A Tibetan medicine composition with bacteriostatic, antioxidant and anti-allergic effects, and a preparation method and application thereof

Abstract

This invention belongs to the field of Tibetan medicine technology, and specifically relates to a Tibetan medicine composition with antibacterial, antioxidant, and anti-allergic effects, its preparation method, and its application. The raw materials of this Tibetan medicine composition include Phyllanthus emblica, Terminalia chebula, and Terminalia chebula in a mass ratio of (6-8):(6-8):1. Within this mass range, the raw materials can produce a synergistic effect, not only achieving excellent antibacterial and anti-allergic efficacy but also possessing antioxidant properties, reducing free radical damage to the skin or oral cavity. Therefore, it can be used to prepare skin or oral products, expanding the application scope of Tibetan medicine compound formulas and enhancing their development value.

Description

Technical Field

[0001] This invention belongs to the field of Tibetan medicine technology, and in particular relates to a Tibetan medicine composition with antibacterial, antioxidant and anti-allergic effects, its preparation method and application. Background Technology

[0002] In recent years, with the rapid development of the social economy and the continuous improvement of people's living standards, health awareness has also been constantly strengthened. People's comprehensive requirements for daily chemical products are also constantly increasing. On the one hand, people hope that products have multiple functions, such as good antibacterial, anti-allergic, moisturizing, and skin-brightening effects. On the other hand, people hope that products have higher safety, such as natural raw materials, gentle formulas, and green and environmentally friendly production processes. Therefore, in oral care and skin cleansing products, products with good antibacterial and anti-allergic effects and natural raw materials are increasingly favored by consumers.

[0003] Skin, the tissue covering the muscles on the body's surface, is the largest organ in the human body, primarily responsible for protecting the body, sweating, and sensing temperature and pressure. Covering the entire body, skin protects various tissues and organs from physical, mechanical, chemical, and pathogenic microbial attacks. Human skin contains two main groups of microorganisms: resident flora and transient flora. Resident flora are hierarchical and orderly colonizing the skin, multiplying on the surface of normal human skin, and generally not pathogenic; they can only cause disease when systemic or local immunity is weakened. Transient flora refers to a group of bacteria acquired from the external environment through contact, mainly including Staphylococcus aureus, Streptococcus spp., Micrococcus spp., anaerobic Staphylococcus spp., Neisseria spp., dermatophytes, and some Gram-negative bacilli (such as Escherichia coli, Pseudomonas aeruginosa, Proteus spp., and Alcaligenes spp.). Transient flora, generally originating from the environment, may cause skin or other diseases due to their unique pathogenic potential. Effectively inhibiting harmful bacteria and maintaining a healthy skin microecological environment has become a new requirement for consumers' skin cleansing and care products. Furthermore, with societal development, people have more opportunities to come into contact with various allergens in their daily lives, making their skin more sensitive. Sensitive skin is typically prone to thinning, incomplete stratum corneum, peeling, tightness, and dryness, and is more susceptible to pathogenic bacteria. Therefore, how to resolve and balance the effects of antibacterial and anti-allergic reactions has become a challenge that current skin cleansing and care products need to address.

[0004] The oral cavity is the starting point of the digestive tract, and its main functions include chewing, sucking, swallowing, speech, sensation, facial expression, food intake, and respiration. The human oral cavity contains a large number of microorganisms that play important roles in digestion, metabolism, and maintaining tooth structure. Normally, the oral cavity's microecology maintains a relatively dynamic balance, resisting external pathogens, preventing their colonization, and protecting oral health. However, this defense is limited. Once the balance is disrupted, the excessive proliferation of oral pathogens can lead to oral and systemic diseases, and these diseases, in turn, exacerbate the imbalance of the oral microecology. Numerous studies have demonstrated that oral diseases (such as dental caries, periodontal disease, and oral mucosal diseases) are related to oral microecological imbalance. An increase in the proportion of Gram-negative bacteria and anaerobic bacteria leads to an increase in endotoxin production, which in turn triggers inflammatory responses. Furthermore, oral diseases caused by oral microecological dysbiosis may also increase the sensitivity of teeth to external stimuli, including mechanical, chemical, temperature, and osmotic pressure stimuli, resulting in transient, sharp pain or discomfort. Therefore, oral care needs to take into account both antibacterial and anti-allergic properties, which is also a new demand for oral care products.

