Anti-allergic composition containing leukocyte extract as well as preparation method and application of anti-allergic composition

Abstract

The invention belongs to the technical field of cosmetics, and discloses an anti-allergic composition containing a leukocyte extract as well as a preparation method and application of the anti-allergic composition. The antiallergic composition comprises the following components: a leukocyte extract, polysaccharide, a tremella sporocarp extract and taurine, wherein the polysaccharide comprises fructo-oligosaccharide and inulin. According to the anti-allergic composition, on the basis of containing the leukocyte extract, the polysaccharide, the tremella sporocarp extract and the taurine are compounded, so that the synergistic effect can be achieved, the anti-allergic effect of the leukocyte extract is effectively improved, and the usage amount of the leukocyte extract in the field of skin care products can be reduced.

Description

Technical Field

[0001] The invention belongs to the technical field of cosmetics, and particularly relates to an anti-allergic composition containing a leukocyte extract, and a preparation method and application thereof. Background Art

[0002] Sensitive skin can cause a variety of skin problems: dehydration, wrinkles, spots and pustules, etc. Before solving these skin problems, it is necessary to repair the skin's internal skin barrier first. Anti-allergy is the most important means and key point. At present, sensitive skin care and anti-allergy technology have become hot spots in the research and development of related cosmetics. How to develop a safe, gentle and efficient anti-allergy product has become the research focus of related cosmetics.

[0003] Leukocyte extract is a cell-secreted complex extracted from leukocytes. It is one of the sources of cosmetic raw materials clearly listed in the "Catalogue of Used Cosmetic Raw Materials (2021 Edition)" issued by the State Food and Drug Administration. Leukocyte extract contains a large number of cell-specific proteins (PDGF, VEGF, HGF, PDGF, bFGF, TGF-β, EGF and hundreds of other repair factors), lipids, nucleic acids (DNA, RNA), etc., which can directly act on the mucosal layer. It has a high transdermal absorption rate and has the effects of promoting proliferation and division of fibroblasts, epithelial cells, endothelial cells, etc., promoting angiogenesis, mucosal repair and cell regeneration. These ingredients can effectively activate aging cells, promote cell regeneration, repair inflammatory damage, improve skin immunity, etc., to help cells in the body restore normal function and vitality.

[0004] At present, products containing leukocyte extracts, such as the invention patent with publication number CN117582402A, disclose a gel containing leukocyte extracts, which has a significant effect on postherpetic pain. However, the content of leukocyte extracts in the gel can be as high as 10%. The high content of leukocyte extracts not only leads to high costs, but also exceeds the historical usage amount clearly stated in the "Catalogue of Used Cosmetic Raw Materials (2021 Edition)" (the highest historical usage amount of resident products: 0.16%). Its toxicity and safety as a regular daily skin care product need to be investigated.

[0005] Therefore, developing a product with good anti-allergic effect and capable of reducing the amount of leukocyte extract used is a technical problem to be solved urgently in this field. Summary of the invention

[0006] The present invention aims to solve at least one of the technical problems existing in the above-mentioned prior art. To this end, the present invention provides an anti-allergic composition containing leukocyte extract and a preparation method and application thereof, wherein the content of leukocyte extract in the anti-allergic composition is low and has good anti-allergic effect.

[0007] The first aspect of the present invention provides an antiallergic composition, which comprises components: leukocyte extract, polysaccharide, Tremella fuciformis fruiting body extract and taurine, wherein the polysaccharide comprises oligofructose and inulin.

[0008] Specifically, oligofructose with a lower degree of polymerization has better skin permeability and can more easily reach the dermis of the skin, while inulin with a higher degree of polymerization stays more in the epidermis and stratum corneum of the skin. The addition of these two polysaccharides, perhaps due to the hydrated hydrogen bonds therein, will greatly enhance the hydration level of the skin surface, thereby improving the moisturizing property. This improved moisturizing property will enable the white blood cell extract to exert a better anti-allergic effect on the skin. It is speculated that this is because a high hydration environment is more conducive to the life movement of "biological" cells. On this basis, the ability of the white blood cell extract to enhance the skin cells' own ability to resist external stimuli is enhanced.

