TW202235069A - A composition containing lactobacillus spp. and its use for preventing and/or improving anti-aging of skin - Google Patents

Abstract

The present invention provides a composition containing Lactobacillus spp. and its use for preventing and/or improving anti-aging of skin. Specifically, the present invention provides a composition containing Lactobacillus plantarum and Bifidobacterium lactis for preventing and/or improving anti-aging of skin. The present invention inhibit or alleviate Glycation by way of administering a subject the composition containing Lactobacillus spp..

Description

Composition containing lactobacilli and its use for preventing and/or improving skin aging

The present invention relates to a composition comprising Lactobacillus and its use for preventing and/or improving skin aging; more specifically, it relates to a composition comprising Lactobacillus plantarum , Bifidobacterium lactis or a combination thereof It is used to prevent and/or improve wrinkles, fine lines, spots or sagging on the skin caused by the glycation stress that increases with aging.

Mechanism of Glycation

Common saccharification reactions can be divided into three stages: (1) The first stage is the process in which carbohydrate molecules interact with substances with amine groups to produce Amadori products; (2) The second stage is the process of Amadori The product is degraded to produce highly active carbonyl compounds, such as glyoxal (GO), methylglyoxal (MG), etc.; (3) The third stage is the highly active carbonyl compound and protein or The process of amino acid interaction reaction to generate stable substances, in which the substance formed by reacting with protein is called glycated protein, and the small molecular substance formed by reaction with amino acid or cleavage of glycated residues (residues) of glycated protein is called glycated molecule. The products of the interaction between sugar molecules and active carbonyl substances produced in the process of sugar metabolism and molecules with amino groups such as proteins, are advanced glycation end products (AGEs). The formation pathway and mechanism of the saccharification reaction and its products are shown in Figure 1.

The physiological effects of AGEs are called glycation stress. The physiological effects of AGEs include: (1) It has the property of promoting protein cross-linking. AGEs can act as bridge molecules between different peptide chains in proteins to promote protein aggregation or tissue stiffness, causing proteins to lose their original functional properties. (2) Protein-bound AGEs can activate the receptor for AGEs (RAGE), oligosaccharyl transferase complex protein 48 (OST-48, generally known as AGER1), 80 K-H protein (AGER2), galectin-3 (AGER3) and other receptors may directly promote ROS production and affect cell physiology. Since RAGE activation is positively correlated with many diseases, AGEs interact with RAGE and play an important role in diseases.

Glycation and Skin Aging

Especially for the physiological effects of AGEs in (1) above, it is often studied in related diseases such as cardiovascular disease, aging, skin disease and neurodegeneration complicated by diabetes and chronic kidney disease. Many studies have confirmed that glycation stress is associated with various disease processes, including diabetes, obesity; neurological and cognitive diseases, cardiovascular diseases, kidney diseases, liver diseases, and some cancers (breast cancer, colorectal cancer, pancreatic cancer, etc.). Many studies have confirmed that glycation stress is also highly correlated with the aging of organisms. In the internal physiological response, including atherosclerosis, cataract or other related inflammation, etc. In terms of external manifestations, glycation stress will cause the human body's skin texture to become rough or loose, dark complexion with spots, dryness and peeling around the lips, etc. This is because AGEs deposit and attach to skin tissue, and bind to receptors called RAGE, causing inflammatory changes in the skin or other tissues. At the same time, in the process of generating AGEs, disordered cross-links will be formed within and between molecules of collagen, causing physical and physiological changes (modifications) to collagen. That is, it causes skin aging such as wrinkles, rough skin, dullness, or loss of firmness. In addition, due to the saccharification reaction, the collagen with disordered cross-links formed in the molecule and between the molecules will not only lose its elasticity, but also become difficult to be decomposed by the proteolytic enzymes in the living body, and the result may lead to Delay in skin transition speed. Therefore, in the field of medicine, based on the research on the relationship between glycation reaction and aging, it is attempted to prevent skin aging by inhibiting glycation reaction to develop another alternative treatment approach.

Existing anti-glycation raw materials

Raw materials currently on the market that are researched to have anti-glycation effects are divided into chemically synthesized structures or natural products derived from plants. Plant sources such as cinnamon ( Cinnamomum cassia ), cumin ( Cuminum cyminum ), rosemary (Rosemary), Houttuynia cordata ( Houttuynia cordata ), etc., extracted natural products such as curcumin (curcumin), rutin (Rutin) , quercitrin (quercitrin).

Consumers generally do not accept chemically synthesized anti-glycation products, and there are still health concerns. However, anti-glycation natural products derived from plants are often extracted with a large amount of solvents, so it is difficult to avoid the doubts of solvent residues. Or, the method and equipment threshold for making low-dose solvent residues is extremely high, resulting in high product prices. Therefore, neither chemically synthesized nor naturally extracted products can fully meet the needs of the market public.

Moreover, since glycation stress is related to skin aging, roughness, sagging, peeling and many other diseases, the development of anti-glycation raw materials is of great market value. And the market is also looking forward to using the anti-glycation functional composition derived from microorganisms as a novel material to expand the diversity of anti-glycation product choices on the market.

Lactobacillus

Lactobacillus spp. exists in the general environment and can ferment carbohydrates into lactic acid, so it is mostly used in the manufacture of fermented foods. As early as the 1900s, it was discovered that this bacterium has multiple benefits for human health, such as helping digestion and improving intestinal health, and it is advocated to eat more Lactobacillus to improve the digestive environment and promote intestinal peristalsis. After research, it was found that because Lactobacillus can decompose sugars (such as lactose, glucose, sucrose, fructose), it can produce lactic acid and acetic acid, and thereby acidify the intestinal environment, thereby inhibiting the proliferation of harmful bacteria in the intestinal tract , and the beneficial effect of adjusting the balance of intestinal flora.

Even if they belong to the same genus of Lactobacillus, if they are of different species, or even of the same species but different strains have different characteristics from each other, they will have different effects on the human body. For example, Lactobacillus plantarum ( Lactobacillus plantarum ), some literature pointed out that L. plantarum PH04 has the ability to lower blood cholesterol; another literature confirmed that L. plantarum 299V can reduce the symptoms of colitis in IL-10-deficient mice; The literature pointed out that L.plantarum 10hk2 can increase pro-inflammatory mediators, such as IL-1β, IL-6 and TNF-α, and anti-inflammatory interleukin-10 (IL-10), to achieve anti-inflammatory effects; there are even literatures confirmed L. plantarum K21 can lower blood cholesterol and triglyceride levels, and has anti-inflammatory effects. The various specific strains listed above belong to the same species of Lactobacillus plantarum, but they can be applied to different diseases or diseases because of the different active ingredients therein.

Lactobacillus plantarum

Lactobacillus plantarum is found in saliva and many fermented foods. Lactobacillus plantarum is highly adaptable, a Gram-positive anaerobic bacterium that can grow at temperatures ranging from 10 to 45 degrees Celsius and is adapted to pH levels of 3.2-4.2 or higher. Lactobacillus plantarum was isolated from saliva for the first time, and subsequent studies found that some specific strains of this bacterium can be used as probiotics for anti-oxidation, maintaining intestinal permeability, inhibiting the growth of gas-producing bacteria in the intestinal tract, and increasing hippocampal The body's brain-derived neurotrophic factor can be applied in the treatment of depression.

Bifidobacterium

Bifidobacterium ( Bifidobacterium ) widely exists in the digestive tract, vagina and oral cavity of humans and animals. It is Gram-positive, immobile, has rod-shaped cells, sometimes forked at one end, and is strictly anaerobic. This bacterium was isolated from the feces of healthy people as early as 1899, and subsequent research found that some specific strains of this bacterium can be used as probiotics in the fields of food, medicine and feed.

