NOVEL BIFIDOBACTERIUM LONGUM STRAIN OR COSMETIC COMPOSITION COMPRISING SAME
20220143112 · 2022-05-12
Assignee
Inventors
- Yong Hyun Lee (Daejeon, KR)
- Bo Ram BECK (Daejeon, KR)
- Ji Hee KANG (Daejeon, KR)
- Do Yeun JEONG (Daejeon, CN)
- Gun Seok PARK (Daejeon, KR)
- Sung Hoon IM (Daejeon, KR)
- Won Ho SONG (Daejeon, KR)
Cpc classification
A61P29/00
HUMAN NECESSITIES
A61K8/99
HUMAN NECESSITIES
C12R2001/01
CHEMISTRY; METALLURGY
A23L33/135
HUMAN NECESSITIES
International classification
A23L33/135
HUMAN NECESSITIES
A61P29/00
HUMAN NECESSITIES
Abstract
The present invention relates to a novel Bifidobacterium longum ATG-F5 strain. The F5 strain is a functional beneficial bacterium which is safe from antibiotic resistance, improves skin health through antibacterial activity against the skin pathogen Cutibacterium acnes, an antioxidant effect on radicals, skin barrier improvement, and anti-inflammatory effects, and enhances skin beauty through a skin whitening function, a wrinkle improvement function, and a dry skin alleviation function. The strain may be used in a cosmetic composition or health functional food.
Claims
1. A Bifidobacterium longum ATG-F5 strain deposited under accession no. KCTC13828BP.
2. The Bifidobacterium longum ATG-F5 strain of claim 1, wherein the strain has a skin barrier strengthening function or a dry skin alleviation function.
3. The Bifidobacterium longum ATG-F5 strain of claim 1, wherein the strain has an anti-acne function.
4. The Bifidobacterium longum ATG-F5 strain of claim 1, wherein the strain has an antioxidant function.
5. The Bifidobacterium longum ATG-F5 strain of claim 1, wherein the strain has a skin whitening function.
6. The Bifidobacterium longum ATG-F5 strain of claim 1, wherein the strain has a wrinkle improvement function.
7. The Bifidobacterium longum ATG-F5 strain of claim 1, wherein the strain has anti-inflammatory function.
8. A cosmetic composition comprising the Bifidobacterium longum ATG-F5 strain of claim 1 which has functions comprising skin barrier strengthening, dry skin alleviation, anti-inflammatory, anti-acne, antioxidant, and wrinkle improvement.
9. The cosmetic composition of claim 8, wherein a formulation of the cosmetic composition is selected from the group consisting of skin lotion, skin softener, skin toner, astringent, milk lotion, moisture lotion, nourishing lotion, massage cream, nourishing cream, moisture cream, hand cream, foot cream, neck cream, foundation, essence, pack, soap, cleansing foam, cleansing lotion, cleansing cream, body lotion, hair shampoo, hair treatment, hair conditioner, and body cleanser.
10. A health functional food comprising the Bifidobacterium longum ATG-F5 strain of claim 1 which has functions comprising skin barrier strengthening, dry skin alleviation, anti-inflammatory, anti-acne, antioxidant, and wrinkle improvement.
11. The health functional food of claim 10, wherein the health functional food is selected from the group consisting of meat, sausage, bread, candies, snacks, noodles, ice cream, dairy products, fermented milk, soup, ionized beverage, beverage, alcoholic beverage, gum, tea, and vitamin complex.
12. A composition for preventing or treating a disease selected from the group consisting of dry skin, acne, and an inflammatory disease comprising the Bifidobacterium longum ATG-F5 strain of claim 1.
13. The composition of claim 12, wherein the inflammatory disease is selected from the group consisting of inflammatory bowel disease, inflammatory collagen vascular disease, glomerulonephritis, inflammatory skin diseases, sarcoidosis, retinitis, gastritis, hepatitis, enteritis, arthritis, tonsillitis, sore throat, bronchitis, pneumonia, pancreatitis, blood poisoning, cystitis, nephritis, and neuritis.