[0009] Specifically, the Tremella fruiting body extract is extracted from Tremella fruiting body, is a Tremella extract commonly found in the cosmetics related industry, has a molecular weight greater than one million Daltons, and has no other specific requirements. Tremella fruiting body extract, as a natural substance with film-forming properties, has a good moisturizing effect on the outer layer / stratum corneum of the skin. The present invention is based on a preliminary theory discovered and derived from a large number of experimental phenomena. In "highly hydrated" skin, the white blood cell extract has a more efficient anti-allergic effect, and through a large amount of experimental data, it is found that the Tremella fruiting body extract is helpful to improve the anti-allergic property of the white blood cell extract.

[0010] Specifically, the chemical name of taurine is 2-aminoethanesulfonic acid. Taurine itself is widely used as an auxiliary drug for analgesia in pharmacy. However, in the field of cosmetics, its external application effect is not significant. The present invention can greatly enhance the inhibitory effect of leukocyte extract on histamine by compounding taurine. It is believed that it can effectively enhance the anti-allergic effect of leukocyte extract and is one of the core components of the anti-allergic composition of the present invention.

[0011] In some embodiments of the present invention, the leukocyte extract accounts for 1-3% of the mass of the anti-allergic composition.

[0012] In some embodiments of the present invention, the polysaccharide accounts for 5-10% by weight of the anti-allergic composition.

[0013] In some embodiments of the present invention, the Tremella fruiting body extract accounts for 0.05-0.3% of the mass of the anti-allergic composition.

[0014] In some embodiments of the present invention, the taurine accounts for 0.1-1.2% by weight of the anti-allergic composition.

[0015] In some embodiments of the present invention, the mass ratio of the oligofructose to inulin is (4-9):1.

[0016] In some embodiments of the present invention, the degree of polymerization of the oligofructose is 2-9.

[0017] In some embodiments of the present invention, the degree of polymerization of the inulin is 30-60.

[0018] In some embodiments of the present invention, the antiallergic composition further comprises at least one of a pH adjuster, a preservative and water.

[0019] In some embodiments of the present invention, the pH adjuster includes sodium dihydrogen phosphate and / or disodium hydrogen phosphate.

[0020] In some embodiments of the present invention, the preservative includes at least one of phenoxyethanol, 1,2-pentanediol, 1,2-hexanediol and p-hydroxyacetophenone.

[0021] In some embodiments of the present invention, the pH value of the anti-allergic composition is 7.0-8.5. In the present invention, the pH value of the anti-allergic composition is adjusted to between 7.0-8.5, and the anti-allergic effect of the anti-allergic composition will be better and more stable.

[0022] The second aspect of the present invention provides a method for preparing the antiallergic composition according to the first aspect of the present invention, comprising the following steps:

[0023] The components are mixed to obtain the antiallergic composition.

[0024] In some embodiments of the present invention, the method for preparing the leukocyte extract comprises the following steps:

[0025] (1) Collect umbilical cord blood from rabbits and perform infectious disease antibody testing, virus testing, and endotoxin testing;

[0026] (2) adding an anticoagulant to the umbilical cord blood that has passed the test, centrifuging it, and taking the supernatant;

[0027] (3) Separating the supernatant by density gradient centrifugation to obtain the leukocyte extract.

[0028] It is understandable that the above-mentioned various tests include routine tests for hepatitis B, hepatitis C, AIDS, syphilis, cytomegalovirus, etc.; and can be detected by conventional detection methods including colloidal gold, fluorescent PCR, horseshoe crab reagent, etc.

[0029] In some embodiments of the present invention, the anticoagulant comprises EDTA (ethylenediaminetetraacetic acid) and / or ACD (acid citrate-dextrose solution).

[0030] In some embodiments of the present invention, in step (2), the centrifugal rotation speed is ≥2000 rpm, preferably 3000-4000 rpm.

[0031] In some embodiments of the present invention, the method for preparing the leukocyte extract further comprises step (5): washing the separated leukocyte extract with PBS buffer to remove residual separation solution and impurities.

[0032] In some embodiments of the present invention, the method for preparing the leukocyte extract further comprises step (6): after washing in step (5), suspending the leukocyte extract in PBS buffer and refrigerating for later use.

[0033] In some embodiments of the present invention, in the method for preparing the leukocyte extract, the PBS buffer used is sodium dihydrogen phosphate and disodium hydrogen phosphate buffer, and the pH value of the PBS buffer is 7.0-8.5.

