An acne gel containing lysozyme

Abstract

The present invention relates to an acne-removing gel preparation, wherein the weight ratio of the active ingredients lysozyme, puerarin and formononetin is 1:(1-5):(1-5). The preparation method of the acne-removing gel preparation of the present invention is simple and the quality is stable. While the three active ingredients play their respective functions, they can also synergistically treat acne caused by bacteria such as Propionibacterium acnes. In addition, lysozyme, puerarin and formononetin are naturally sourced and have no toxic side effects.

Description

Technical Field

[0001] The present invention relates to the field of medicine, and particularly to an acne-removing gel containing lysozyme. Background Art

[0002] Acne is a multi-factorial skin disease, mainly related to increased sebum production, hyperkeratosis of the epithelial cells at the hair follicle orifice, and proliferation of Propionibacterium acnes in the hair follicle. In addition, the selective expression of genes affects organ development, so it is also related to genetics. The development of sebaceous glands and the production of sebum are controlled by androgens, and the increase in androgens is affected by factors such as endocrine, living environment, and age. Sebaceous glands in acne patients are larger, and the sebaceous glands secrete more than normal people. Due to the increase in sebaceous gland secretion, the level of linoleic acid in sebum is relatively reduced, affecting the synthesis of fat, resulting in a lack of fatty acids in the follicular epithelium, thereby inducing excessive follicular keratinization, preventing the normal shedding of epithelial cells, leading to an excessive reduction in the hair follicle orifice, and the sebum cannot be discharged smoothly, accumulating at the hair follicle orifice, forming comedones.

[0003] Propionibacterium acnes is an anaerobic bacterium, mainly parasitizing in the hair follicle, feeding on oil, and is the main characteristic bacterium of acne. With the increase in free fatty acids, the accumulated fatty acids cause a non-specific inflammatory reaction around the hair follicle. In addition, the main pathogenic bacteria of acne also include Malassezia furfur and Staphylococcus epidermidis, etc. Propionibacterium acnes can also secrete low-molecular polypeptides, attracting neutrophils into the hair follicle, resulting in the release of their hydrolytic enzymes, causing the leakage and rupture of the hair follicle epithelium, and causing the hair follicle contents to enter the surrounding dermal tissue, resulting in a series of clinical manifestations from simple inflammation to papules, abscesses, nodules, and cysts.

[0004] Currently, most of the reported acne-removing products in the literature use the cream dosage form. However, one of the reasons for inducing acne is excessive oil secretion. Therefore, the treatment effect of acne-removing creams is not good, and the use feeling is also not ideal. The key points of daily care for acne patients are to control oil and replenish water, and regulate the balance of oil secretion. Cosmetic dosage forms with less oil content and good moisturizing properties should be selected. A gel is a thick liquid or semi-solid preparation in the form of a homogeneous, suspension or emulsion made from a drug and an excipient capable of forming a gel. It has the advantages of wide biocompatibility, slow release and controlled release of drugs, etc. Gel preparations are mainly made by simply mixing a gel matrix after swelling with a moisturizer, a skin conditioner, etc. They have less oil content and can form a water-locking film on the skin surface. Therefore, gel preparations are an ideal dosage form for developing acne-removing products.

[0005] Lysozyme, also known as muramidase or N-acetylmuramoyl glycanohydrolase, is an alkaline enzyme that can hydrolyze mucopolysaccharides in pathogenic bacteria. It mainly destroys the β-1,4 glycosidic bond between N-acetylmuramic acid and N-acetylglucosamine in the cell wall, decomposes the insoluble mucopolysaccharide in the cell wall into soluble glycopeptides, and causes the cell wall to rupture and the contents to escape, resulting in the dissolution of bacteria.

[0006] Pueraria lobata (Willd.) Ohwi is the dried root of the leguminous plant Pueraria lobata, which has the effects of relieving muscle fever, inducing measles, promoting fluid production to quench thirst, and lifting yang to stop diarrhea. Existing pharmacological animal experiments have shown that the active ingredients in Pueraria lobata have the effects of improving myocardial metabolism and microcirculation of blood vessels, dilating coronary blood vessels, relieving hypertension, preventing arrhythmia, improving cerebrovascular circulation, exerting estrogen-like effects, protecting the liver and kidneys, scavenging free radicals, relieving symptoms of hypoxia, anti-tumor, regulating the body's immune function, and relieving alcohol intoxication. Most of the flavonoid compounds in Pueraria lobata are isoflavonoids. So far, researchers have isolated dozens of flavonoid compounds from Pueraria lobata, including puerarin, daidzin, daidzein, genistein, etc.