In order to improve skin problems that occur with age, such as wrinkles, fine lines, spots, sagging, etc., and to develop new materials to break through the limitations of existing materials, the present invention provides a composition for preventing and/or improving skin aging , comprising Lactobacillus plantarum , Bifidobacterium lactis or a combination thereof.

Preferably, the Lactobacillus plantarum strain GKM3 is deposited at the Food Industry Development Research Institute of the Foundation with the registration number BCRC-910787; the Bifidobacterium lactis strain GKK2 is deposited at the Food Industry Development Research Institute of the Foundation with the registration number BCRC910826.

Preferably, the composition comprises medium components for cultivating and/or fermenting the Lactobacillus

Preferably, the composition further includes plant extracts or compounds that have the effect of improving skin transformation and/or inhibiting glycation reaction.

Preferably, the plant extract having the effect of improving skin transformation and/or inhibiting saccharification reaction includes plant extracts of Cupressaceae, Ginkgoceae, Citrus, Perillaceae, Houttuyniaceae, and Eucommiaceae.

Preferably, the plants of the family Cupressaceae include Podocarpus; the plants of the Ginkgo family include Ginkgo biloba; the plants of the Citrus family include corkberry, orange, grapefruit, pomelo and lemon; the plants of the Perilla family include mint, peppermint, perilla and rosemary; the plant of Houttuyniaceae comprises Houttuynia cordata; the plant of Eucommiaceae comprises Eucommia.

Preferably, the compound having the effect of improving skin transformation and/or inhibiting glycation reaction includes equol.

Preferably, each unit of the composition contains more than 0.1% by weight of lactobacillus, and a total of more than 0.001% by weight of plant extracts or compounds that can improve skin transformation and/or inhibit glycation reaction.

Preferably, the aforementioned composition is a pharmaceutical composition, cosmetic, skin care product, nutritional product or food.

Preferably, the cosmetics include lotion, facial cleanser, beauty lotion, lotion or cream.

In order to make it easier to administer the composition containing lactobacilli to recipients in need, the present invention provides a use of the aforementioned composition containing lactobacilli for preventing and/or improving skin aging.

Preferably, the composition comprising Lactobacillus is prepared by the following method: (a) Inoculate a colony of Lactobacillus in the culture medium for solid-state culture to obtain bacteria; (b) inoculating the bacteria in step (a) into a liquid medium for liquid culture to obtain a mixture of liquid medium and bacteria; and (c) Fermenting the mixture of the liquid medium and the cells in step (b) to obtain a complete fermentation solution.

Preferably, the method further comprises the following steps: (d) centrifuging the whole fermentation liquid in step (c) to obtain the sludge and/or supernatant; and (e) Freeze-drying the sludge or the supernatant obtained in step (d) to obtain freeze-dried powder.

Preferably, the skin aging is improved by administering the composition comprising lactobacilli to inhibit or slow down the glycation reaction of the recipient.

Preferably, the inhibiting or slowing down of the subject's glycation response refers to the reduction of the content of pre-glycation products in the subject.

Preferably, the pre-saccharification product comprises Amadori.

Preferably, the inhibiting or slowing down of the glycation reaction of the recipient refers to the reduction of the content of dicarbonyl compounds in the mid-glycation stage of the recipient.

Preferably, the mid-saccharification dicarbonyl compound comprises glyoxal (GO), methylglyoxal (MG) or a combination thereof.

Preferably, the inhibiting or slowing down of the recipient's glycation reaction refers to the reduction of the content of the post-glycation product in the recipient, or the improvement of the content of the functional group contained in the product.

Preferably, the post-glycation products comprise advanced glycation end products (AGEs).

Preferably, the improvement of the content of the functional group includes the reduction of the content of carbonyl groups, the increase of the content of ε-amine groups, and the increase of the content of free sulfhydryl groups.

Preferably, the inhibiting or slowing down of the recipient's glycation response means that the subject's binding rate of advanced glycation end products (AGEs) to receptors for advanced glycation end products (RAGE) is reduced.

Preferably, the symptoms of skin aging include wrinkles, fine lines, spots, sagging or a combination thereof.

Preferably, the improving skin aging comprises increasing the recipient's skin elasticity, increasing skin moisture content or a combination thereof.

Strain culture

The above-mentioned Lactobacillus brevis colony (colony) was picked and inoculated on the solid medium to activate the strain. In a preferred embodiment, the solid medium is MRS agar. After the growth of the bacteria is completed, the fresh bacteria and the solid medium are inserted into the Erlenmeyer flask containing the liquid medium for liquid culture. In a preferred embodiment, the culture is carried out in a liquid state at a temperature of 25 to 40° C., an aeration rate of 0 to 1 vvm nitrogen or carbon dioxide, and a rate of 250 to 1000 rpm. In a preferred embodiment, the time for liquid culture is 16 to 24 hours, more preferably 18 hours for the number of viable bacteria to reach the highest point. In a preferred embodiment, the liquid medium is MRS liquid medium. In a preferred embodiment, the formulation of the liquid medium is shown in Table 1 below. After the liquid culture is completed, the fermentation culture is carried out. In one embodiment, the fermentation culture is an aerobic or anaerobic environment. In one embodiment, the culture temperature of the fermentation culture is controlled at 30°C to 37°C. In one embodiment, the time of fermentation culture is as long as 16 hours to 48 hours.

Table 1 Element Ratio (weight percent) carbohydrate 1~20% yeast extract 0.5~10% Protein Potion 0.5~10% Inorganic salts 0.01~5% amino acid 0.01~0.1%

Preparation of Lactobacillus Ferment

After the lactobacilli are fermented and grown to complete the growth, the whole fermentation liquid including the bacteria and the culture liquid is collected and centrifuged to separate the bacteria sludge and the supernatant. In a preferred embodiment, the whole fermentation solution is put into 15% by weight skim milk powder and left to stand for 1 to 4 hours to agglomerate the bacterial cells into agglomerates to increase the recovery rate. In a preferred embodiment, the centrifugal speed is 1000 to 15000 rpm. In the following experiments, the supernatant obtained after the fermentation broth was centrifuged was used as the lactobacillus fermentation product. The aspect of the lactobacillus of the present invention includes, but is not limited to, the above-mentioned fermentation whole liquid containing bacteria and culture liquid, the bacteria sludge after the fermentation whole liquid is centrifuged to remove the supernatant, and the fermentation whole liquid centrifugation to remove the bacteria sludge supernatant. The content of the lactic acid bacteria of the present invention can be usually 0.1% by weight or more, preferably 1 to 50% by weight, more preferably 5 to 30% by weight, relative to the total amount of the composition containing lactic acid bacteria of the present invention. , More preferably in the range of 10 to 15% by weight.

Plant extracts or compounds with skin transformation and improvement effects

In the composition of the present invention, the following plant extracts or compounds having skin transformation improvement effects are added as needed. By adding plant extracts or compounds that have the effect of improving skin transformation, the anti-wrinkle effect, anti-fine line effect, whitening effect, and even anti-spot effect and anti-relaxation effect achieved by Lactobacillus can be greatly improved Beyond just the additive effect of the two.