[0005] Currently, commonly used antibacterial agents in daily skin cleansing and care products include: chlorometaxylenol, triclosan, chlorhexidine, phenoxyethanol, benzyl alcohol, imidazolidinyl urea, parabens, bropol, and Kathon. These chemical antibacterial agents are effective and relatively inexpensive, leading to their widespread use in products. However, recent studies have found that while these chemical antibacterial agents inhibit harmful bacteria on the skin, they also pose a strong risk of skin irritation, making them unacceptable to sensitive skin. Therefore, they do not meet consumer expectations. Similarly, existing oral care products struggle to simultaneously achieve both antibacterial and anti-allergic effects. Summary of the Invention

[0006] To address the above-mentioned technical problems, this invention provides a Tibetan medicine composition with antibacterial, antioxidant, and anti-allergic effects, its preparation method, and its applications. This Tibetan medicine composition has advantages such as antibacterial and anti-allergic properties, and is green and safe, overcoming the shortcomings of existing antibacterial products that cannot simultaneously provide anti-allergic effects. Furthermore, it also possesses antioxidant properties, reducing free radical damage to the skin or oral cavity, making it suitable for preparing skin and oral care products.

[0007] To achieve the above-mentioned objectives, the present invention adopts the following technical solution:

[0008] In a first aspect, the present invention provides a Tibetan medicine composition with antibacterial, antioxidant and anti-allergic effects, the raw materials of which include Phyllanthus emblica, Terminalia chebula and Terminalia chebula in a mass ratio of (6-8):(6-8):1.

[0009] Phyllanthus emblica is the dried, ripe fruit of the plant Phyllanthus emblica, belonging to the Euphorbiaceae family. It has a sweet and sour taste and is cool in nature. It possesses the effects of clearing heat and cooling the blood, promoting digestion and strengthening the stomach, and generating fluids and relieving cough. It can be used for blood heat and blood stasis, indigestion, abdominal distension, cough, dry throat and sore throat, thirst due to heat, and fever from colds. It is a medicinal and edible variety and an economic crop with wide applications in the food and pharmaceutical fields, and is also a traditional Tibetan medicine. Phyllanthus emblica mainly contains tannins and phenolic acids such as gallic acid, chebulic acid, corilagin, and chebulin, as well as flavonoids such as quercetin, catechin, gallocatechin, and rutin, and other terpenoids and sterols. Studies have shown that Phyllanthus emblica has pharmacological activities such as anti-oxidation, anti-aging, anti-diabetic and its complications, anti-hyperlipidemia, antibacterial, anti-inflammatory, and hepatoprotective effects.

[0010] Terminalia chebula, the dried, ripe fruit of the Terminalia chebula plant (family Combretaceae), is a traditional Tibetan medicine used to treat various diseases. It contains tannins and phenolic acids such as gallic acid, chebulic acid, and chebulic acid; triterpenoids such as ajangalamin, ajangalaminic acid, and fenestrated baicalein; and flavonoids such as rutin and quercetin. Pharmacological studies have shown that Terminalia chebula possesses anti-inflammatory, analgesic, antimicrobial, antioxidant, hypoglycemic, anticholinesterase, and anti-gastric ulcer effects.

[0011] Terminalia chebula, the dried, ripe fruit of the Combretaceae plant Terminalia chebula, has a bitter and astringent taste and is neutral in nature. It possesses properties of clearing heat and detoxifying, astringing and nourishing blood, and harmonizing other medicinal ingredients. It is used for various febrile diseases, diarrhea, jaundice, liver and gallbladder diseases, and post-illness weakness, and is a frequently used ingredient in Tibetan medicine compound prescriptions. Terminalia chebula mainly contains tannins and phenolic acids such as gallic acid, chebulic acid, ellagic acid, and chebulic acid. Pharmacological experiments have shown that Terminalia chebula has antioxidant, anti-diabetic, anti-atherosclerotic, and hepatoprotective effects.