14. A method of preventing or treating a disease selected from the group consisting of dry skin, acne, and an inflammatory disease by administering a composition comprising the Bifidobacterium longum ATG-F5 strain of claim 1 to a subject in need thereof.
15. The method of claim 14, wherein the inflammatory disease is selected from the group consisting of inflammatory bowel disease, inflammatory collagen vascular disease, glomerulonephritis, inflammatory skin diseases, sarcoidosis, retinitis, gastritis, hepatitis, enteritis, arthritis, tonsillitis, sore throat, bronchitis, pneumonia, pancreatitis, blood poisoning, cystitis, nephritis, and neuritis.
Description
DESCRIPTION OF DRAWINGS
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BEST MODE
[0104] Hereinafter, preferred embodiments of the present invention will be described in detail. However, the present invention is not limited to the embodiments described herein and may be embodied in other forms. The embodiments are provided so that the present disclosure will be thorough and complete and will fully convey the spirit of the invention to those skilled in the art.
Example 1. Culture of Microorganisms for Functional Evaluation
[0105] Bifidobacterium longum (hereinafter, referred to as F5) was isolated from donated neonatal feces (of babies born in 2018, Daejeon, Korea). To this end, neonatal feces was diluted with 0.9% (w/v) saline by 10-fold serial dilution method, the resulting solution was smeared on a Bifidobacterium selective (BS, MBcell Seoul, Korea) solid medium, and bacteria were incubated at 37° C. for 48 hours. The colonies of bacteria generated in the BS medium were observed with a microscope. Through the observation, bacteria not exhibiting a catalase reaction and bacillus-type bacteria were selected and named ATG-F5 strain (hereinafter simply referred to as F5). For experiment, the F5 strain was first cultured in a BL or MRS-cys agar solid medium, followed by inoculation of purely isolated colony in a broth liquid medium. The microorganisms were cultured at 37° C. overnight (i.e., for 16 to 20 hours).
Example 2. Confirmation of Characteristics of F5 Strain
Example 2-1. 16S rRNA Sequencing of F5 Strain
[0106] The 16S rRNA sequence analysis of the F5 strain was performed by Solgent Co., Ltd. (located in Dajeon, Korea) at request. Primers for sequencing analysis were 27F (5′-AGA GTT TGA TCC TGGCTC AG-3′), 518F (5′-CCA GCA GCC GCG GTA ATA C-3′), 907R (5′-CCGTCA ATT CMT TTR AGT TT-3′), 1492R (5′-GGT TAC CTT GTT ACG ACT T-3′), and the nucleotide sequences were read for a total of 4 times. The contig nucleotide sequence derived through the nucleotide sequence alignment of each reading was analyzed using the BLAST online tool (https://blast.ncbi.nlm.nih.gov/Blast.cgi) of National Center For Biotechnology Information (NCBI).
[0107] The nucleotide sequence of SEQ ID NO: 1 (see
[0108] Accordingly, on Mar. 18, 2019, the strain of the present invention was deposited with the Korea Institute of Biological Resources under accession number: KCTC13828BP.
Example 2-3. Sugar Fermentation Pattern of F5 Strain
[0109] In addition, a slightly modified API50 CHL test (BioMerieux, France) was performed to determine the sugar fermentation pattern. Briefly, L-cystein was added to a 10 mL API 50CHL medium (BioMerieux, France) at a concentration of 0.5 μg/ml, and the pH was adjusted to about 6.7. Next, the purely cultured F5 strain was suspended to have an OD.sub.600 absorbance of about 0.5, and the suspension culture solution was inoculated into each cupule of the API 50CH test strip and cultured at 37° C. The sugar fermentation results were checked when 48 hours and 72 hours passed after the inoculation.
[0110] As shown in Table 1, the result of the evaluation of sugar fermentation using an API kit reveals that the F5 strain degrades L-arabinose, ribose, galactose, glucose, fructose, mannose, mannitol, sorbitol, methyl-αD-mannopyranoside, N-acetylglucosamine, amygdalin, arbutin, esculin, salicin, cellobiose, maltose, lactose, melibiose, trehalose, melezitose, raffinose, and turanose. The F5 strain also weakly degrades D-arabinose, methyl-αD-glucopyranoside, gentibiose, L-fucose, and gluconate.