The aforementioned plant extracts or compounds that have the effect of improving skin transformation, such as sesame, rehmannia glutinosa, peach, licorice, cactus, wheat germ, wild strawberry, sage, mint, peppermint, houttuynia cordata, uncaria, perilla , Pipa, Japanese Camellia, Jujube, Polygonum indigo, Akebia, Ashitaba, Luo Hanbo, Acerola, Hollyhock, Aloe Vera, Ginkgo, Phellodendron, Forsythia, Olive, Orange, Chamomile, Quince, Grapefruit, Cinnamon, Moon peach, Burnet, White birch, Carambola, Swertia, Morus alba, Tea, Clove, Carrot, Witch hazel, Rose, Himalayan raspberry, False leaf tree, Gorse, Peony root bark , horse chestnut, evening primrose, cornflower, eucalyptus, saxifrage, grapefruit, coix seed, orchid, lemon, vetch, rosehip, rosemary, cherry, staghorn, chamaema chamaejasme, golden fruit olive, cnidium, Bordeaux Tree, castor, sassafras, angelica, daphne, frankincense, kansui, impatiens, forsythia, alfalfa, myrrh, tallow, stone pine, golden grass, xiongzi, fig, magnolia, white magnolia, Birch, quinoa, honeysuckle, gardenia, peach kernel, jujube and other plant extracts, seaweed, green algae, royal jelly, tangerine peel.

The aforementioned compounds that have the effect of improving skin transformation include carnitine, niacinamide, pyridoxine hydrochloride (vitamin B6), hyaluronic acid, glycolic acid, citric acid, lactic acid, malic acid, tartaric acid, 2 -Hydroxybutyric acid, 3-hydroxybutyric acid, tartronic acid, vitamin A, glycyrrhizic acid derivatives, allantoin, diclofenac, felbinac, ibuprofen, ketoprofen, rosso Profen and its salts.

Among the above-mentioned plant extracts that have the effect of improving skin transformation, there are plants of Cupressaceae (Tumba chinensis, etc.), plants of Ginkgoceae (Ginkgo biloba, etc.), plants of Citrus family (corkberry, orange, grapefruit, pomelo, lemon, etc.) ), plants of Perillaceae (mint, peppermint, perilla, rosemary, etc.), plants of Houttuyniaceae (Houttuynia cordata, etc.), and extracts of plants of Eucommiaceae (Eucommia, etc.) are more suitable. In addition, extracts of Luo Hanbo, Ginkgo biloba, Phellodendron barkberry, Orange extract, Perilla extract, Houttuynia cordata extract and Eucommia tea extract can neutralize lactic acid in the present invention due to their inhibitory effect on the saccharification reaction described later. Combinations of bacilli are most suitable.

With regard to the preferred contents of the above-mentioned plant extracts or compounds with skin transformation improvement effects, relative to the total amount of the composition containing lactobacilli of the present invention, it can usually be more than 0.001% by weight, preferably 0.01 to 5% by weight %, more preferably in the range of 0.1 to 20% by weight, more preferably in the range of 1 to 10% by weight.

Plant extracts or compounds with saccharification reaction inhibitory effect

In the composition of the present invention, the following plant extract or compound having a saccharification reaction inhibitory effect is added as needed. By adding plant extracts or compounds that have the effect of improving skin transformation, the anti-wrinkle effect, anti-fine line effect, whitening effect, and even anti-spot effect and anti-relaxation effect achieved by Lactobacillus can be greatly improved Beyond just the additive effect of the two.

The above-mentioned plant extracts that have the effect of inhibiting the saccharification reaction, such as artichoke, eyebright, cold water hemp, lobster, asparagus, vitex fruit, akebia, cabbage coconut fruit, nightshade, arhat Cypress, catechin, cork oak, gynostemma, fennel, witch hazel, aristotle, epimedium, knotweed, ginkgo, Japanese wintergreen, strawberry, shandan, seaweed, false loquat, yellow bellwood bark, Red leaf maple, Pansandan, fennel, American witch hazel, dwarf holly, turmeric, plum, oak angustifolia, epimedium shorthorn, camphor, coneflower, eleuthero, gorse, gentian, elderberry Wood, holly, fan palm, amurberry, berberine, milk thistle, oats, catkins, saffron, rose lily, yellow quail, ghost lily, oregano, orange, pumpkin seeds, catuaba bark, Deer lily, guarana, roselle, asarum, Jehol dandelion, platycodon, sorrel, Haizhou Changshan, Philippine island tree, Zhejiang lily, walnut, lingonberry, duck tongue, Saga brown algae, pomegranate, sweet potato, Saar Sand Root, Wild Hawthorn, Sanshope, Perilla, Wing Bean, Mosquito, Peony, Rhubarb palmatum, White Mountain Chrysanthemum, Quinoa, Mountain Pineland, Great Plantain, Centella Asiatica, Buckwheat, Musk Lily, Orina, Grapevine (Davilla rugosa), noble lily, Cordia salicifolia, clove, senna angustifolia, gotu kola, daisy, dill, chain hook, mountain chrysanthemum, iron cannon lily, chain hook root, hanging Hook, peach kernel, houttuynia cordata, prickly pear, Eucommia tea leaves, tomato, Potentilla, sorrel leaf, elderberry, nasturtium, chrysanthemum, mint, Hakata lily, sage, yam, passionflower , passion fruit, Brazilian ginseng root, jackfruit, scarlet cherry, sunflower, Hime boa grass, Hime lily, yellow quail, betel nut, coltsfoot, magpie bean, cohosh raceme, Showa grass, Polygonum chinensis, incense Nan, pine, saber leaf vertebra, yerba mate, white lily, pheasant pheasant fruit, province oil, wild papaya, mozuku, willow, daylilies, soursop, mountain lily, water spinach, weeding vegetables, lychee seeds , Minjiang Lily, Ryukyu Yazhu, Ryukyu Rose Strawberry, Unripe Apple Fruit, Basswood Flower, Osmanthus, Lettuce, Lemon, Lemongrass, Thyme, Lemon Leaf Verbena, Lemon Balm, Rosehip, Pale Red Cinnamon, Myth Rosemary, briar rose, laurel, rhodiola rosea, Mexican sumac, Burnet, milk vetch, persimmon leaf, licorice leaf, black soybean seed coat, purple rice seed, moon peach leaf, lily lily, western willow, Eucommia leaf, tomorrow Extracts of plants such as leaves of leaves, extracts of seaweed such as seaweed, seaweed, and antler seaweed, extracts of green coffee beans, sweet potato shochu, and agaric mycelium.

The above-mentioned compounds with saccharification reaction inhibitory effect, such as equol, isoflavones, 1,4-anthraquinone, 1-amino-2-hydroxymethylanthraquinone, 4-aminophenol, 1,3,5 -Trihydroxybenzene, kojic acid, 3,4-dihydroxyphenylacetic acid, caffeic acid, ifenprodil, 6-hydroxy-2,5,7,8-tetramethyldihydrobenzopyran -2-Carboxylic acid, 6-oxindole, 7-hydroxy-4,6-dimethylphthalide, α-lipoic acid, 4-hydroxychalcone, pearl protein hydrolyzate, aminoguanidine, red algae Octyl sodium, ergothiol, resveratrol, hydroxystilbenes such as 3,3',5,5'-tetrahydroxystilbene, oxindole, carnosine, salicylic acid, norporcine Ericaine bibromide, sinapic acid, nicotinic acid tocopheryl ester, nicotinic acid amide, nordihydroguaiaretic acid, anthocyanins, mannitol, hydrolyzed casein, hydrolyzed tannin, Catechol, chlorogenic acid, isochlorogenic acid, neochlorogenic acid, cryptochlorogenic acid, caffeoylquinic acid and other chlorogenic acids, cyanidin, betulin, procyanine glycol oligomers, Glucosylrutin, etc.