[0012] The classic Tibetan medicine formula "Three Fruit Soup" consists of the above three medicinal herbs. Known in Tibetan medicine as Zhebu Song Tang, it has the effects of clearing heat, regulating qi and blood, and resolving scurvy. Clinically, it is often used to treat sore throat, cough, asthma, indigestion, anemia, liver dysfunction, and cardiovascular diseases. While there are many modern products developed using Three Fruit Soup, its application in skin and oral care products has not yet been explored. This invention unexpectedly discovered that "Three Fruit Soup" possesses antibacterial, anti-allergic, and antioxidant properties, and can be used as an active ingredient in skin and oral care products to exert corresponding effects.

[0013] Furthermore, regardless of whether the "Three Fruit Soup" is extracted with water or ethanol, its extract contains a complex composition, affecting the expected efficacy. If it is added as a raw material to skin or oral care products, a large amount is required to achieve the desired effect. This invention creatively and significantly adjusts the raw material ratio based on the traditional "Three Fruit Soup," unexpectedly discovering that Phyllanthus emblica, Terminalia chebula, and Terminalia chebula can produce a significant synergistic effect within the aforementioned mass ratio range, significantly enhancing antibacterial, antioxidant, and anti-allergic effects. This allows for the production of ideal antibacterial, antioxidant, and anti-allergic effects with a smaller dosage, making it more suitable for preparing skin and oral care products.

[0014] Preferably, the mass ratio of Phyllanthus emblica, Terminalia chebula and Terminalia chebula is (6.5-7.5):(6.5-7.5)1.

[0015] Preferably, the mass ratio of Phyllanthus emblica, Terminalia chebula, and Terminalia chebula is 7:7:1. This mass ratio produces the most ideal antibacterial, antioxidant, and anti-allergic effects.

[0016] Secondly, the present invention also provides a method for preparing the above-mentioned Tibetan medicine composition, specifically including the following steps:

[0017] S1. After crushing the raw material, soak it in water for extraction at a temperature of 40-80℃. Filter the resulting extract through a 0.22μm-10μm filter membrane and concentrate it to 0.1-1.0g crude drug / ml to obtain a concentrated solution.

[0018] S2. Adjust the pH of the concentrated solution obtained in S1 to 7.0-7.5 with 1-50 g / L sodium hydroxide solution, then add hydrogen peroxide solution to a hydrogen peroxide concentration of 0.1%-3% v / v, and stir at 20-80℃ for 10-60 min.

[0019] S3. Add 0.5% to 5% m / m activated carbon to the solution obtained in S2, stir and treat at a temperature of 20 to 80℃ for 1 to 5 hours, and filter to remove the activated carbon.

[0020] This preparation method uses water as the extraction solvent and treats the concentrated extract obtained from the water extraction with sodium hydroxide, hydrogen peroxide, and activated carbon. This significantly enhances the antibacterial, antioxidant, and anti-allergic effects of the resulting Tibetan medicine composition compared to preparation processes using other extraction solvents or without treatment with sodium hydroxide, hydrogen peroxide, and activated carbon. Consequently, it can be better applied to skin or oral products, exhibiting more significant efficacy. Furthermore, after filtering to remove activated carbon in S3, the resulting liquid is lighter in color (appearing as a yellowish-white powder after drying), which does not affect its effectiveness in skin or oral products.

[0021] Preferably, the pulverization is performed to a particle size ≤0.85mm.

[0022] Preferably, the mass of the water is 5 to 20 times the mass of the raw material.

[0023] Preferably, the extraction is performed 1 to 2 times, each time for 2 to 4 hours. After extraction, solid-liquid separation can be carried out by centrifugation, filtration, or other methods to obtain the extracts from each extraction, which are then combined.

[0024] Preferably, the concentration temperature is 50–80°C.

[0025] Preferably, the preparation method further includes drying the filtrate obtained by S3 filtration at a temperature not exceeding 80°C for transportation, storage and use.

[0026] Optionally, the drying process includes, but is not limited to, freeze drying, low-temperature drying, reduced-pressure low-temperature drying, and spray drying.

[0027] Thirdly, the present invention also provides the application of the above-mentioned Tibetan medicine composition or the Tibetan medicine composition prepared by the above-mentioned preparation method in the preparation of products with antibacterial, antioxidant and anti-allergic effects.