TABLE-US-00001 TABLE 1 Carbohydrates Bifidobacterium longam ATG-F5 Glycerol − Erythritol − D-Arabinose w L-Arabinose + Ribose + D-Xylose − L-Xylose − Adonitol − Methyl-βD-Xylopyranoside − Galactose + Glucose + Fructose + Mannose + Sorbose − Rhamnose − Dulcitol − Inositol − Mannitol + Sorbitol + Methyl-αD-Mannopyranoside + Methyl-αD-Glucopyranoside w N-Acetylglucosamine + Amygdalin + Arbutin + Esculin + Salicin + Cellobiose + Maltose + Lactose + Melibiose + Sucrose + Trehalose + Inulin − Melezitose + Raffinose + Starch − Glycogen − Xylitol − Gentiobiose w Turanose + Lyxose − Tagatose − D-Fucose − L-Fucose w D-Arabitol − L-Arabitol − Gluconate w 2-keto-glugonate − 5-keto-gluconate − * Sugar fermentation pattern of Bifidobacterium longum ATG-F5 (Positive: +, Weakly positive: w, Negative: −)
Example 3. Evaluation of Antibacterial Activity of F5 Strain
[0111] In order to check the antibacterial activity of the F5 strain among various functional evaluation items associated with skin irritation, the antibacterial activity of the F5 strain against a total of two types of infectious or opportunistic bacteria, specifically, acne causative bacteria (Cutibacterium acnes, KCTC5012 and KCTC3314) was evaluated through a disc test. Through the disc test, a clear zone was identified. Two types of bacteria cultured overnight in BL broth (MBcell Seoul, Korea) plate media were each suspended in 1× phosphate buffered saline (PBS) at an OD.sub.600 absorbance of about 0.8. Each suspension was absorbed with a sterile cotton swab, was spread and dried over on an agar medium in which BL and MRS were mixed in a ratio of 1:1 for an antibacterial activity test, and an 8 mm paper disc (Advantec, Japan) was attached to the dried agar medium for testing. The F5 bacterial solutions cultured in the BL broth for 18 to 20 hours was inoculated on paper discs, by 35 μl per disc, were dried for about 3 minutes, and incubated at 37° C. The diameter of the clear zone generated after the incubation was measured, and 8 mm was subtracted from the measured clear zone diameter to obtain a final value.
[0112] Regarding antibacterial activity, the antibacterial activity of the F5 strain was evaluated using Cutibacterium acnes (KCTC 5012 and 3314) strains. The F5 strain exhibited an antibacterial activity of a 9 to 10 mm clear zone and an antibacterial activity of a 5 to 6 mm clear zone with respect to KCTC5012 and KCTC3314, respectively (see Table 2 and
TABLE-US-00002 TABLE 2 Clear zone against Clear zone against Experiment Cutibacterium acnes Cutibacterium acnes round “KCTC 5012” “KCTC 3314” First round 9 mm 6 mm Second 9 mm 5 mm round Third round 10 mm 6 mm Average 9.3 mm 5.7 mm
Example 4. Antibiotic Resistance Safety of F5 Strain
[0113] An antibiotic test was performed using an E-test strip (BioMerieux, France) or an MIC test strip (Liofilchem, Italy) of 9 antibiotics including ampicillin, vancomycin, gentamicin, kanamycin, streptomycin, clindamycin, erythromycin, tetracycline, and chloramphenicol, and the minimum inhibitory concentration (MIC) values were obtained.
[0114] For this, the F5 strain was suspended to be at an OD.sub.600 absorbance of about 0.8 and smeared on a BL solid medium using a sterilized cotton swab. The solid medium on which the F5 strain was smeared was dried for about 3 minutes, and the E-test strip or MIC test strip was mounted and incubated at 37° C. for 24 to 48 hours.