[0034] The third aspect of the present invention provides a cosmetic comprising the antiallergic composition according to the first aspect of the present invention.

[0035] In some embodiments of the present invention, the anti-allergic composition accounts for 0.01-5% of the mass of the cosmetic.

[0036] In some embodiments of the present invention, the cosmetic comprises any one of lotion, essence, emulsion, cream, mask or gel.

[0037] Compared with the prior art, the beneficial effects of the present invention are as follows:

[0038] (1) The anti-allergic composition of the present invention, on the basis of containing leukocyte extract, can exert a synergistic effect by compounding polysaccharide, Tremella fruiting body extract and taurine, and effectively improves the anti-allergic effect of leukocyte extract, which can reduce the usage of leukocyte extract in the field of skin care products.

[0039] (2) The leukocyte extract in the anti-allergic composition of the present invention is derived from rabbit umbilical cord blood. Rabbits have the advantage of fast reproduction and lower production cost. In addition, the leukocyte extract of the present invention is optimized in terms of the process of anticoagulant type, pH value of PBS buffer, and centrifugal speed, which can further improve the quality of the leukocyte extract and is conducive to industrial development. DETAILED DESCRIPTION

[0040] In order to make the technical scheme of the present invention more clearly understood by those skilled in the art, the following embodiments are listed for illustration. It should be pointed out that the following embodiments do not limit the protection scope of the present invention.

[0041] Unless otherwise specified, the raw materials, reagents or devices used in the present invention can be obtained from conventional commercial channels or by existing known methods.

[0042] Examples 1-4

[0043] Examples 1-4 respectively provide an antiallergic composition, and the specific formula is shown in Table 1.

[0044] Table 1

[0045]

[0046]

[0047] Among them, the polysaccharide is composed of oligofructose and inulin in a mass ratio of 6:1.

[0048] The leukocyte extract is extracted from rabbit umbilical cord blood, and the preparation method thereof comprises the following steps:

[0049] (1) Collect rabbit umbilical cord blood safely and aseptically, add EDTA (ethylenediaminetetraacetic acid) anti-gel, and refrigerate for later use;

[0050] (2) Conduct infectious disease antibody testing, virus testing, and endotoxin testing on the collected umbilical cord blood. If the test result is negative, centrifuge the blood, take the supernatant, and refrigerate it for later use; the centrifugation speed is 3500 rpm and the time is 30 min;

[0051] (3) using density gradient centrifugation to further separate the supernatant to obtain a leukocyte extract;

[0052] (4) washing the leukocyte extract with PBS buffer (pH 7.5) to remove residual separation solution and impurities;

[0053] (5) After washing, add the leukocyte extract to PBS buffer and refrigerate until use.

[0054] The antiallergic compositions of Examples 1-4 above are prepared by conventional mixing.

[0055] Comparative Example 1

[0056] The only difference from Example 1 is that the leukocyte extract is missing, and the missing amount is evenly distributed among the polysaccharide, Tremella fruiting body extract and taurine. The remaining components, contents and preparation method are the same as those in Example 1.

[0057] Comparative Example 2

[0058] The only difference from Example 1 is that polysaccharide, Tremella fruiting body extract and taurine are missing, and the missing amounts are supplemented by leukocyte extract. The other components, contents and preparation method are the same as Example 1.

[0059] Comparative Example 3

[0060] The only difference from Example 1 is that oligofructose is missing and the missing amount is supplemented by inulin. The other components, contents and preparation method are the same as those in Example 1.

[0061] Comparative Example 4

[0062] The only difference from Example 1 is that an equal amount of Tremella fuciformis fruiting body extract is replaced with sodium hyaluronate with similar efficacy, and the remaining components, contents and preparation method are the same as in Example 1.

[0063] Comparative Example 5

[0064] The only difference from Example 1 is that taurine is replaced with glutamic acid with similar efficacy in equal amount, and the other components, contents and preparation method are the same as those in Example 1.

[0065] Comparative Example 6

[0066] This example uses a dipotassium glycyrrhizinate aqueous solution with a mass concentration of 1%.

[0067] Application Examples 1-4

[0068] Application Examples 1-4 respectively provide a gel, and the specific formula is shown in Table 2.

[0069] Table 2

[0070]

[0071] The gels of the above application examples 1-4 are prepared by conventional mixing.