[0028] Preferably, the product is a skin product or an oral product.

[0029] Optionally, the skin products include, but are not limited to, topical skin medications and daily skin chemical products, such as cosmetics and skin cleansing products.

[0030] Optionally, the oral care products include, but are not limited to, dental medications and oral daily chemical products.

[0031] The beneficial effects of this invention are as follows:

[0032] This invention represents a deep development and application of the classic Tibetan medicine formula "Sanguo Tang" (Three Fruit Soup). Based on extensive experimental research, this invention has determined the formulation ratio and preparation method for external use of "Sanguo Tang." Compared to other formulation ratios and preparation methods, the Tibetan medicine composition provided by this invention exhibits superior antibacterial, antioxidant, and anti-allergic effects. It can be used in skin and oral products, providing a comprehensive range of antibacterial, antioxidant, and anti-allergic effects, thus expanding the application scope of Tibetan medicine compound formulas and enhancing their development value. Attached Figure Description

[0033] Figure 1 The results of the inhibition zone determination for Staphylococcus aureus in Example 1 are shown.

[0034] Figure 2 The results of the inhibition zone determination for Pseudomonas aeruginosa in Example 1 are shown.

[0035] Figure 3The results of the inhibition zone assay for Escherichia coli in Example 1 are shown.

[0036] Figure 4 The results of the inhibition zone assay for Porphyromonas gingivalis in Example 1 are shown.

[0037] Figure 5 The results of the inhibition zone determination for Candida albicans in Example 1 are shown.

[0038] Figure 6 The results of the DPPH removal rate test in Example 2;

[0039] Figure 7 The results of the ABTS clearance rate test in Example 2;

[0040] Figure 8 The results of the TPTZ reduction capability test are shown in Example 2. Detailed Implementation

[0041] To make the objectives, technical solutions, and advantages of this invention clearer, the invention will be further described in detail below with reference to specific embodiments. It should be understood that the specific embodiments described herein are merely illustrative and not intended to limit the invention. All other embodiments obtained by those skilled in the art based on the implementation methods of this invention without inventive effort fall within the protection scope of this invention.

[0042] Based on the characteristics of the skin and oral cavity and their physiological functions, people desire daily chemical products for oral care and skin cleansing that possess both good antibacterial and anti-allergic effects. However, existing antibacterial agents, while inhibiting harmful bacteria, are highly irritating and do not meet consumer needs. Traditional Chinese medicine is a treasure of my country's traditional culture, playing a crucial role in disease prevention and treatment for thousands of years, and possessing advantages such as fewer side effects and less likelihood of causing secondary diseases. Developing new daily chemical products using traditional Chinese medicine has become a new hot topic and trend in this field. This invention uses traditional Chinese medicine as raw materials and, through extensive research, has obtained a Tibetan medicine composition with antibacterial, antioxidant, and anti-allergic effects, as well as its preparation method. The raw materials of this Tibetan medicine composition include Phyllanthus emblica, Terminalia chebula, and Terminalia chebula in a mass ratio of (6-8):(6-8):1. Phyllanthus emblica, Terminalia chebula, and Terminalia chebula in the above ratio can produce a multi-component synergistic effect, not only producing good antibacterial effects but also significant anti-allergic effects.

[0043] This invention also provides a method for preparing the Tibetan medicine composition, specifically including the following steps:

[0044] S1. After crushing the raw material, soak it in water for extraction at a temperature of 40-80℃. Filter the resulting extract through a 0.22μm-10μm filter membrane and concentrate it to 0.1-1.0g crude drug / ml to obtain a concentrated solution.

[0045] S2. Adjust the pH of the concentrated solution obtained in S1 to 7.0-7.5 with 1-50 g / L sodium hydroxide solution, then add hydrogen peroxide solution to a hydrogen peroxide concentration of 0.1%-3% v / v, and stir at 20-80℃ for 10-60 min.

[0046] S3. Add 0.5% to 5% m / m activated carbon to the solution obtained in S2, stir and treat at a temperature of 20 to 80℃ for 1 to 5 hours, and filter to remove the activated carbon.