[0007] Puerarin is the main active ingredient extracted from Pueraria lobata. Its structural formula is 8-β-D-glucopyranosyl-4',7-dihydroxyisoflavone. It is a white needle crystal, slightly soluble in water, and has the effects of protecting the cardiovascular and cerebrovascular systems and nerve cells. It can dilate blood vessels, lower blood pressure, lower blood sugar, anti-tumor, improve the body's immunity, antioxidant, anti-inflammatory, and regulate bone metabolism. Currently, it is widely used in the clinical treatment of diseases such as coronary heart disease, angina pectoris, diabetes and its complications, gastric cancer, colon cancer, arthritis, osteoporosis, etc.

[0008] Formononetin is an isoflavonoid phytoestrogen, which is the main active ingredient of the leguminous plant Trifolium pratense L. Strong evidence of formononetin in promoting apoptosis and anti-proliferation of tumor cells indicates that it can be used as a drug against various cancers, and these anti-cancer properties can be observed in various in vitro cancer cell models such as breast cancer, colorectal cancer, and prostate cancer. It is also found in various in vivo studies that formononetin has the effect of inhibiting tumor growth and metastasis. Further research found that formononetin can also inhibit tumor cell migration and invasion and induce cell cycle arrest.

[0009] Currently, there is no report on the combination of lysozyme, puerarin, and formononetin for use in the field of acne gel. Summary of the Invention

[0010] The technical problem to be solved by the present invention is to provide an acne gel preparation that can effectively treat acne and whose active ingredients are derived from natural sources.

[0011] The technical solution adopted by the present invention to solve the above problems is to provide an acne gel preparation, in which the weight ratio of the active ingredients lysozyme, puerarin, and formononetin is 1:(1 - 5):(1 - 5).

[0012] Preferably, in the acne gel preparation, the weight ratio of lysozyme, puerarin, and formononetin is 1:(1 - 3):(1 - 3).

[0013] More preferably, in the acne-removing gel preparation, the weight ratio of lysozyme, puerarin, and formononetin is 1:3:1.

[0014] Preferably, the lysozyme is extracted from egg white.

[0015] Preferably, the acne-removing gel preparation further comprises pharmaceutically acceptable excipients.

[0016] More preferably, the pharmaceutically acceptable excipients are selected from one or more of a gel matrix, a humectant, a solubilizer, and a solvent.

[0017] Even more preferably, the pharmaceutically acceptable excipients are a gel matrix, a humectant, a solubilizer, and a solvent.

[0018] Preferably, the gel matrix is selected from one or more of carbomer, poloxamer, and hydroxypropyl methylcellulose; more preferably, the gel matrix is a composition of carbomer and poloxamer; even more preferably, the gel matrix is a composition of carbomer 940 and poloxamer 407, and the weight ratio of carbomer 940 to poloxamer 407 is (1:5)-(5:1); most preferably, the gel matrix is a composition of carbomer 940 and poloxamer 407 with a weight ratio of 2:1.

[0019] Preferably, the humectant is glycerol or propylene glycol; more preferably, the humectant is glycerol.

[0020] Preferably, the solubilizer is Tween 80 or Tween 60; more preferably, the solubilizer is Tween 80.

[0021] Preferably, the solvent is distilled water.

[0022] More preferably, the pharmaceutically acceptable excipients are carbomer 940, poloxamer 407, glycerol, Tween 80, and distilled water.

[0023] Even more preferably, the acne-removing gel preparation is made from the following components by weight percentage: 0.2% lysozyme, 0.6% puerarin, 0.2% formononetin, 6% carbomer 940, 3% poloxamer 407, 15% glycerol, 5% Tween 80, and 70% distilled water.

[0024] The second aspect of the present invention provides a preparation method of the above acne-removing gel preparation, comprising the following steps:

[0025] (1) Mix lysozyme, puerarin, and formononetin with a solubilizer uniformly to obtain a liquid medicine;

[0026] (2) Add a humectant to the gel matrix, mix evenly, and then mix with the liquid medicine prepared in step (1);

[0027] (3) Add a solvent to the solution obtained in step (2), stir to swell, and let stand for 24 hours to obtain the product.

