arranged more neatly than those in the model group, and the number of sebaceous glands and hair follicles increased. The D-galactose was purchased from Shanghai Haoyuan Biomedical Technology Co., Ltd., and the item number is HY-N0210. The Masson staining kit was purchased from Beijing Solebow Technology Co., Ltd., and the item number is G1340.

[0037] In the present invention, the product preferably includes any one of the following: medicines, health products, health foods and cosmetics. The Guangxi Akkermansia, Guangxi Akkermansia probiotics or the metabolites of Guangxi Akkermansia can be used to prepare skin whitening and anti-skin aging medicines, health products and health foods, and the effective viable count is not less than 11×10 8 CFU / mL or 1×10 8 CFU / g, can be 2×10 8 CFU / mL or 2×10 8 CFU / g, 4×10 8 CFU / mL or 4×10 8 CFU / g, 5×10 8 CFU / mL or 5×10 8 CFU / g, 7×10 8 CFU / mL or 7×10 8 CFU / g, 10×10 8 CFU / mL or 10×10 8 CFU / g, 20×10 8 CFU / mL or 20×10 8 CFU / g. The metabolites of Akkermansia guangxi can be used to prepare cosmetics. The metabolites of Akkermansia guangxi include extracellular metabolites and / or intracellular metabolites of the bacteria, which may be short-chain fatty acids (such as butyric acid, butyrate), total superoxide dismutase and glutathione peroxidase. The present invention has no special restrictions on the preparation method of the medicine, health care product, health food and cosmetics, and they can be prepared by conventional methods well known in the art.

[0038] The application of Akkermansia guangxi provided by the present invention in the preparation of skin whitening and / or anti-skin aging products is described in detail below in conjunction with the examples, but they should not be construed as limiting the scope of protection of the present invention.

[0039] Example 1

[0040] Experimental study on tolerance of Akkermansia spp. in Guangxi to bile salts and artificial simulated gastrointestinal fluid

[0041] 1. Strain simulated gastric juice tolerance test

[0042] After three rounds of activation of the strain using a liquid composite medium that replaces mucin components, the bacterial solution was mixed with artificial simulated gastric fluid at a volume ratio of 1:10, and the mixed solution at 0h and 3h was taken for plate coating and colony counting. The survival rate of the strain in simulated artificial gastric fluid was calculated according to Formula I.

[0043] Simulated artificial gastric fluid survival rate (%) = N3 / N0×100% Formula Ⅰ.

[0044] Wherein, N3 represents the number of viable bacteria after 3 h of culture, and N0 represents the number of viable bacteria after 0 h of culture.

[0045] 2. Strain simulated gastric juice tolerance test

[0046] The artificial gastric juice mixture after 3 hours of culture was mixed with artificial simulated intestinal juice at a volume ratio of 1:10, and the mixture at 0h and 4h was taken for plate coating and colony counting. The survival rate of the strain in simulated artificial intestinal juice was calculated according to formula II.

[0047] Simulated artificial intestinal fluid survival rate (%) = N4 / N0×100% Formula II.

[0048] Wherein, N4 represents the number of viable bacteria after 4 hours of culture, and N0 represents the number of viable bacteria after 0 hours of culture.

[0049] 3. Bile salt tolerance test

[0050] Prepare a liquid synthetic medium containing 0.2% bile salts, mix the strain in the liquid synthetic medium containing 0.2% bile salts at a volume ratio of 1:10, take the mixed solution at 0h and 4h for plate coating and colony counting. Calculate the survival rate of the strain in the simulated artificial intestinal fluid according to formula III.

[0051] Bile salt survival rate (%) = N4 / N0×100% Formula III.

[0052] Wherein, N4 represents the number of viable bacteria after 4 hours of culture, and N0 represents the number of viable bacteria after 0 hours of culture.