[0047] The above preparation method uses water as the extraction solvent and treats the concentrated extract obtained from water extraction with sodium hydroxide, hydrogen peroxide, and activated carbon. This significantly enhances the antibacterial, antioxidant, and anti-allergic effects of the Tibetan medicine composition, allowing it to be better applied to skin or oral products and exert more significant efficacy.

[0048] The present invention will be described below through specific embodiments.

[0049] Unless otherwise specified, all reagents, pharmaceuticals, and instruments used in the following examples were obtained commercially.

[0050] The Phyllanthus emblica, Terminalia chebula, and Terminalia chebula used in the following examples all conform to national or local standards.

[0051] Example 1

[0052] This invention provides a Tibetan medicine composition with antibacterial, antioxidant and anti-allergic effects, which is prepared from Phyllanthus emblica, Terminalia chebula and Terminalia chebula in a mass ratio of 7:7:1.

[0053] The preparation method is as follows:

[0054] The medicinal materials were weighed according to the mass ratio of Phyllanthus emblica: Terminalia chebula: Terminalia chebula = 7:7:1, mixed, pulverized, and passed through a 20-mesh sieve to obtain powdered raw materials. Deionized water was added at a mass-to-volume ratio of 1:10 for extraction at 60℃, twice, 2.5 hours each time. The extract was filtered through a 100-mesh filter cloth. The combined filtrates were further filtered through a 5μm pore size filter membrane to obtain a clear and transparent filtrate. The clear and transparent filtrate was concentrated under reduced pressure at 60℃ until the concentration of the medicinal solution was [missing information]. 0.5 g crude drug / ml was used to obtain a concentrated solution; the pH of the concentrated solution was adjusted to 7.0 with 10 g / L sodium hydroxide solution; 7.5% v / v hydrogen peroxide solution was added to the pH-adjusted solution until the hydrogen peroxide concentration was 1.5% v / v, and the solution was stirred at 50℃ for 35 min, then 2.5% (m / m) activated carbon was added, and the solution was stirred at 60℃ for 3.5 h, and the activated carbon was removed by filtration; the resulting solution was dried under reduced pressure at 60-80℃ to obtain the final product.

[0055] Example 2

[0056] This invention provides a Tibetan medicine composition with antibacterial, antioxidant and anti-allergic effects, which is prepared from Phyllanthus emblica, Terminalia chebula and Terminalia chebula in a mass ratio of 7:7:1.

[0057] The preparation method is as follows:

[0058] The medicinal materials were weighed according to the mass ratio of Phyllanthus emblica: Terminalia chebula: Terminalia chebula = 7:7:1, mixed, pulverized, and passed through a 10-mesh sieve to obtain powdered raw materials. Deionized water was added at a mass-to-volume ratio of 1:5 for extraction at 60℃, twice, for 2 hours each time. The extract was filtered through a 50-mesh filter cloth. The combined filtrates were further refined through a 0.22μm pore size filter membrane to obtain a clear and transparent filtrate. The clear and transparent filtrate was concentrated under reduced pressure at 50℃ until the concentration of the medicinal liquid was reached. The concentration was 0.1 g crude drug / ml to obtain a concentrated solution; the pH of the concentrated solution was adjusted to 7.2 with 1 g / L sodium hydroxide solution; 7.5% v / v hydrogen peroxide solution was added to the pH-adjusted solution until the hydrogen peroxide concentration was 0.1% v / v, and the solution was stirred at 20℃ for 60 min, then 0.5% (m / m) activated carbon was added, and the solution was stirred at 20℃ for 5 h, and the activated carbon was removed by filtration; the resulting solution was dried under reduced pressure at 60-80℃ to obtain the final product.

[0059] Example 3

[0060] This invention provides a Tibetan medicine composition with antibacterial, antioxidant and anti-allergic effects, which is prepared from Phyllanthus emblica, Terminalia chebula and Terminalia chebula in a mass ratio of 7:7:1.