Among the aforementioned ingredients, withania, luohanbo, catechin, ginkgo, dwarf holly, amurberry, orange, pumpkin seeds, catuaba bark, phenanthrene, glucosyl rutin, lingonberry, perilla, buckwheat leaves, grape Rattan (Davilla rugosa), Houttuynia cordata, Hime boa constrictor grass, Showa grass, Polygonum japonica, Ryukyu arrow bamboo, Ryukyu rose strawberry, unripe apple fruit, persimmon leaf, licorice leaf, moon peach leaf, Eucommia leaf extract, and female Equor, isoflavones, ifenprodil, pearl protein hydrolyzate, catechol, caffeic acid, and betulin, because they can effectively improve the anti-wrinkle effect and anti-aging effect caused by the composition containing lactobacillus. Fine-grain effect, whitening effect, even anti-spot effect and anti-relaxation effect can be used as a better combination in the present invention.

Among the above-mentioned plant extracts with saccharification reaction inhibitory effect, there are also plants of Cupressaceae (Podocarpus, etc.), plants of Ginkgoceae (Ginkgo, etc.), plants of Citrus (corkberry, orange, lemon, etc.), plants of Perillaceae Extracts of plants (perilla, rosemary, etc.), plants of the Houttuyniaceae family (Houttuynia cordata, etc.), and plants of the Eucommiaceae family (Eucommia, etc.) are more suitable. Among them, Luo Hanbo extract, Ginkgo biloba extract, Phellodendron bark extract, Orange extract, Perilla extract, Houttuynia cordata extract, and Eucommia leaf extract have skin conversion and improvement effects. In the present invention, they are based on Combination with Lactobacillus works best.

When these plant extracts or compounds with saccharification reaction inhibitory effect are used together with Lactobacillus, it can effectively inhibit the formation of saccharification end products (advanced glycation end products: AGEs), in addition to reducing or even poorly inhibiting AGEs on In addition to the precipitation and adhesion of skin tissue, at the same time, in the process of generating AGEs, it can form disordered cross-links in the molecules and between molecules of collagen, and can reduce or even inhibit the physical and physiological effects caused by collagen. Change (modification), so it can effectively prevent or even improve skin aging such as wrinkles, skin roughness, dullness, or reduction of firmness. Furthermore, when the plant extracts having the saccharification reaction inhibitory effect are used together with lactobacilli, the following advantages can be exerted: the odor and odor caused by the plant extracts are blocked, and the storage stability is further improved.

Regarding the preferred contents of the above-mentioned plant extracts or compounds having a saccharification reaction inhibitory effect, relative to the total amount of the composition containing lactobacilli of the present invention, it can usually be more than 0.001% by weight, preferably 0.01 to 5% by weight. %, more preferably in the range of 0.1 to 20% by weight, more preferably in the range of 1 to 10% by weight.

Concrete products of the composition and their additives

The composition of the present invention can be applied to the field of cosmetics such as cosmetics and make-up cosmetics through external use, especially when added to cosmetics such as lotion, facial cleanser, beauty lotion, emulsion, and cream that are directly applied to the skin, it has excellent anti-aging properties. aging effect.

lotion

The composition of the present invention can use an aqueous medium as a base, and further blend ethanol and a moisturizing agent into it to form a lotion. The lotion system can be used to supply moisture or moisturizing ingredients to the stratum corneum of the skin, and can also be used to assist the physiological functions of the skin. Lotions include cleansing lotion, astringent lotion, softening lotion, multi-layered lotion, etc., and also include skin lotion, body lotion, hand lotion, moisturizing lotion, and the like.

For the lotion prepared by blending the composition of the present invention, it is better to use a moisturizer that has an excellent moisturizing effect and can more effectively exert the effect of improving rough skin by combining Lactobacillus. Humectants such as glycerin, ethylene glycol, propylene glycol, 1,3-butanediol, 3-methyl-1,3-butanediol, pentaerythritol, diglycerin, polyglycerin, diethylene glycol, triethylene glycol Alcohol, polyethylene glycol, dipropylene glycol, trehalose, ceramides, 2-methacryloxyethylphosphocholine, hyaluronic acid, chondroitin sulfate, hydrolyzed hyaluronic acid, yeast extract, coix seed essence, Peony essence, seaweed essence, gentian essence, trimethylglycine, N-methyl-L-serine, niacinamide, royal jelly essence. The amount of the moisturizer is generally 0.0001% by weight or more, preferably 0.001 to 50% by weight, more preferably 0.01 to 25% by weight, and even more preferably 0.1 to 10% by weight, based on the total amount of the lotion of the present invention blended within the range.

For the lotion prepared by blending the composition of the present invention, it is better to use an astringent with excellent astringent effect. Astringents include acid agents such as citric acid, succinic acid, tartaric acid, retinoic acid, malic acid, etc., retinol, zinc chloride, aluminum powder, aluminum compounds such as aluminum sulfate, alum and modified alcohols, ethanol, European Hollyhock Essence, Rabbit Chrysanthemum Essence, Polygonum Essence, Biting Cat Essence, Orris Root Essence, Fennel Essence, Oolong Tea Essence, Ethanol, Crape Myrtle Essence, Forsythia Essence, Orange Blossom Water . Essence, white birch essence, honeysuckle essence, quince essence, ivy essence, western wild hawthorn essence, thyme thyme essence, tea essence, houttuynia cordata essence, parsley essence, North American gold Hazel essence, grape leaf essence, pine essence, lemon balm essence, cornflower essence, mugwort essence, lemon juice, vetch essence, burnet elm essence. The astringent is usually 0.0001% by weight or more, preferably 0.001-50% by weight, more preferably 0.01-25% by weight, and even more preferably 0.1-10% by weight, relative to the total weight of the lotion of the present invention. Blended under range.

For the lotion prepared by blending the composition of the present invention, it is preferable to use a cuticle softening agent with excellent cuticle softening effect. Cuticle softening agents include, for example, urea, sulfur, ethanol, lactic acid, glycolic acid, salicylic acid, sodium sulfate, and the like.

In terms of softening (softening) agents, oily components similar to sebum are preferred. For the lotion prepared by blending the composition of the present invention, it is better to use a softener (softening) agent that has excellent softening effect and can be widely used in the field of cosmetics. Softening (softening) agents such as cetyl alcohol, lauryl alcohol, cetearyl alcohol, stearyl alcohol, oleyl alcohol, behenyl alcohol, lanolin alcohol, hydrogenated lanolin alcohol, hexyldecanol and octyldodecyl alcohol Alcohol, squalane, vegetable squalane, cetyl 2-ethylhexanoate, α-olefin oligomer, isotridecyl isononanoate, isononyl isononanoate, cetyl isononanoate , Ethyl Oleate, Oleyl Oleate, Ethylhexyl Palmitate, Capryl Glycol, Hydrogenated Soy Phospholipids.