[0028] The third aspect of the present invention also provides the use of the above acne treatment gel preparation in the preparation of a drug for inhibiting bacteria.

[0029] Preferably, the bacteria are Propionibacterium acnes.

[0030] The fourth aspect of the present invention also provides the use of the above acne treatment gel preparation in the preparation of a drug for treating acne.

[0031] The fifth aspect of the present invention also provides the use of the above acne treatment gel preparation in the preparation of a drug for reducing the content of TNF-α and / or IL-6 in serum.

[0032] Preferably, the present invention also provides the use of the above acne treatment gel preparation in the preparation of a drug for simultaneously reducing the content of TNF-α and IL-6 in serum.

[0033] The present invention has positive and beneficial effects:

[0034] Surprisingly, through repeated experiments, the present invention unexpectedly found that the combined use of lysozyme, puerarin, and formononetin has a synergistic effect on inhibiting bacterial growth. The preparation method of the acne treatment gel preparation of the present invention is simple and the quality is stable. While the three active ingredients exert their respective effects, they can synergistically treat acne caused by bacteria such as Propionibacterium acnes. In addition, lysozyme, puerarin, and formononetin are naturally sourced and have no toxic side effects. Specific Embodiments

[0035] The following further illustrates the present invention with reference to examples, but the embodiments of the present invention are not limited thereto. The experimental methods used in the following examples are all conventional methods unless otherwise specified. The lysozyme used in the examples and test examples is all extracted from egg white.

[0036] Example 1. Acne Treatment Gel Preparation G1 Containing Lysozyme

[0037] Mix 0.2 g of lysozyme, 0.6 g of puerarin, and 0.2 g of formononetin with 5 g of Tween 80 to obtain a liquid medicine; add 15 g of glycerol to a mixture of 6 g of Carbomer 940 and 3 g of Poloxamer 407, mix well and then mix with the liquid medicine; continue to add 70 g of distilled water, stir to swell, and let stand for 24 hours to obtain the acne treatment gel preparation G1.

[0038] Example 2. Acne Treatment Gel Preparation G2 Containing Lysozyme

[0039] Mix 0.2 g of lysozyme, 0.4 g of puerarin, and 0.4 g of formononetin uniformly with 5 g of Tween 80 to obtain a liquid medicine; add 15 g of glycerol to a mixture of 6 g of carbomer 940 and 3 g of poloxamer 407, mix well, and then mix with the liquid medicine; continue to add 70 g of distilled water, stir to swell, and let stand for 24 hours to obtain the acne-removing gel preparation G2.

[0040] Example 3. Acne-removing gel preparation G3 containing lysozyme

[0041] Mix 0.2 g of lysozyme, 0.2 g of puerarin, and 0.6 g of formononetin uniformly with 5 g of Tween 80 to obtain a liquid medicine; add 15 g of glycerol to a mixture of 6 g of carbomer 940 and 3 g of poloxamer 407, mix well, and then mix with the liquid medicine; continue to add 70 g of distilled water, stir to swell, and let stand for 24 hours to obtain the acne-removing gel preparation G3.

[0042] Example 4. Acne-removing gel preparation G4 containing lysozyme

[0043] Mix 0.2 g of lysozyme, 0.6 g of puerarin, and 0.2 g of formononetin uniformly with 5 g of Tween 80 to obtain a liquid medicine; add 15 g of glycerol to a mixture of 4.5 g of carbomer 940 and 4.5 g of poloxamer 407, mix well, and then mix with the liquid medicine; continue to add 70 g of distilled water, stir to swell, and let stand for 24 hours to obtain the acne-removing gel preparation G4.

[0044] Example 5. Acne-removing gel preparation G5 containing lysozyme

[0045] Mix 0.2 g of lysozyme, 0.6 g of puerarin, and 0.2 g of formononetin uniformly with 5 g of Tween 80 to obtain a liquid medicine; add 15 g of glycerol to a mixture of 3 g of carbomer 940 and 6 g of poloxamer 407, mix well, and then mix with the liquid medicine; continue to add 70 g of distilled water, stir to swell, and let stand for 24 hours to obtain the acne-removing gel preparation G5.