[0061] The preparation method is as follows:

[0062] Weigh the medicinal materials according to the mass ratio of Phyllanthus emblica: Terminalia chebula: Terminalia chebula = 7:7:1, mix and pulverize, pass through a 20-mesh sieve to obtain powder raw material, add deionized water at a mass-to-volume ratio of 1:15 for soaking and extraction, extraction temperature is 50℃, extraction is performed once, extraction time is 4 hours, the extract is filtered through a 200-mesh filter cloth, the filtrates are combined and further filtered through a filter membrane with a pore size of 1μm to obtain a clear and transparent filtrate; concentrate the clear and transparent filtrate under reduced pressure at 70℃ until the medicinal solution concentration is reached. The concentration was 1.0 g crude drug / ml to obtain a concentrated solution; the pH of the concentrated solution was adjusted to 7.5 with 50 g / L sodium hydroxide solution; 7.5% v / v hydrogen peroxide solution was added to the pH-adjusted solution until the hydrogen peroxide concentration was 3% v / v, and the solution was stirred at 80℃ for 10 min, then 5% (m / m) activated carbon was added, and the solution was stirred at 80℃ for 1 h, and the activated carbon was removed by filtration; the resulting solution was dried under reduced pressure at 60-80℃ to obtain the final product.

[0063] Example 4

[0064] This invention provides a Tibetan medicine composition with antibacterial, antioxidant and anti-allergic effects, which is prepared from Phyllanthus emblica, Terminalia chebula and Terminalia chebula in a mass ratio of 7:7:1.

[0065] The preparation method is as follows:

[0066] The medicinal materials were weighed according to the mass ratio of Phyllanthus emblica: Terminalia chebula: Terminalia chebula = 7:7:1, mixed, pulverized, and passed through a 10-mesh sieve to obtain powdered raw materials. Deionized water was added at a mass-to-volume ratio of 1:20 for extraction at 70℃, twice, for 3 hours each time. The extract was filtered through a 300-mesh filter cloth, and the combined filtrates were further refined through a 3μm pore size filter membrane to obtain a clear and transparent filtrate. The clear and transparent filtrate was concentrated under reduced pressure at 50℃ until the concentration of the medicinal liquid was reached. The concentration was 0.3 g crude drug / ml to obtain a concentrated solution; the pH of the concentrated solution was adjusted to 7.3 with 25 g / L sodium hydroxide solution; 7.5% v / v hydrogen peroxide solution was added to the pH-adjusted solution to bring the hydrogen peroxide concentration to 1.5% v / v, and the solution was stirred at 70℃ for 20 min, then 2% (m / m) activated carbon was added, and the solution was stirred at 70℃ for 3 h, and the activated carbon was removed by filtration; the resulting solution was dried under reduced pressure at 60-80℃ to obtain the final product.

[0067] Example 5

[0068] This invention provides a Tibetan medicine composition with antibacterial, antioxidant and anti-allergic effects, which is prepared from Phyllanthus emblica, Terminalia chebula and Terminalia chebula in a mass ratio of 7:7:1.

[0069] The preparation method is as follows:

[0070] The medicinal materials were weighed according to the mass ratio of Phyllanthus emblica: Terminalia chebula: Terminalia chebula = 7:7:1, mixed, pulverized, and passed through a 20-mesh sieve to obtain powder raw materials. Deionized water was added at a mass-to-volume ratio of 1:12 for extraction at 65℃, twice, 2.5 hours each time. The extract was filtered through a 500-mesh filter cloth, and the combined filtrates were further refined through an 8μm pore size filter membrane to obtain a clear and transparent filtrate. The clear and transparent filtrate was concentrated under reduced pressure at 55℃ until the concentration of the medicinal liquid was reached. The concentration was 0.6 g crude drug / ml to obtain a concentrated solution; the pH of the concentrated solution was adjusted to 7.2 with 5 g / L sodium hydroxide solution; 7.5% v / v hydrogen peroxide solution was added to the pH-adjusted solution to bring the hydrogen peroxide concentration to 2.5% v / v, and the solution was stirred at 55℃ for 50 min, then 3.5% (m / m) activated carbon was added, and the solution was stirred at 76℃ for 3 h, and the activated carbon was removed by filtration; the resulting solution was dried under reduced pressure at 60-80℃ to obtain the final product.

[0071] Example 6

[0072] This invention provides a Tibetan medicine composition with antibacterial, antioxidant and anti-allergic effects, prepared from Phyllanthus emblica, Terminalia chebula and Terminalia chebula in a mass ratio of 6.5:7.5:1.