For the lotion prepared from the composition of the present invention, it is preferable to use a surfactant that has a high effect of inhibiting the turbidity and precipitation of cosmetics over time and is widely used in the field of cosmetics. Surfactants such as L-arginine ethyl. DL-pyrrolidone carboxylate (L-arginine ethyl⋅DL-pyrrolidone carboxylic acid salt), polyoxyethylene glyceryl pyrroglutamic acid isostearate, PEG-9 polydimethylsiloxy ethyl dimethyl Silicone, coconut oil fatty acid sorbitan, polyoxyethylene polyoxypropylene decyl tetradecyl ether, polyoxyethylene cetyl ether, polyoxyethylene stearyl ether, polyoxyethylene methyl glucoside, Polyoxyethylene lauryl ether, polyoxyethylene hardened castor oil, polyoxyethylene hydrogenated castor oil, sorbitan isostearate, polyoxyethylene isostearate hardened castor oil, fatty acid esters of glycerol , polyglyceryl diisostearate, sucrose fatty acid ester, sodium stearyl glutamate, sodium stearyl lactate, sorbitan sesquiisostearate, sorbitan sesquioleate, polyoxygen Vinyl alkyl (C12-15) ether phosphoric acid, polyoxyethylene cholesteryl ether, polyoxyethylene cetyl ether phosphoric acid, polyoxyethylene cetearyl ether, polyoxyethylene phytosterol, polyoxyethylene mountain Alcohol ether, polyoxyethylene sorbitan monooleate (20E.O), glycerin monostearate, polyethylene glycol monostearate, polyoxyethylene glycerol monostearate, polyoxyethylene monolaurate Sorbitan, dioctyldodecyl laurylglutamate, hydrolyzed collagen solution, glyceryl monostearate, hydrogenated soybean lysophospholipids, hydrogenated soybean phospholipids, and water-soluble collagen.

Furthermore, in order to improve the quality and stability of the lotion, fragrances, pigments, preservatives, bactericides, ultraviolet absorbers, metal blocking agents and buffers can be blended. These substances are usually 0.0001% by weight or more, preferably 0.001 to 50% by weight, more preferably 0.01 to 25% by weight, and more preferably 0.1 to 10% by weight, relative to the total amount of the lotion of the present invention. Blend down.

facial cleanser

The composition for external use on skin of the present invention can be used as a facial cleanser by blending ingredients having a cleansing effect. As far as the dirt on the skin is concerned, it is the dirt (dirty) produced by the original physiological function of the skin, that is, in addition to the rancidity and deterioration of the sebum membrane and those whose cuticle has been peeled off, there are also dust in the air, etc. Dirt and daily use of cosmetics, facial cleanser can be used almost every day in order to wash away these dirt and keep the skin clean. Specific examples of face cleansers include soaps, body washes, face cleansers, and make-up removers. In terms of ingredients having a cleansing effect, alkali salts (soaps) of higher fatty acids can be mentioned, and the alkali salts of higher fatty acids are mainly sodium salts, potassium salts, and amine salts of C10~C18 fatty acids.

In the facial cleanser, in order to improve and stabilize the quality, one or more selected from fragrances, pigments, antioxidants and metal blocking agents can be further blended. These substances are relative to the total weight of the facial cleanser of the present invention. , usually at least 0.0001% by weight, preferably 0.001 to 50% by weight, more preferably 0.01 to 25% by weight, and more preferably 0.1 to 10% by weight. In the face cleanser, one or two or more selected from glycerin, ethanol, sugar, polyhydric alcohols, and germicides may be further blended. Bactericides In soap-based detergents such as facial cleansers to achieve the purpose of disinfection, sterilization, and deodorization, the use of clothexidine hydrochloride, isopropyl methylphenol, photosensitive element 101, cresol, and chlorine dichloride Cresol, trichlorocarbonylaminobenzene, halocarban, phenol, chlorocresol, phenoxyethanol, trichlorohydroxydiphenyl ether, etc. are preferred.

Beauty Essence

The composition for external use on skin of the present invention can be used as a beauty essence by blending a moisturizing agent and a softening agent. Generally speaking, the so-called beauty essence is different from lotion, and it is a type of cosmetic that has viscosity, excellent usability and moisturizing properties, and is considered to have moisturizing and softening functions. Known dosage forms include dissolvable type, O/W emulsified type, and oily liquid type.

In general, most of the beauty essences have a moisturizing function and a softening function by further blending one or more kinds selected from fragrances, ultraviolet absorbers, whitening agents, anti-inflammatory agents, preservatives, and bactericides. the functional person. The anti-inflammatory agent is preferably used alone or in combination with glycyrrhizic acid or its derivatives such as stearyl glycyrrhizinate and dipotassium glycyrrhizinate, gentian essence, perilla essence, sage essence, Western Elderberry essence, gotu kola essence, Houttuynia cordata essence, angelica essence, peony essence, evening primrose essence, Sapinberry essence, saxifrage essence, mugwort essence, apple essence or one 2 or more are preferred. These substances are usually in the range of 0.0001% by weight or more, preferably 0.001-50% by weight, more preferably 0.01-25% by weight, and more preferably 0.1-10% by weight, relative to the total amount of the beauty essence of the present invention Blend down.

lotion

The composition for external use on the skin of the present invention uses an aqueous medium as a base, and can be formed into an emulsion by blending an oily component and a surfactant. The purpose of the lotion is to clean the skin, remove makeup, protect the skin, moisturize and soften it, and promote the blood circulation of the skin. Except for special circumstances, emulsion is a fluid emulsion because the amount of oil phase is less than that of cream, so it is also called "liquid cream". There are two types of emulsification: O/W type and W/O type, but generally speaking, it is O/W type. Because the ratio of water phase is large, when applied to the skin, it can be stretched lightly and give a refreshing feeling. For the oily component and surfactant, various components described in detail in the section of the aforementioned lotion can be used appropriately.

In the emulsion of the present invention, one or two or more selected from moisturizing agents, softeners, blood flow promoters and polymer substances can be blended in accordance with the purpose and function. These substances are compared with the emulsion of the present invention The total amount is usually at least 0.0001% by weight, preferably 0.001 to 50% by weight, more preferably 0.01 to 25% by weight, and more preferably 0.1 to 10% by weight. Generally speaking, emulsions are mostly pre-added polymer substances in the water phase for the purpose of protecting colloids. Examples of polymer substances include carboxyvinyl polymers, sanxian gum, acrylic acid polymers, sodium carboxymethyl cellulose (CMC) )Wait.

cream

The composition for external use on skin of the present invention uses an aqueous medium as a base, and can be formed into a cream by blending oily components and surfactants. The type of emulsification is the same as that of the emulsion, there are both O/W type and W/O type, but the cream is generally a non-fluid emulsion because the oil phase is more than the emulsion. The purpose of the cream is the same as that of the emulsion, which is to clean the skin, remove makeup, protect the skin, moisturize and soften it, and promote the blood circulation of the skin. More specifically, cleansing cream, softening cream, nourishing cream, night cream, primer, dissipating cream, moisturizing cream, massage cream, cold cream, lip care cream, etc. are mentioned.

The oil-based ingredients and surfactants used in the cream can be applied to the same ingredients as the aforementioned lotion. The cream system can further blend one or more kinds selected from cetyl wax, cetyl alcohol, lanolin, liquid paraffin, vaseline, glycerin and squalane as supplementary ingredients of oily ingredients. The total amount of cream is usually 0.0001% by weight or more, preferably 0.001 to 50% by weight, more preferably 0.01 to 25% by weight, and more preferably 0.1 to 10% by weight.

other ingredients

The composition of the present invention can be applied to lotion, facial cleanser, beauty lotion, lotion or cream, and other commonly used ingredients such as thickeners, vitamins, Amino acids, anti-wrinkle agents, seaweed extracts, cell activators, transdermal absorption accelerators, foaming agents, solubilizers, keratolytic agents, hormones, pigments, plasticizers, inorganic powders, organic powders, etc. .

The composition of the present invention can also be used in the fields of medicine or diet through internal use, especially when added to medicines, feeds, beverages, nutritional supplements, dairy products, food or health foods that are directly suitable for oral administration, it has excellent anti-aging effect.