[0115] The guidelines published by the European food safety authority (EFSA) were referenced for the types of antibiotics and the minimum inhibitory concentration that can be considered safe (EFSA Panel on Additives and Products or Substances used in Animal Feed, 2012).
[0116] As shown in Table 3, the results of measuring the susceptibility of the 9 antibiotics were found to be consistent with the standard values of the guidelines presented by the EFSA. The unit of each numerical value in Table 3 is μg/ml.
TABLE-US-00003 TABLE 3 Strains AMP VAN GEN KAN STR CD ERY TET CM Bifidobacterium 0.75 0.75 8 NR 8 0.016 0.023 0.5 0.38 longum ATG-F5 EFSA standard 2 2 64 NR 128 1 1 8 4 AMP, ampicillin; VAN, vancomycin; GEN, gentamicin; KAN, kanamycin; STR, streptomycin; CD, clindamycin; ERY, erythromycin; TET, tetracycline; CM, chloramphenicol; NR, not required.
Example 5. Preparation of Cell Lysate of F5 Strain
[0117] To evaluate each function, a 10-fold concentrated cell free supernatant (CFCS) was prepared.
[0118] To this end, the culture medium of the F5 strain was centrifuged at 4,000 rpm for 25 minutes so that the culture medium is separated into the F5 strain and the culture supernatant. The pH of the supernatant was adjusted to 7 to 8 and filtered using a 0.2 μm-pore syringe filter (Satorious, Germany). The filtered solution was stored at −20° C. The cells were suspended in 1×PBS, washed with vortex, centrifuged at 4° C. and 4,000 rpm for 5 minutes to remove the remaining culture medium, and suspended again in 1×PBS. Lysozyme (Sigma-Aldrich, Germany) was dispensed to the suspended cells at a concentration of 500 μg/ml, and the resulting suspension was incubated for 2 hours in an incubator at 37° C., and then was introduced into a sonicator (70 W, 30 seconds, and 30 cycles) to undergo ultrasonic cytolysis. As a result, a lysate was prepared. Next, the lysate was heat-treated at 60° C. for 45 minutes to kill bacteria that may still remain alive in the lysate. The 60° C. heat treatment was performed in a water bath to minimize protein denaturation.
[0119] Thereafter, the cell lysate was stored at −20° C. and thawed for the experiment. The cell lysate was used for all subsequent experiments. In addition, the moisture content of the suspension was measured to dispense the suspension at different concentrations relative to the solid content during use, and the solid content was calculated for dilution.
Example 6. Confirmation of Antioxidant Function of F5 Strain
[0120] A radical scavenging experiment using ABTS ([2, 2′-Azino-Bis (3-Ethylbenzthiazoline-6-sulfonic acid], Sigma-Aldrich, Germany) was planned to measure the antioxidant function of the F5 strain. This is an experiment using the principle of discoloration from turquoise to colorless when the cationic radicals of the ABTS are removed by reacting with antioxidant materials (refer to Nam et al., 2015).
[0121] In this experiment, the antioxidant ability of the lysate of the F5 strain was measured using the ABTS solution. To prepare for the ABTS experiment, a mixed solution was prepared by mixing 14 mM ABTS stock solution and 4.9 mM potassium persulfate in a 1:1 ratio, and a dark reaction was carried out overnight so that the mixed solution turns into blue-green. After that, a working solution was prepared by diluting the solution to exhibit an absorbance of about 0.7 at an OD 734 wavelength. Next, 10% (v/v) of the lysate sample of the F5 strain was added to the working solution, and reacted in the dark for 10 minutes. Next, the absorbance of the resulting solution was measured at a wavelength of 734 nm, and each measurement value was calculated by the following formula.