[0046] Comparative Example 1. Acne-removing gel preparation C1 containing lysozyme

[0047] Mix 0.2 g of lysozyme and 0.8 g of puerarin uniformly with 5 g of Tween 80 to obtain a liquid medicine; add 15 g of glycerol to a mixture of 6 g of carbomer 940 and 3 g of poloxamer 407, mix well, and then mix with the liquid medicine; continue to add 70 g of distilled water, stir to swell, and let stand for 24 hours to obtain the acne-removing gel preparation C1.

[0048] Comparative Example 2. Acne-removing gel preparation C2 containing lysozyme

[0049] Mix 0.2 g of lysozyme, 0.8 g of formononetin and 5 g of Tween 80 evenly to obtain a liquid medicine; add 15 g of glycerol to a mixture of 6 g of Carbomer 940 and 3 g of Poloxamer 407, mix evenly and then mix with the liquid medicine; continue to add 70 g of distilled water, stir and swell, and let stand for 24 hours to obtain the anti-acne gel preparation C2.

[0050] Comparative Example 3, anti-acne gel preparation C3 containing lysozyme

[0051] Mix 0.2 g of lysozyme, 0.6 g of puerarin and 0.2 g of formononetin with 5 g of Tween 80 evenly to obtain a liquid medicine; add 15 g of glycerol to 9 g of Carbomer 940, mix evenly and then mix with the liquid medicine; continue to add 70 g of distilled water, stir and swell, and let stand for 24 hours to obtain the anti-acne gel preparation C3.

[0052] Comparative Example 4, anti-acne gel preparation C4 containing lysozyme

[0053] Mix 0.2 g of lysozyme, 0.6 g of puerarin and 0.2 g of formononetin with 5 g of Tween 80 evenly to obtain a liquid medicine; add 15 g of glycerol to 9 g of Poloxamer 407, mix evenly and then mix with the liquid medicine; continue to add 70 g of distilled water, stir and swell, and let stand for 24 hours to obtain the anti-acne gel preparation C4.

[0054] Test Example 1, efficacy of the anti-acne gel preparation of the present invention on the rabbit ear acne model

[0055] 1. Test method

[0056] On the inner side of the left ear of the rabbits in the model group, apply 2% coal tar solution once a day at an area of about 2 cm × 2 cm at the opening of the ear duct, 0.5 mL each time, for 14 consecutive days. The blank group (10 rabbits) was given an equal dose of normal saline. After 14 days, in the model group, inoculate 50 μL of Propionibacterium acnes suspension with a 0.5 McFarland turbidity standard intradermally into the rabbit ear every other day. If visible cysts appear, the inoculation can be stopped. After injection, measure the auricle thickness once a day for 7 consecutive days. The rabbits with established models were randomly divided into a model group, the groups of Examples 1-3 of the present invention and the groups of Comparative Examples 1-2, with 10 rabbits in each group. Apply the corresponding drug / matrix at an area of 2 cm × 2 cm at the modeling site, and administer the drug continuously for 14 days, once a day. Among them, the groups of Examples 1-3 and the groups of Comparative Examples 1-2 apply the anti-acne gels prepared in Examples 1-3 and Comparative Examples 1-2 at a dose of 0.5 g / kg body weight per day. The blank group and the model group apply an equal dose of blank gel matrix without active ingredients (the matrix formula is the same as that of Example 1) every day. 24 hours after the last administration of each test group of rabbits, take blood from the heart, centrifuge at 3200 r / min at 4 °C for 20 min, take the supernatant, and determine the contents of TNF-α and IL-6 in the serum by ELISA.

[0057] 2. Test results

[0058] The effects of each experimental group of the present invention on the rabbit ear acne model are shown in Table 1 below.

[0059] Table 1 Efficacy of the anti-acne gel of the present invention on the rabbit ear acne model

[0060]

[0061]

[0062] The test results in Table 1 above show that the levels of TNF-α and IL-6 in the serum of the model rabbits treated with the anti-acne gels of Examples 1-3 of the present invention are significantly lower than those of the model group (with significant differences compared to the model group), indicating that the anti-acne gel of the present invention can effectively treat acne caused by Propionibacterium acnes and has good clinical anti-acne effects. In addition, the treatment effects of Examples 1-3 are also better than those of Comparative Examples 1-2 (formononetin and puerarin were omitted from the active ingredients respectively), demonstrating that on the basis of the same total amount of active ingredients (the total amount of active ingredients is 1 g / 100 g gel preparation), the combination of the three active ingredients of the present invention is significantly superior to the combination of two active ingredients in the treatment of acne, indicating that the three active ingredients produce a synergistic effect. It should be noted that the anti-acne gel containing lysozyme, puerarin and formononetin in a weight ratio of 1:3:1 (Example 1) has an even better treatment effect than the preparations with other active drug ratios (Examples 2-3), producing unexpectedly excellent effects.