The composition of the present invention may further contain additives according to the desired product form according to the application. The additive may be an excipient, preservative, diluent, filler, absorption enhancer, or sweetener. The excipient may be selected from sodium citrate, calcium carbonate, calcium phosphate, sucrose or combinations thereof. The preservatives prolong the shelf life of the composition, eg benzyl alcohol, parabens. The diluent may be selected from water, ethanol, propylene glycol, glycerin or combinations thereof. Fillers may be selected from lactose, high molecular weight polyethylene glycols or combinations thereof. The absorption enhancer may be selected from dimethylsulfoxide (DMSO), laurocapram, propylene glycol, glycerin, polyethylene glycol, or combinations thereof. The sweetener may be selected from Acesulfame K, aspartame, saccharin, sucralose, neotame or combinations thereof. In addition to the above-listed additives, other suitable additives can be selected as required, provided that the medicinal effect of the composition is not affected.

On the premise that the drug form does not affect the desired effect of the composition, it can be prepared as powder, lozenge, suppository, microcapsule, ampule (ampoule/ampule), liquid spray or Suppository.

The route of administration is based on the form of the composition and the appropriate dose for the recipient, for example, oral or parenteral, and also includes intravenous, intramuscular, intraperitoneal, subcutaneous, transdermal, respiratory (aerosol) ), rectal, vaginal or topical (including oral and sublingual). The timing of administration depends on the form of the composition and the appropriate dose for the recipient, for example, every meal, every day, every week. The recipient is a human or an animal.

Experimental design of saccharification reaction

Glucose (glucose) was used as the glycation molecule, methylglyoxal (MG), and bovine serum albumin (BSA), the albumin with the highest content in the mammalian circulatory system, were used in the experiment of saccharification reaction. The reactions were carried out in the following different experiments. The concentrations of glucose, methylglyoxal, and bovine serum albumin were 10 mM, 25 mM, and 5 mg/mL, respectively. The strains selected as Lactobacillus can be purchased from BCRC as shown in Table 2, and the Lactobacillus fermented product was prepared by the aforementioned method. The following in vitro experimental results are calculated in triplicate.

Table 2 serial number strain name scientific name Registration number 1 Lactobacillus bulgaricus Lactobacillus bulgaricus ATCC SD 6833 2 Short Tits GKJOY Lactobacillus brevis BCRC-910920 3 Bifidobacterium bifidum Bifidobacterium bifidum BCRC-910986 4 Pediococcus pentosacea Pediococcus pentosaceus BCRC-910879 5 Lactobacillus plantarum Lactobacillus plantarum BCRC-910787 6 Bifidobacterium lactis Bifidobacterium lactis BCRC-910826 7 Lactobacillus plantarum Lactobacillus plantarum BCRC-910877 8 Lactococcus lactis Lactococcus lactis BCRC-12312 9 Lactobacillus rhamnosus Lactobacillus rhamnosus BCRC-910918 10 Pediococcus lactis Pediococcus acidilactici BCRC-910876 11 Bifidobacterium breve Bifidobacterium breve BCRC-910989 12 Lactobacillus casei Lactobacillus casei BCRC-910825 13 Lactobacillus reuteri Lactobacillus reuteri BCRC-910827 14 Lactobacillus johnsonii Lactobacillus johnsonii BCRC-910999 15 Lactobacillus paracasei Lactobacillus paracasei BCRC-910788 16 Lactobacillus plantarum Lactobacillus plantarum BCRC-910858 17 Bifidobacterium animalis Bifidobacterium animalis BCRC-910917 18 Lactobacillus fermentum Lactobacillus fermentum BCRC-910824 19 Lactobacillus salivarius Lactobacillus salivarius FERM BP-7974 20 Lactobacillus plantarum Lactobacillus plantarum BCRC-910919 twenty one Lactobacillus rhamnosus Lactobacillus rhamnosus BCRC-910828 twenty two Corynebacterium glutamicum Corynebacterium glutamicum ATCC-14619 twenty three Lactobacillus rhamnosus Lactobacillus rhamnosus BCRC-16000 twenty four Lactobacillus brevis Lactobacillus brevis BCRC-910859 25 Bifidobacterium longum Bifidobacterium longum BCRC-910988

Experiment 1: Determination of early glycation products

Glucose was selected as the glycosylated molecule and bovine serum albumin (BSA) to prepare the saccharification reaction solution. The saccharification reaction was carried out for 5 days in total, and the reaction temperature was controlled at 37°C. In this experiment, the control group was BSA containing glycosylated molecules, and the experimental group was BSA containing glycosylated molecules added to the fermentation product of Lactobacillus to compare glycosylated BSA (Glycated BSA) or BSA containing glycosylated molecules intervened in the fermentation product of Lactobacillus. The first day of saccharification reaction was taken, and the content of Amadori products in the early stage of saccharification was determined by using nitroblue tetrazolium (NBT) reagent with reference to the method of Armbruster et al. in 1987.

Figure 2 shows the effect of Lactobacillus fermentation on the formation of Amadori products in the early stage of protein saccharification. The control group was glycosylated bovine serum albumin (Glycated BSA) as a glycosylated protein. The concentration of ketoamine (ketoamine) as a product of Amadori was measured to be 10.72 nmoles/mg without adding any lactobacillus fermentation product , marking the vertical line to the right of the figure. Compared with the control group, the other groups supplemented with Lactobacillus fermentation products had lower concentrations of ketamines produced in the early stage of glycation, most of which were between 2 and 9 nmoles/mg. The fermentation products of Lactobacillus GKM3 and GKK2 numbered 5 and 6 respectively reduced the concentration of ketamine to 2.82 nmoles/mg and 3.47 nmoles/mg. This result shows that the Lactobacillus of the present invention or its fermented product are all helpful to the formation of amadori products in the early glycation stage in the anti-glycation reaction.

Experiment 2: Evaluation of the capture efficiency of dicarbonyl compounds in the mid-saccharification stage

1 mL of the lactobacillus fermentation product was mixed with 1 mL of methylglyoxal (MG) at a concentration of 2 mM, and reacted at 37° C. for 1 hour. Then add 1mL o-phenylenediamine (OPD) with a concentration of 12Mm, and react at 37°C for 15 minutes to derivatize MG into 2-methylquinoxaline (2-MQ), followed by Centrifuge at 14,000 xg for five minutes. In this experiment, the control group contained only MG, and the experimental group was MG added with Lactobacillus ferment to compare MG or MG added with Lactobacillus ferment. Referring to the method of Lo et al. in 2011, HPLC analysis was carried out to determine the ability of Lactobacillus fermentation to capture the dicarbonyl compound MG in the mid-saccharification stage.