ABTS radical scavenging activity (%)={1−(OD sample/OD control)}×100
[0122] Using this method, the antioxidant activity of the Bifidobacterium longum ATG-F5 lysate measured by the ABTS radical removal rate is shown in
[0123] Referring to
Example 7. Evaluation of Melanin Inhibition and Whitening Function Using Tyrosinase Inhibitor of F5 Strain
[0124] To evaluate whitening function, B16F10 Melanoma cells, which is a cell line of a mouse model, was used. B16F10 is a representative cell line for evaluation of the whitening function. The whitening function is evaluated using a mechanism in which in the cell signaling pathway, L-Tyrosine starts from the tyrosinase activation pathway and increases melanin production at the last reaction point.
[0125] For this evaluation, B16F10 was cultured in a 75 cm 2 flask (Thermo fisher, USA) and in a culture medium prepared by adding 10% (v/v) fetal bovine serum (Sigma-Aldrich, Germany) and 1% (v/v) penicillin/streptomycin cocktail (Sigma-Aldrich, Germany) to Gibco® Dulbecco modified eagle medium (DMEM, Gibco, USA). Then, when the cells grew about 80%, the cells were harvested and subcultured. The cells were seeded in a 6-well plate at a concentration of 2×10.sup.6 cells/ml, and then incubated in an incubator under 5% CO.sub.2 conditions for 24 hours.
[0126] After that, the culture medium was removed and the plate was washed using Dulbecco's Phosphate Buffered Saline (DPBS, Gibco, USA). Then, a control group (untreated) and a test group treated with α-melanocyte stimulating hormone (α-MSH, Sigma-Aldrich, USA) were prepared. The α-MSH increases the production of melanin, thereby causing skin discoloration. To the test group, 200 nmol/ml of α-MSH and 50, 100, and 500 μg/ml of the cell lysate were dispensed at each concentration. The treatment time of the α-MSH and the cell lysate of the strain was 24 hours, and the cells were incubated at 37° C. in an incubator in an 5% C.sub.2 environment.
[0127] After 24 hours, the supernatant in each well was removed, and washing was performed using DPBS. 500 μl of 0.1M PBS added with 1% (v/v) triton X-100 (Daejung, Korea) was dispensed into the washed wells to obtain cells using a microtube. The cells were centrifuged, the supernatant was separately stored, and the pellets were collected. Next, using the collected cell pellets, melanin production was evaluated. 1N NaOH was added to pure pellets. The cell pellets were melted in an 80° C. water bath for about 1 hour, then put into ice to cool, and was subjected to absorbance measurement at a wavelength of 405 nm. The analysis was performed through relative comparison.
[0128] As a result, when melanin production was induced by stimulating tyrosinase activity with α-MSH using Mouse B16F10 melanoma cells, melanin production was reduced according to the treatment concentration of the cell lysate of the F5 strain as shown in
[0129] After collecting the cell pellets, the supernatant collected separately was used for tyrosinase inhibitor measurement. 40 μl of the supernatant and 160 μl of dihydroxyphenylalanine (L-DOPA, Sigma, USA) dissolved at 2 μg/ml in 0.1M PBS were mixed and reacted for 1 hour in an incubator at 37° C. After that, an ELISA reader (BioTek, USA) was used to measure the absorbance at 475 nm to show relative the inhibitory ability of the tyrosinase inhibitor.
[0130] Accordingly, as shown in the result of
Example 8. Evaluation of Pro-Collagen Increase and Wrinkle Improvement Function Through Inhibition of MMP-1 by F5 Strain
[0131] To evaluate a wrinkle improvement function, a human model cell line, Human Dermal Fibroblast (HDF) cell line, was used. A Gibco® Dulbecco modified eagle medium (DMEM, Gibco, Germany) supplemented with 10% (v/v) fetal bovine serum (Sigma-Aldrich, Germany) and 1% (v/v) penicillin/streptomycin cocktail (Sigma-Aldrich, Germany) USA) was used to culture the cells. In addition, for the culture of the cells, a 75 cm 2 Flask (Thermo fisher, USA) was used. When the cells grew about 80% in the 75 cm 2 flask, the cells were harvested and subcultured.