[0072] Comparative application example 1

[0073] The only difference from Application Example 1 is that the antiallergic composition of Example 1 is replaced by the antiallergic composition of Comparative Example 1 in equal amounts, and the remaining components, contents and preparation methods are the same as those of Application Example 1.

[0074] Comparative Application Example 2

[0075] The only difference from Application Example 1 is that the antiallergic composition of Example 1 is replaced by the antiallergic composition of Comparative Example 2 in equal amounts, and the remaining components, contents and preparation methods are the same as those of Application Example 1.

[0076] Comparative Application Example 3

[0077] The only difference from Application Example 1 is that the antiallergic composition of Example 1 is replaced by the antiallergic composition of Comparative Example 3 in equal amounts, and the remaining components, contents and preparation methods are the same as those of Application Example 1.

[0078] Comparative Application Example 4

[0079] The only difference from Application Example 1 is that the antiallergic composition of Example 1 is replaced by the antiallergic composition of Comparative Example 4 in equal amounts, and the remaining components, contents and preparation methods are the same as those of Application Example 1.

[0080] Comparative Application Example 5

[0081] The only difference from Application Example 1 is that the antiallergic composition of Example 1 is replaced by the antiallergic composition of Comparative Example 5 in equal amounts, and the remaining components, contents and preparation methods are the same as those of Application Example 1.

[0082] Comparative Application Example 6

[0083] The only difference from Application Example 1 is that the antiallergic composition of Example 1 is replaced by an equal amount of the dipotassium glycyrrhizinate aqueous solution of Comparative Example 6, and the remaining components, contents and preparation methods are the same as those of Application Example 1.

[0084] Product effect testing

[0085] Test Example 1

[0086] Modern pharmacological studies have shown that the level of hyaluronidase increases rapidly during inflammation, further inducing the occurrence and development of inflammation. At this time, if hyaluronidase inhibitors are used, the occurrence and development of inflammation can be prevented.

[0087] Test method:

[0088] 1. The gels of Application Examples 1-4 and Comparative Application Examples 1-6 were respectively prepared into dilute solutions with a mass concentration of 20% as samples to be tested;

[0089] 2. Add 100 μL of the sample to be tested to each reaction tube, add 100 μL of double distilled water to the blank control tube, add 50 μL of hyaluronidase to each tube, and incubate in a 37°C water bath for 20 minutes; add 100 μL of calcium chloride with a concentration of 12.5 mM / L to each tube, and incubate in a 37°C water bath for 20 minutes;

[0090] 3. After hyaluronidase is activated, add 250 μL of sodium hyaluronate, mix well, and place in a water bath at 37°C for 40 minutes. Add 100 μL each of 0.4 mM / L sodium hydroxide and 0.4 mM / L potassium borate, place in a water bath in boiling water for 3 minutes, cool to room temperature, and pipette 200 μL from each tube into a 96-well microplate test well. Measure 3 wells in each tube and take the average value.

[0091] 4. Use Shimadzu CS-9301PC ELISA analyzer to measure OD value at wavelength 585nm, use double distilled water as blank control group, and calculate hyaluronidase activity inhibition rate by OD value: hyaluronidase activity inhibition rate % = [(OD 空白对照组 -OD 样品组 ) / OD 空白对照组 ]*100%.

[0092] The test results are shown in Table 3.

[0093] Table 3

[0094] sample The average value of three test data Inhibition rate Application Example 1 0.254 77.01％ Application Example 2 0.235 78.73％ Application Example 3 0.1898 82.82％ Application Example 4 0.224 79.73％ Comparative application example 1 0.987 10.68％ Comparative Application Example 2 0.568 48.60％ Comparative Application Example 3 0.425 61.54％ Comparative Application Example 4 0.487 55.93％ Comparative Application Example 5 0.435 60.63％ Comparative Application Example 6 0.358 67.60％ Blank control group 1.105 -

[0095] As can be seen from Table 3, the gels provided by Application Examples 1-4 of the present invention all have excellent hyaluronidase inhibitory effects, and the inhibitory effects are significantly better than those of Comparative Application Examples 1-6, indicating that the gels prepared by the anti-allergic composition of the present invention have efficient anti-allergic ability. Compared with traditional antiallergic agents (dipotassium glycyrrhizinate), single leukocyte extracts, lack of leukocyte extracts, and other component replacement combinations, the anti-allergic composition of the present invention is compounded with leukocyte extracts, polysaccharides, Tremella fruiting body extracts, and taurine, which can play a synergistic role and significantly and effectively improve the anti-allergic effect of the leukocyte extracts.