Figure 3 is an evaluation of the ability of Lactobacillus fermentation to capture dicarbonyl compounds in the middle stage of protein saccharification. Comparing the MG without adding any lactobacillus fermentation product with the group adding lactobacillus fermentation product, taking the MG display value of 0 without adding any lactobacillus fermentation product as a benchmark, the value of the MG capture ability of the adding lactobacillus fermentation product group is higher The higher the value, the stronger the ability to capture MG. The values of each group of Lactobacillus fermentation products were significantly improved, and the values of the fermentation products of Lactobacillus GKM3 and GKK2 respectively numbered 5 and 6 were higher, showing better MG capture ability. This result shows that the Lactobacillus or its fermented product of the present invention is helpful for the capture of dicarbonyl compounds in the mid-glycation stage in the anti-glycation reaction.

experiment three : Evaluation of the degree of saccharification and its functional groups in the late stage of saccharification

In this experiment, the control group was BSA without glycosylated molecules and BSA with glycosylated molecules, and the experimental group was BSA with glycosylated molecules added with Lactobacillus fermentation products to compare glycosylated BSA (Glycated BSA). Take the saccharification reaction solution as configured in Experiment 1, and use the sodium dodecyl sulfate polyacrylamide gel electrophoresis (sodium dodecyl sulfate Polyacrylamide gel electrophoresis, SDS-PAGE) was used to analyze the effect of various Lactobacillus fermentation groups on the degree of protein glycosylation. And with reference to the method of Levine et al. in 1990, use 2,4-dinitrophenylhydrazine (2,4-dinitrophenylhydrazine, DNPH) to measure the specific functional group carbonyl content (carbonyl content) in glycosylated protein; refer to 2013 Hashimoto et al. Human's method, using 2,4,6-trinitrobenzenesulphonic acid (2,4,6-trinitrobenzenesulphonic acid, TNBS) to determine the reduction of specific functional groups ε-amino groups (free ε-amino groups) in glycated proteins content; and refer to Ellman's method in 1959, with Ellman's reagent (ie 5,5'-dithiobis-(2-nitrobenzoic acid), 5,5'-dithiobis-(2-nitrobenzoic acid), DTNB) Determination of changes in the content of free sulfhydryl groups.

Fig. 4 is the SDS-PAGE profile of the late product of protein saccharification by the fermentation product of Lactobacillus. The left column of the figure shows that the glycosylated protein (Glycated BSA) has a significantly larger molecular weight, more diffuse band distribution, and a lower concentration than the unglycosylated protein (BSA). This is because the functional groups of the protein will be cross-linked during the saccharification process of the protein. The upper right column in the figure shows the group in which glycosylated protein is added with lactobacillus fermented product, and the lower column on the right shows the group without saccharified protein added with lactobacillus fermented product. The lower column on the right shows that the protein before unsaccharification is added with Lactobacillus fermentation product and the left column is compared with the protein before unsaccharification. Both band distributions have small diffusion, high concentration and no obvious difference. The results indicated that the proteins in these two groups were not glycosylated, so no matter whether the Lactobacillus fermentation product was added or not, the protein would not be cross-linked. The upper right column of the figure shows the group of glycosylated proteins added with Lactobacillus fermented products, compared with the group of glycosylated proteins added with Lactobacillus fermented products in the SDS-PAGE pattern. Higher, showing that the cross-linking of glycosylated proteins is reduced. Especially for the groups of Lactobacillus GKM3 and GKK2 numbered 5 and 6 respectively, the distribution of the bands has a small diffusion and a high degree of concentration, and is obviously concentrated on the lower side, and the degree of molecular cross-linking or aggregation is more judged. small. This result shows that the strains of the present invention all help to inhibit the cross-linking polymerization reaction derived from protein glycation in the late stage of glycation reaction.

Figure 5 shows the effect of Lactobacillus fermentation product on the carbonyl content of the late product of protein saccharification. In the control group, after co-reaction of BSA (5 mg/mL) with glucose (10 mM) and MG (25 mM), the carbonyl content of Glycated BSA increased to 69.74 nmoles/mg, indicated by the vertical line on the right side of the graph. On the other hand, in the group added with Lactobacillus fermentation product, the increase in carbonyl content was lower than that of the control group, and ranged from 25 to 65 nmoles/mg. Among them, the fermentation products of Lactobacillus GKM3 and GKK2 numbered 5 and 6 increased less carbonyl groups, even reaching below 30nmoles/mg. This result shows that the lactobacillus or its fermented product of the present invention is helpful to resist the reduction of the carbonyl content of the late product of the saccharification reaction.

Figure 6 shows the effect of lactobacillus fermentation product on the content of ε-amine group in the late product of protein saccharification. In the control group, after the co-reaction of BSA (5 mg/mL) with glucose (10 mM) and MG (25 mM), the ε-amine content of Glycated BSA decreased by 37.28%, indicated by the vertical line on the middle side of the figure Wire. In contrast, the reduction of ε-amine group was improved in some groups added with Lactobacillus fermentation product. Among them, the fermentation products of Lactobacillus GKM3 and GKK2 numbered 5 and 6, compared with other strains, reduced the reduction of the ε-amine group the most. This result shows that the Lactobacillus or its fermented product of the present invention is helpful to improve the reduction of ε-amine content in the late stage of the anti-glycation reaction.

Fig. 7 shows the effect of lactobacillus fermentation product on the free sulfhydryl content of late product of protein saccharification. In the control group, after the co-reaction of BSA (5 mg/mL) with glucose (10 mM) and MG (25 mM), the free sulfhydryl content of Glycated BSA was 0.97 nmoles/mg, indicated by the middle side in the figure Vertical line. In contrast, in some groups added with Lactobacillus fermentation product, the free sulfhydryl content was higher than 0.97 nmoles/mg, indicating that the Lactobacillus fermentation product reduced the free sulfhydryl group of oxidized Glycated BSA. Among them, the fermentation products of Lactobacillus GKM3 and GKK2 numbered 5 and 6 have a stronger ability to reduce sulfhydryl groups of glycated proteins than other strains. This result shows that the lactobacillus or its fermented product of the present invention has a strong ability to reduce sulfhydryl groups in the late stage of the anti-glycation reaction.

Experiment 4: Highly glycosylated end products in the late stage of saccharification (AGEs) Determination of content

Taking the saccharification reaction solution as configured in Experiment 1, the reaction solution on the fifth day after the end of the reaction was used to measure the content of highly glycosylated end products (AGEs) derived from MG by referring to the method of Sero et al. in 2013.

Figure 8 shows the effect of lactobacillus fermentation on the formation of advanced glycation end products (AGEs) in the late stage of protein glycation. In a specific embodiment, one of the typical AGEs products derived from MG is argpyrimidine. In the control group, after the co-reaction of BSA (5 mg/mL) with glucose (10 mM) and MG (25 mM), the content of argpyrimidine increased 84.32 times compared with that before unsaccharification, indicated by the vertical line on the right side of the figure (near the frame line on the right side of Figure 8). On the other hand, when the lactobacillus fermented product was added, most of the argpyrimidine content increase ratios were greatly reduced. This result shows that the Lactobacillus or its fermented product of the present invention helps to reduce the generation of advanced glycated end products (AGEs) in the later stage of the saccharification reaction.

Experiment 5: Evaluation of the binding ability of glycated protein to receptors of highly glycated end products in the late stage of glycation

Take the saccharification reaction solution as configured in Experiment 1, the reaction solution on the fifth day after the end of the reaction, refer to the method of Du Yan et al. in 1997, use CircuLex AGE-RAGE in vitro Binding Assay Kit (No. CY-8151 , MBL Medical & Biological Laboratories Co. Ltd.) To analyze the effect of Lactobacillus fermentation on the combination of MG-BSA and RAGE.

Figure 9 shows the effect of lactobacillus fermentation on the combination of advanced glycation end products (AGEs) and receptors for advanced glycation end products (RAGE) in the saccharification reaction. Because AGEs combine with RAGE on the cell membrane to initiate specific cellular biological responses, related responses play an important role in many diseases and aging. Compared with the AGE-RAGE binding rate of the control group without adding lactobacillus fermentation product as a benchmark, the AGE-RAGE binding rate during the saccharification process decreased in the experimental group added with lactobacillus fermentation product. This result shows that the Lactobacillus or its fermented product of the present invention helps to reduce the binding rate of advanced glycation end products (AGEs) and receptors for advanced glycation end products (RAGE) in the saccharification reaction.