[0132] Cells for the experiment were seeded in a 24-well plate at a concentration of 2×10.sup.5 cells/ml, and then cultured using an incubator under 5% CO.sub.2 conditions for 24 hours. After that, the culture medium was removed and the plate was washed using Dulbecco's Phosphate Buffered Saline (DPBS, Gibco, USA). A control group (untreated group) and a test group treated with 10 ng/ml of TNF-α (Human Tumor necrosis factor alpha, Sigma-Aldrich, USA) were prepared. The TNF-α increases MMP-1 to create skin wrinkles.
[0133] For the test group, 50, 100, and 500 μg/ml of the cell lysate of the F5 strain along with 10 ng/ml of TNF-α were dispensed at each concentration. The lysate of the TNF-α and the F5 strain were treated for 24 hours, and incubated at 37° C. in a 5% CO.sub.2 condition using an incubator.
[0134] After 24 hours, the supernatant in each well was collected, and enzyme-linked immunosorbent assay (ELISA) for matrix metalloproteinase-1 (MMP-1) and Type 1 collagen was performed. MMP-1 was relatively quantified using Human Pro-MMP-1 Quantikine ELISA Kit (R&D systems, USA), and Type 1 collagen was relatively quantified using Human Procollagen Type I C-peptide (PIP) EIA Kit (Takara, JAPAN).
[0135] As a result, as shown in
[0136] In particular, as shown in
Example 9. Evaluation of Skin Barrier Strengthening or Skin Dryness Relief Function of F5 Strain
[0137] Human keratinocyte, HaCaT cell line, was used for skin function evaluation. The cells were cultured in the same way as described above. When the cells grew about 80% in a 75 cm 2 flask, the cells were harvested and subcultured. The cells were seeded on a 10 cm round plate at a concentration of ×10.sup.6 cells/ml, and then cultured using an incubator in a 5% CO.sub.2 condition for 24 hours. Thereafter, the culture medium was removed and the plate was washed with Dulbecco's Phosphate Buffered Saline (DPBS, Gibco, USA). The cell lysate of the F5 strain was dispensed at concentrations of 50, 100, and 500 μg/ml. The control group was not treated at all. The lysate of the F5 strain was treated for 24 hours, and cultured using an incubator at 37° C. in a 5% CO.sub.2 condition.
[0138] Next, the plate was washed, and the total RNA of the cells was isolated using TRIzol® Reagent (Ambion, USA). After the isolated RNA was quantified to synthesize cDNA, it was synthesized into cDNA using SuperScript™ IV First-Strand Synthesis System (Invitrogen, USA).
[0139] After that, to identify skin-related functional genes, quantitative real-time PCR (qRT-PCR) was performed using oligo primers for occludin (OCLN) and claudin 4 (CLDN4) related to skin barrier, and Hyaluronan synthase 2 (HAS2) related to skin dryness. For the PCR analysis, a fast real-time PCR system, Applied Biosystems 7500, was used. In this case, β-actin was used as a housekeeping gene. The list of primers used in the experiment is shown in Table 4.
TABLE-US-00004 TABLE 4 Base sequence of primers used to measure gene expression Sense Anti-sense Gene (5′.fwdarw.3′) (5′.fwdarw.3′) Size (bp) β-actin TCTACGAGGGG GGATGCCACAG 330 TATGCCCTCC GACTCCATGC OCLN GACTTCAGGCA GCCAGTTGTGT 132 GCCTCGTTAC AGTCTGTCTCA CLDN4 TGGGGCTACAG GGTCTGCGAGG 145 GTAATGGG TGACAATGTT HAS2 CTCTTTTGGAC AGGGTAGGTTA TGTATGGTGCC GCCTTTTCACA 205
[0140] As a result, as shown in
[0141] Therefore, the lysate of the F5 strain of the present invention has a function of remarkably strengthening the skin barrier by engaging in contact between skin cells due to the increase in the expression of OCLN and CLDN4. Similarly, it can be confirmed that the cell lysate of the F5 strain stimulates the expression of Hyaluronan synthase 2 (HAS2) to increase the production of Hyaluronic acid (HA) in the skin, thereby alleviating skin dryness.