[0053] Results Figure 1 and Figure 2 .Depend on Figure 1 and Figure 2 It can be seen that compared with the Akk model strain (ATCC BAA-835), Guangxi Akkermansia N21116 has a stronger tolerance to simulated gastrointestinal fluid. After 3 hours of simulated gastric fluid treatment, the survival rate reached more than 80%, while the survival rate of ATCC BAA-835 was about 65%; then after 4 hours of simulated intestinal fluid treatment, the survival rate of N21116 was about 26%, and the survival rate of ATCC BAA-835 was about 22%. Guangxi Akkermansia N21116 and N21169 have a stronger tolerance to bile salts than ATCC BAA-835, and the survival rate reached 70% after 4 hours of treatment, while the survival rate of ATCC BAA-835 was about 40%. Guangxi Akkermansia has a certain tolerance to gastrointestinal fluid and bile salts.

[0054] Example 2

[0055] Toxic effects of Akkermansia guangxiensis on B16 melanoma cells

[0056] 1. CCK-8 method to detect the survival rate of B16 melanoma cells

[0057] When B16 melanoma cells grow to the logarithmic growth phase, they are digested and resuspended with trypsin and added to a 96-well plate at a number of 5,000 cells per well. They are cultured overnight in a cell culture incubator at 37°C and 5% CO2. The activated strain is taken, and the bacterial concentration is 1×10 8 CFU / mL, centrifuge at 12000rpm for 15min, and take the fermentation supernatant for use. Discard the cell culture supernatant, add 10% cell supernatant and 90% cell culture medium to the wells, incubate for 48h, add 10μL CCK-8 solution to each well, incubate the culture plate in the incubator for 4h, and measure the absorbance at 450nm with an enzyme marker. Set Akk model strain (ATCC BAA-835) and rhamnosus LGG as controls. Calculate the cell proliferation rate according to formula IV.

[0058] Cell viability (%) = [A (added bacteria) - A (blank)] / [A (0 added bacteria) - A (blank)] × 100% Formula IV.

[0059] Wherein, A(with bacteria) represents the absorbance of the wells with cells, CCK-8 solution and bacterial supernatant, A(blank) represents the absorbance of the wells with culture medium and CCK-8 solution but no cells, and A(0 with bacteria) represents the absorbance of the wells with cells, CCK-8 solution but no bacterial supernatant.

[0060] 2. Effects of Akkermansia guangxiensis on the morphology of B16 melanoma cells

[0061] When B16 melanoma cells grow to the logarithmic growth phase, they are digested and resuspended with trypsin and added to a 6-well plate at a cell count of 500,000 per well. They are cultured overnight in a 37°C, 5% CO2 cell culture incubator. 600 nm nM α-MSH is added to each well for incubation. The activated strain is taken, and the bacterial concentration is 1×10 8 CFU / mL, centrifuged at 12000rpm for 15min, and the fermentation supernatant was taken for use. After the cells were incubated with α-MSH for 24h, 10% cell supernatant and 90% cell culture medium were added to the wells, and after incubation for 48h, the cell morphology was observed using an inverted biological microscope.

[0062] Results Figure 3 and Figure 4 .Depend on Figure 3 and Figure 4It can be seen that after 48 hours of intervention of 10% Guangxi Akkermansia fermentation supernatant on B16 melanoma cells, there was no statistical difference in cell activity between the blank group and the control group, indicating that 10% Guangxi Akkermansia fermentation supernatant had no toxic effect on the growth of B16 melanoma cells. In addition, B16 melanoma cells grew well before α-MSH stimulation. After adding α-MSH stimulation, the cells in the stimulated group were Figure 4 The cells in the blank wells shown in (f) all became slightly rounded. After adding the supernatants of different bacteria for 48 hours, Figure 4 There was no significant difference in the cell morphology between (a) N21116 group, (b) N21169 group, (c) ATCC BAA-835 group and (d) LGG group and (e) the single positive stimulation well cell morphology, and no obvious cell shedding was found.