Experiment 6: Detection experiment of human skin elasticity

Experimental design The subjects first analyzed the skin elasticity on the flexion side of the forearm with a moisture elasticity tester (ANENG brand, Shengwang Electronics Co., Ltd.). The evaluation scale of the skin elasticity score is divided into 1 to 5 points, and the skin elasticity status represented by each score is shown in the following table:

table 3 Fraction skin condition 1 rough 2 Rough 3 general 4 soft 5 soft

After obtaining the skin elasticity value before the experiment, apply 100 μL of Lactobacillus fermented liquid on the skin of the flexion side of the forearm every day and record the daily skin elasticity. The experiment process lasted for ten days.

Figure 10 is the evaluation result of the improvement of human skin elasticity by the fermentation product of lactobacillus. According to the actual measurement, the elasticity of the lactobacillus fermented liquid on human skin increases with the increase of use time. This result shows that the lactobacillus or its fermented product of the present invention helps to increase skin elasticity and improve the aging phenomenon of human skin.

Experiment 7: Human skin moisture detection experiment

Experimental Design The subjects first analyzed the skin moisture on the flexion side of the forearm with a moisture elasticity tester (ANENG brand, Shengwang Electronics Co., Ltd.). The normal range of arm moisture will vary with the seasons. Summer is generally 35~60%, while winter is generally 30~45%. The detection season of this experiment is winter. After obtaining the skin moisture value before the experiment, apply 100 μL of Lactobacillus fermented liquid on the skin of the flexion side of the forearm and record the skin moisture again after 10 minutes.

Figure 11 is the result of the water-retaining properties of the lactobacillus fermented product on human skin. According to the actual measurement, the lactobacillus fermented liquid has the effect of improving the water-locking property of human skin. It shows that the lactobacillus or its fermented product of the present invention helps to maintain better skin elasticity, and can improve the water loss phenomenon, which is one of the aging phenomena of human skin.

none

Figure 1 shows the formation pathway and mechanism of the saccharification reaction and its products.

Figure 2 shows the effect of Lactobacillus on pre-glycation products.

Figure 3 shows the evaluation of the ability of Lactobacillus to capture dicarbonyl compounds in the mid-saccharification stage.

Figure 4 shows the electropherogram showing the effect of Lactobacillus on the cross-linking reaction of late glycation products.

Figure 5 shows the effect of Lactobacillus on the carbonyl content of late glycation products.

Figure 6 shows the effect of Lactobacillus on the content of ε-amine groups in late glycation products.

Figure 7 shows the effect of Lactobacillus on the free sulfhydryl content of late saccharification products.

Figure 8 shows the effect of Lactobacillus on advanced glycation end products (AGEs) in advanced glycation.

Figure 9 shows the effect of Lactobacillus on the binding of advanced glycation end products (AGEs) and receptors for advanced glycation end products (RAGE) in the glycation reaction.

Figure 10 shows the evaluation of lactobacillus on the improvement of human skin elasticity.

Figure 11 shows the results of the water retention of Lactobacillus on human skin.

Lactobacillus plantarum GKM3: BCRC-910787

Bifidobacterium lactis GKK2: BCRC-910826

Claims (24)

A composition for preventing and/or improving skin aging, comprising Lactobacillus plantarum , Bifidobacterium lactis or a combination thereof. The composition as described in claim 1, wherein the Lactobacillus plantarum strain GKM3 is deposited in the Food Industry Development Research Institute of the Foundation with the deposit number BCRC-910787; the Bifidobacterium lactis strain GKK2 is deposited with the Food Industry Foundation with the deposit number BCRC910826 Industrial Development Institute. The composition as described in Claim 1, which comprises culture medium components for cultivating and/or fermenting the Lactobacillus plantarum, Bifidobacterium lactis or a combination thereof. The composition according to any one of Claims 1 to 3, further comprising plant extracts or compounds that have the effect of improving skin transformation and/or inhibiting glycation reaction. The composition as described in claim 4, wherein the plant extracts that have the effect of improving skin transformation and/or inhibiting saccharification reaction include Cupressaceae, Ginkgoceae, Citrus, Perillaceae, Houttuyniaceae and/or Eucommiaceae Extracts of plants. The composition as described in claim 5, wherein the plant of the Cupressaceae comprises Podocarpus chinensis; the plant of the Ginkgo family comprises Ginkgo biloba; the plant of the Citrus family comprises one or more of corkberry, orange, grapefruit, pomelo and lemon; the The plants of the perilla family include one or more of mint, peppermint, perilla and rosemary; the plants of the family Houttuynia include Houttuynia cordata; and the plants of the Eucommia family include Eucommia. The composition as described in claim 4, wherein the compound having the effect of improving skin transformation and/or inhibiting glycation reaction includes equol. The composition as described in any one of Claims 1 to 7, which contains 0.1% by weight or more of Lactobacillus per unit of the composition, and a total of 0.001% by weight or more of the lactic acid bacteria that have the effect of improving skin transformation and/or inhibiting saccharification Reactive plant extracts or compounds. The composition described in any one of Claims 1 to 8 is a pharmaceutical composition, cosmetic, skin care product, nutritional product or food. The composition as described in claim 9, wherein the cosmetics include lotion, facial cleanser, beauty lotion, lotion or cream. A use of the composition described in any one of Claims 1 to 7 for preventing and/or improving skin aging. The use as described in claim 11, wherein the composition comprising Lactobacillus plantarum, Bifidobacterium lactis or a combination thereof is prepared by the following method: (a) Inoculate the colony of Lactobacillus in the medium for solid-state culture to obtain the bacteria; (b) inoculating the bacteria in step (a) into a liquid medium for liquid culture to obtain a mixture of liquid medium and bacteria; and (c) Fermenting the mixture of the liquid medium and the cells in step (b) to obtain a complete fermentation solution. The use as described in claim 12, wherein the method further includes the following steps: (d) centrifuging the whole fermentation liquid in step (c) to obtain the sludge and/or supernatant; and (e) Freeze-drying the sludge or the supernatant obtained in step (d) to obtain freeze-dried powder. The use as described in claim 11, wherein the improvement of skin aging includes inhibiting or slowing down the glycation reaction of the recipient by administering the composition comprising Lactobacillus plantarum, Bifidobacterium lactis or a combination thereof. The use as described in Claim 14, wherein said inhibiting or slowing down the glycation reaction of the recipient refers to reducing the content of pre-glycation products in the recipient. The use as described in Claim 15, wherein the pre-saccharification product contains Amadori. The use as described in Claim 14, wherein the inhibiting or slowing down of the glycation reaction of the recipient means that the content of dicarbonyl compounds in the mid-glycation stage of the recipient is reduced. The use as described in claim 17, wherein the mid-saccharification dicarbonyl compound comprises glyoxal (GO), methylglyoxal (MG) or a combination thereof. The use as described in claim 14, wherein the inhibiting or slowing down the glycation reaction of the recipient means that the content of the late glycation product of the recipient is reduced, or the content of the functional group contained in the product is improved. The use as described in claim 19, wherein the late glycation products include advanced glycation end products (AGEs). The use as described in claim 19, wherein the improvement of the content of the functional group includes a reduction of the content of carbonyl groups, an increase of the content of ε-amine groups or an increase of the content of free sulfhydryl groups. The use as described in claim 14, wherein the inhibiting or slowing down of the recipient's glycation response means that the recipient's high glycation end products (AGEs) and receptors for advanced glycation end products (RAGE) binding rate is reduced. The use as described in any one of claims 11 to 22, wherein the symptoms of skin aging include wrinkles, fine lines, spots, sagging or a combination thereof. The use as described in any one of claims 11 to 22, wherein the improvement of skin aging includes increasing the recipient's skin elasticity, increasing the skin moisture content, or a combination thereof.

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