Example 10. Evaluation of Anti-Inflammatory Function of F5 Strain
[0142] For the evaluation of the anti-inflammatory function, the expression level of IL-10 was checked. The RAW264.7 cell line, which is a murine macrophage, was used. As the culture method, the HDF cell culture method described above was used. The cells were harvested and subcultured when they grew to about 80% in a 75 cm 2 flask. The cells were seeded in a 24-well plate at a concentration of 1×10.sup.6 cells/ml, and then were incubated in an incubator in a 5% CO.sub.2 condition for 24 hours. First, as a control group, the untreated cells were prepared. As a positive control group, the cells with treated with only LPS (Lipopolysaccharide, Sigma-Aldrich, USA) was prepared. In addition, a group treated with the cell lysate of the F5 strain alone and a group treated with 100 μg/ml of the cell lysate of the F5 strain and 1 μg/ml of LPS were also prepared. After 24 hours, the supernatant in each well was collected and relatively quantified using Interleukin 10 (IL-10, R&D, USA).
[0143] The results are shown in
Cosmetic Formulation Example 1. Preparation of Skinner Toner −1
[0144] Using the composition of Table 5 below, a skin toner (100 g) containing a lyophilized product of the cell lysate of the Bifidobacterium longum ATG-F5 strain was prepared according to a conventional method.
TABLE-US-00005 TABLE 5 Raw material Content (g) Lyophilized product of cell lysate of 3.0 Bifidobacterium longum ATG-F5 strain Butylene glycol 2.0 Propylene glycol 2.0 Polyoxyethylene (60) hydrogenated castor oil 1.0 Ethanol 10.0 Triethanolamine 0.1 Antiseptic Trace Pigment Trace Perfume Trace Purified water Remainder
Cosmetic Formulation Example 2. Preparation of Moisture Lotion
[0145] After extracting the lyophilized product of the cell lysate of the Bifidobacterium longum ATG-F5 strain with 70% (v/v) aqueous ethanol solution, a solvent-free concentrate was obtained, and a moisture lotion (100 g) containing the concentrate as shown in Table 6 was prepared according to a conventional method.
TABLE-US-00006 TABLE 6 Raw material Content (g) Lyophilized product of cell lysate of 1.0 Bifidobacterium longum ATG-F5 strain Sitosterol 1.7 Polyglyceryl 2-oleate 1.5 Ceteares 1.2 Cholesterol 1.5 dicetyl phosphate 0.4 Concentrated glycerin 5.0 Sunflower Oil 10.0 Carboxyvinyl Polymer 0.2 Xanthan Gum 0.3 Antiseptic Trace Perfume Trace Purified water Remainder
Cosmetic Formulation Example 3. Preparation of Nutrient Cream
[0146] Using the composition of Table 7 below, a nutrient cream (100 g) containing a lyophilized product of the cell lysate of the Bifidobacterium longum ATG-F5 strain was prepared according to a conventional method.
TABLE-US-00007 TABLE 7 Raw material Content (g) Lyophilized product of cell lysate of 5.0 Bifidobacterium longum ATG-F5 strain Sitosterol 4.0 Polyglyceryl 2-oleate 3.0 Ceramide 0.7 Ceteares-4 2.0 Cholesterol 3.0 Dicetyl phosphate 0.4 Concentrated glycerin 5.0 Sunflower Oil 22.0 Carboxyvinyl Polymer 0.5 Triethanolamine 0.5 Antiseptic Trace Perfume Trace Purified water Remainder
Cosmetic Formulation Example 4. Preparation of Essence
[0147] Using the composition of Table 8 below, an essence (100 g) containing a lyophilized product of the cell lysate of the Bifidobacterium longum ATG-F5 strain was prepared according to a conventional method.