[0063] Test Example 2 Study on the chemical stability of the anti-acne gel preparation of the present invention

[0064] 1. Test method

[0065] In this test, 5 test groups were set up. Among them, in test groups 1-3, the anti-acne gel preparations of the present invention prepared in Examples 1 and 4-5 were used respectively, and in test groups 4-5, the anti-acne gel preparations prepared in Comparative Examples 3-4 were used. Each test group had 3 repeated samples. The above 5 test groups were stored at 60 °C and 75% RH for 2 weeks respectively. After taking them out, the contents of puerarin and formononetin in the preparations of each test group were measured respectively, and the average values of the contents of puerarin and formononetin after storage in each test group were calculated.

[0066] 2. Test results

[0067] The average values of the contents of puerarin and formononetin in each test group are shown in Table 2 below.

[0068] Table 2 Contents of puerarin and formononetin in the anti-acne gel preparation of the present invention under high temperature and high humidity

[0069] Experimental group Puerarin content (%) Formononetin content (%) Experimental group 1 98.42％ 98.58％ Experimental group 2 97.27％ 96.51％ Experimental group 3 96.54％ 97.38％ Experimental group 4 93.60％ 92.75％ Experimental group 5 92.96％ 93.83％

[0070] From the above data on the content of active ingredients, it can be seen that after the gel preparations of Test Groups 1-3 (Examples 1 and 4-5) were stored under high temperature and high humidity conditions, the contents of puerarin and formononetin did not decrease significantly, and the contents of active ingredients could be maintained above 96%. In contrast, in Test Group 4 (Comparative Example 3), poloxamer 407 was omitted, and in Test Group 5 (Comparative Example 4), carbomer 940 was omitted. After storage, the contents of puerarin and formononetin in the gel preparations were less than 94%, indicating that a large amount of impurities were generated from the two active ingredients during storage, and the stability was poor. The above comparative test results show that the gel matrix selected in the present invention can effectively inhibit the generation of impurities of puerarin and formononetin, and it is expected that the gel preparation of the present invention has good stability during production and storage.

Claims

1. A acne treatment gel preparation, characterized in that, The weight ratio of the active ingredients lysozyme, puerarin and formononetin is 1:3:

1.

2. The anti-acne gel preparation according to claim 1, characterized in that, The acne treatment gel preparation further comprises pharmaceutically acceptable excipients selected from one or more of a gel matrix, a humectant, a solubilizer and a solvent.

3. The acne treatment gel preparation according to claim 2, wherein, The gel matrix is selected from one or more of carbomer, poloxamer and hydroxypropyl methylcellulose.

4. The anti-acne gel preparation according to claim 3, wherein The gel matrix is a composition of carbomer and poloxamer, and the weight ratio of carbomer 940 to poloxamer 407 is (1:5)-(5:1).

5. The acne treatment gel preparation according to claim 3, wherein The gel matrix is a composition of carbomer 940 and poloxamer 407 with a weight ratio of 2:

1.

6. The anti-acne gel preparation according to claim 2, wherein The humectant is glycerol or propylene glycol.

7. The acne treatment gel preparation according to claim 2, characterized in that, The solubilizer is Tween 80 or Tween 60.

8. The preparation method of the acne-removing gel preparation according to any one of claims 2-7, characterized in that, It includes the following steps: (1) Mix lysozyme, puerarin and formononetin with the solubilizer evenly to obtain a liquid medicine. (2) Add the humectant to the gel matrix, mix evenly, and then mix with the liquid medicine prepared in step (1). (3) Add the solvent to the solution obtained in step (2), stir and swell, and let it stand for 24 hours to obtain the product.

9. Use of the acne treatment gel preparation according to any one of claims 1-7 in the preparation of a drug for inhibiting bacteria, wherein the bacteria are Propionibacterium acnes.

10. Use of the acne treatment gel preparation according to any one of claims 1-7 in the preparation of a drug for treating acne.

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