[0063] Example 3

[0064] When B16 melanoma cells grow to the logarithmic growth phase, they are digested and resuspended with trypsin and added to a 6-well plate at a cell count of 500,000 per well. They are cultured overnight in a 37°C, 5% CO2 cell culture incubator. 600 nm nM α-MSH is added to each well for incubation. The activated strain is taken, and the bacterial concentration is 1×10 8 CFU / mL, centrifuged at 12000rpm for 15min, and the fermentation supernatant was taken for use. After the cells were incubated with α-MSH for 24h, 10% cell supernatant and 90% cell culture medium were added to the wells. After continuing to incubate for 36h, the cell tyrosinase activity was measured, and 2mM / well kojic acid was used as a control. After washing the cells 3 times with PBS solution, 300μL of 50mM PBS (pH=7.5) buffer containing 1.0% Triton X-100 and 0.1mM PMSF was added to each empty space, placed in -80℃ for 30min, and then thawed at room temperature. After repeated freezing and thawing 3 times, the liquid was centrifuged at 12000rpm for 15min, 100μL of supernatant was taken and added to the ELISA plate, and 100μL of 500μg / mL L-Dopa solution was added to each well, incubated at 37℃ for 40min, and the absorbance was measured at 475nm. The relative activity of tyrosinase was calculated according to formula V.

[0065] Tyrosinase relative activity (%) = A1 / A0×100% Formula V.

[0066] Where A0 is the absorbance of the blank control group of cells without α-MSH stimulation, and A1 is the absorbance of the cells in the sample group.

[0067] Results Figure 5 .Depend on Figure 5It can be seen that compared with the blank group, the tyrosinase activity in the α-MSH stimulation group increased significantly, indicating that the model was successfully established; compared with the positive stimulation group, the tyrosinase activity in the other intervention groups decreased, and the Guangxi Akkermansia showed a certain ability to inhibit tyrosinase activity, and was slightly better than the 2mM kojic acid intervention group. Among them, Guangxi Akkermansia N21169 had a stronger ability to inhibit tyrosinase activity.

[0068] Example 4

[0069] Preparation method of freeze-dried Akkermansia guianensis agent

[0070] Three rounds of activation of Akkermansia guangxiensis N21116 and 21169 were performed using a liquid composite culture medium that replaced mucin components, and the culture was gradually expanded to 5 L according to a 10% inoculation volume. The bacterial liquid was collected and centrifuged at 4°C, the supernatant was discarded, the bacteria were washed with physiological saline, and centrifuged again. The wet weight of the bacteria was weighed, and the whole process was operated under anaerobic conditions.

[0071] Mix the lyophilized protective agent with the bacteria, divide into EP tubes, and freeze in liquid nitrogen for 1 hour. Put the divided into freeze-drying bottles, freeze-dry with freeze-drying agent, and finally obtain freeze-dried powder. Count using the plate count method, store the remaining freeze-dried powder at -80℃, and resuspend with physiological saline when using.

[0072] Example 5

[0073] Animal experiments

[0074] A total of 40 SPF 6-8 week old female C57BL / 6J mice were randomly divided into 4 groups after 1 week of adaptive feeding, with 10 mice in each group. The specific groups are as follows:

[0075] Normal group: basal feed and daily intragastric administration of normal saline;

[0076] Model group: basal diet, daily subcutaneous injection of D-galactose (200 mg / kg) and daily intragastric administration of normal saline;

[0077] N21116 intervention group: basal diet, daily subcutaneous injection of D-galactose (200 mg / kg) and daily oral gavage of N21116 (1×10 8 CFU / mL);

[0078] N21169 intervention group: basal diet, daily subcutaneous injection of D-galactose (200 mg / kg) and daily oral gavage of N21169 (1×10 8 CFU / mL).

[0079] The experiment lasted for 8 weeks. The skin tissues were collected and fixed, dehydrated and embedded, and then Masson staining was performed according to the instructions of the kit.