TABLE-US-00008 TABLE 8 Content Raw material (g) Lyophilized product of cell lysate of 1.0 Bifdobacterium longum ATG-F5 strain Sitosterol 1.7 Polyglyceryl 2-oleate 1.5 Ceteares-4 2.0 Cholesterol 3.0 Dicetyl phosphate 0.4 Concentrated glycerin 5.0 Sunflower Oil 22.0 Carboxyvinyl Polymer 0.5 Triethanolamine 0.5 Antiseptic Trace Perfume Trace Purified water Remainder
Cosmetic Formulation Example 5. Preparation of Foundation
[0148] Using the composition of Table 9 below, a foundation (100 g) containing a lyophilized product of the cell lysate of the Bifidobacterium longum ATG-F5 strain was prepared according to a conventional method.
TABLE-US-00009 TABLE 9 Content Raw material (g) Lyophilized product of cell lysate of 1.0 Bifidobacterium longum ATG-F5 strain Beeswax 2.0 Cyclomethicone 2.0 Liquid paraffin 5.0 Squalane 5.0 Stearic acid 2.0 Lipophilic monostearate glycerin 3.0 Caprylic/Capric Triglycerides 4.0 Glycerin 4.0 Propylene glycol 3.0 Butylene glycol 3.0 Triethanolamine 1.0 Aluminum Magnesium Silicate 0.5 Pigment 12.0 Antiseptic Trace Perfume Trace Purified water Remainder
Cosmetic Formulation Example 6. Preparation of Hair Shampoo
[0149] Using the composition of Table 10 below, a hair shampoo (100 g) containing a lyophilized product of the cell lysate of the Bifidobacterium longum ATG-F5 strain was prepared according to a conventional method.
TABLE-US-00010 TABLE 10 Content Raw material (g) Lyophilized product of cell lysate of 3.0 Bifidobacterium longum ATG-F5 strain Arachidyl Glucoside 4.5 Ethanol 2.0 Butylene glycol 2.0 Citric acid 0.1 Phenoxyethanol 0.02 Purified water Remainder
Formulation Example 1. Pharmaceutical Formulation
Formulation Example 1-1. Preparation of Tablets
[0150] 200 g of a lyophilized product of the cell lysate of the Bifidobacterium longum ATG-F5 strain of the present invention was mixed with 175.9 g of lactose, 180 g of potato starch, and 32 g of colloidal silicic acid. After adding a 10% (w/v) gelatin solution to this mixture, the resulting product was ground and passed through a 14-mesh sieve. The remaining solid was dried, and 160 g of potato starch, 50 g of talc, and 5 g of magnesium stearate were added thereto. The resulting mixture was prepared as an anti-inflammatory tablet.
Formulation Example 1-2. Preparation of Ointments
[0151] 1 g of a lyophilized product of the cell lysate of the Bifidobacterium longum ATG-F5 strain of the present invention was mixed with 99 g of petrolatum to prepare an anti-inflammatory ointment.
Formulation Example 2. Preparation of Food Product
Formulation Example 2-1. Preparation of Cooking Seasonings
[0152] A health-functional cooking seasoning was prepared by adding a lyophilized powder of the cell lysate of the Bifidobacterium longum strain of the present invention to a cooking seasoning in an amount of 1% by weight.
Formulation Example 2-2. Preparation of Dairy Products
[0153] A lyophilized powder of the cell lysate of the Bifidobacterium longum strain of the present invention was added to milk in an amount of 0.1% by weight, and various dairy products such as butter and ice cream were prepared using the milk.
Formulation Example 2-5. Preparation of Vegetable Juice
[0154] A health-functional vegetable juice was prepared by adding 0.5 g of a lyophilized powder of the cell lysate of the Bifidobacterium longum strain of the present invention to 1,000 ml of tomato juice or carrot juice.
Formulation Example 2-6. Preparation of Fruit Juice
[0155] Health-functional fruit juice was prepared by adding 0.1 g of a lyophilized powder of the cell lysate of the Bifidobacterium longum strain of the present invention to 1,000 ml of apple juice or grape juice.
[0156] [Depository Institution]
[0157] Name of Institution: The Korean Collection for Type Cultures
[0158] Accession Number: KCTC13828BP
[0159] Date of Deposit: Mar. 18, 2019
[Amendment by Rule 91 28 Jun. 2019]
[0160]