[0096] Test Example 2

[0097] Test method: 110 mice, half male and half female, were randomly divided into 11 groups, 10 mice in each group; in the experiment, the hair on the back of the mice was cut with scissors, and then washed with depilatory cream to expose the skin, and then the 11 groups of mice were respectively given drugs, all of which were smeared and the dosage was shown in Table 4. Each group was given once a day for 5 consecutive days. One hour after the last dose, 0.2 mL of 0.02% dextran saline injection was injected into the tail vein of each mouse. The number of scratching times within 30 minutes with itching, tail biting, back scratching, etc. as the itching characteristics of the mice was observed, and the average value was obtained and recorded as N. Inhibition rate % = = [(N 纯净水抓痒次数 -N 样品组抓痒次数 ) / N 纯净水抓痒次数 ]*100%.

[0098] The test results are shown in Table 5.

[0099] Table 4

[0100]

[0101] Table 5

[0102]

[0103]

[0104] As shown in Table 5, compared with the data of the traditional dipotassium glycyrrhizinate (Comparative Application Example 6), the gel using the anti-allergic composition of the present invention has a highly effective anti-allergic effect. Compared with the data of the other comparative application examples 1-5, the anti-allergic composition of the present invention can play a synergistic role by compounding leukocyte extract, polysaccharide, Tremella fruiting body extract and taurine, and effectively improves the anti-allergic effect of the leukocyte extract.

[0105] It can be seen that the anti-allergic composition of the present invention is more suitable for use as an anti-allergic agent in the cosmetics field than traditional dipotassium glycyrrhizinate and single leukocyte extract, and can produce more efficient, safe and mild anti-allergic cosmetics.

[0106] The preferred embodiments of the present invention are described in detail above, but the invention is not limited to the embodiments. Those skilled in the art may make various equivalent modifications or substitutions without violating the spirit of the invention, and these equivalent modifications or substitutions are all included in the scope defined by the claims of this application.

Claims

1. An antiallergic composition, characterized in that: The anti-allergic composition comprises components: leukocyte extract, polysaccharide, tremella fruiting body extract and taurine, wherein the polysaccharide comprises oligofructose and inulin.

2. The antiallergic composition according to claim 1, characterized in that The leukocyte extract accounts for 1-3% of the mass of the anti-allergic composition; and / or, the polysaccharide accounts for 5-10% of the mass of the anti-allergic composition; and / or, the Tremella fruiting body extract accounts for 0.05-0.3% of the mass of the anti-allergic composition; and / or, the taurine accounts for 0.1-1.2% of the mass of the anti-allergic composition.

3. The antiallergic composition according to claim 1, characterized in that The mass ratio of the oligofructose to inulin is (4-9):

1.

4. The antiallergic composition according to claim 1, characterized in that: The polymerization degree of the oligofructose is 2-9, and the polymerization degree of the inulin is 30-60.

5. The antiallergic composition according to claim 1, characterized in that The pH value of the antiallergic composition is 7.0-8.

5.

6. The method for preparing the antiallergic composition according to any one of claims 1 to 5, characterized in that: The following steps are involved: The components are mixed to obtain the antiallergic composition.

7. The preparation method according to claim 6, characterized in that: The preparation method of the leukocyte extract comprises the following steps: (1) Collect umbilical cord blood from rabbits and perform infectious disease antibody testing, virus testing, and endotoxin testing; (2) adding an anticoagulant to the umbilical cord blood that has passed the test, centrifuging it, and taking the supernatant; (3) Separating the supernatant by density gradient centrifugation to obtain the leukocyte extract.

8. A cosmetic, characterized in that: The antiallergic composition comprises the antiallergic composition according to any one of claims 1 to 5.

9. The cosmetic according to claim 8, characterized in that: The antiallergic composition accounts for 0.01-5% of the weight of the cosmetic.

10. The cosmetic according to claim 8, characterized in that: The cosmetics include any one of lotion, essence, emulsion, cream, mask or gel.

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