[0080] Results Figure 6 .Depend on Figure 6 It can be seen that Figure 6 (b) Model group mice Figure 6 (a) The collagen fibers in the skin of mice in the blank group were unevenly distributed, with reduced content, broken fibers, and disordered arrangement. There were also relatively fewer hair follicles and sebaceous glands. Figure 6 (c) N21169 intervention group mice and Figure 6 (d) The collagen fibers in the skin of mice in the N21116 intervention group were relatively complete and arranged more neatly than those in the model group. The number of sebaceous glands and hair follicles increased. The collagen fibers in the N21116 intervention group were tighter than those in the N21169 intervention group.

[0081] From the results of the above examples, it can be seen that Akkermansia Guangxi has a certain tolerance to gastrointestinal fluid and bile salts, has the potential to colonize in the intestines of the body, and has probiotic potential. At the same time, Akkermansia Guangxi and its probiotic agents or their metabolites are used in the preparation of functional products for skin whitening and anti-skin aging.

[0082] The above is only a preferred embodiment of the present invention. It should be pointed out that for ordinary technicians in this technical field, several improvements and modifications can be made without departing from the principle of the present invention. These improvements and modifications should also be regarded as the scope of protection of the present invention.

Claims

1. Use of Akkermansia guangxi, Akkermansia guangxi probiotics or metabolites of Akkermansia guangxi in the preparation of skin whitening and / or anti-skin aging products.

2. The use according to claim 1, characterized in that: The Guangxi Akkermansia includes Guangxi Akkermansia strain N21116 and / or Guangxi Akkermansia strain N21169; The deposit number of the Guangxi Akkermansia strain N21116 is GDMCC No: 62888; The deposit number of the Guangxi Akkermansia strain N21169 is GDMCC No: 62889.

3. The application according to claim 1, characterized in that: The skin whitening comprises reducing the activity of tyrosinase in skin cells and inhibiting the formation of melanin.

4. The use according to claim 1, characterized in that: The anti-skin aging includes maintaining the morphology of collagen fibers or promoting the increase in the number of hair follicles and / or sebaceous glands.

5. The use according to claim 1, characterized in that: The products include any of the following: medicines, health products, health foods and cosmetics.

6. The use according to any one of claims 1 to 5, characterized in that: The Guangxi Akkermansia probiotics include at least one of the following dosage forms: powder, tablet, drop, capsule and liquid.

7. The use according to any one of claims 1 to 5, characterized in that: The number of viable bacteria in the Guangxi Akkermansia probiotic agent is not less than 1×10 8 CFU / mL or 1×10 8 CFU / g.

8. The use according to claim 1, characterized in that: The metabolites of Akkermansia Guangxi include at least one of the following forms: culture fluid of Akkermansia Guangxi, culture supernatant of Akkermansia Guangxi and purified culture supernatant of Akkermansia Guangxi.

9. The use according to claim 1, characterized in that: The method for preparing the metabolites of Akkermansia guangxi comprises inoculating activated Akkermansia guangxi into a fermentation medium, and performing anaerobically culturing at 36-38° C. for 70-74 hours to obtain a culture solution of Akkermansia guangxi; Alternatively, the culture solution of Akkermansia guangxi is subjected to bacterial liquid separation, and the liquid phase is collected to obtain the culture supernatant of Akkermansia guangxi; Alternatively, the culture supernatant of Akkermansia Guangxi is free of impurities to obtain a purified product of the culture supernatant of Akkermansia Guangxi.

10. The use according to claim 9, characterized in that: The fermentation medium is an aqueous solution containing the following ingredients: 7.2 g / L glucose, 9.0 g / L yeast extract, 6.0 g / L soy peptone, 0.5 g / L L-cysteine, 0.425 g / L sodium nitrate, 2.0 g / L N-acetylglucosamine, 4.5 g / L sodium chloride, 5.0 g / L disodium hydrogen phosphate and 8.0 g / L threonine.

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