[0073] The preparation method is as follows:

[0074] Weigh the medicinal materials according to the mass ratio of Phyllanthus emblica: Terminalia chebula: Terminalia chebula = 6.5:7.5:1, mix and pulverize, pass through a 20-mesh sieve to obtain powder raw material, add deionized water at a mass-to-volume ratio of 1:15 for soaking and extraction, extraction temperature is 40℃, extraction is performed once, extraction time is 3.5 hours, the extract is filtered through a 500-mesh filter cloth, the filtrates are combined and further filtered through a 10μm pore size filter membrane to obtain a clear and transparent filtrate; the clear and transparent filtrate is concentrated under reduced pressure at 80℃, concentration... The concentration of the drug solution was reduced to 0.8 g crude drug / ml to obtain a concentrated solution. The pH of the concentrated solution was adjusted to 7.3 with 15 g / L sodium hydroxide solution. 7.5% v / v hydrogen peroxide solution was added to the pH-adjusted solution to bring the hydrogen peroxide concentration to 2% v / v. The solution was stirred at 35°C for 40 min, then 1.5% (m / m) activated carbon was added, and the solution was stirred at 55°C for 2.5 h. The activated carbon was removed by filtration. The resulting solution was freeze-dried to obtain the final product.

[0075] Example 7

[0076] This invention provides a Tibetan medicine composition with antibacterial, antioxidant and anti-allergic effects, which is prepared from Phyllanthus emblica, Terminalia chebula and Terminalia chebula in a mass ratio of 7.5:6.5:1.

[0077] The preparation method is as follows:

[0078] Weigh the medicinal materials according to the mass ratio of Phyllanthus emblica: Terminalia chebula: Terminalia chebula = 7.5:6.5:1, mix and pulverize, pass through a 20-mesh sieve to obtain powder raw material, add deionized water at a mass-to-volume ratio of 1:10 for soaking and extraction, extraction temperature is 80℃, extraction is performed twice, extraction time is 3 hours, the extract is filtered through a 500-mesh filter cloth, the filtrates are combined and further filtered through a 10μm pore size filter membrane to obtain a clear and transparent filtrate; the clear and transparent filtrate is concentrated under reduced pressure at 65℃. The concentration of the drug solution was adjusted to 0.7 g crude drug / ml to obtain a concentrated solution. The pH of the concentrated solution was adjusted to 7.2 with 10 g / L sodium hydroxide solution. 7.5% v / v hydrogen peroxide solution was added to the pH-adjusted solution until the hydrogen peroxide concentration was 1% v / v. The solution was stirred at 65℃ for 25 min, then 4% (m / m) activated carbon was added and stirred at 30℃ for 4 h. The activated carbon was removed by filtration. The resulting solution was dried at 60-80℃ to obtain the final product.

[0079] Example 8

[0080] This invention provides a Tibetan medicine composition with antibacterial, antioxidant, and anti-allergic effects, prepared from Phyllanthus emblica, Terminalia chebula, and Terminalia chebula in a mass ratio of 8:6:1. The preparation method is the same as in Example 1.

[0081] Example 9

[0082] This invention provides a Tibetan medicine composition with antibacterial, antioxidant, and anti-allergic effects, prepared from Phyllanthus emblica, Terminalia chebula, and Terminalia chebula in a mass ratio of 6:8:1. The preparation method is the same as in Example 1.

[0083] Comparative Example 1

[0084] This comparative example provides a product made from Phyllanthus emblica alone, and its preparation method is the same as in Example 1.

[0085] Comparative Example 2

[0086] This comparative example provides a product made from Terminalia chebula alone, and its preparation method is the same as in Example 1.

[0087] Comparative Example 3

[0088] This comparative example provides a product made from Terminalia chebula alone, and its preparation method is the same as in Example 1.

[0089] Comparative Example 4

[0090] This comparative example provides a Tibetan medicine composition prepared from Phyllanthus emblica and Terminalia chebula in a mass ratio of 1:1. The preparation method is the same as in Example 1.

[0091] Comparative Example 5

[0092] This comparative example provides a Tibetan medicine composition prepared from Phyllanthus emblica and Terminalia chebula in a mass ratio of 7:1. The preparation method is the same as in Example 1.

[0093] Comparative Example 6

[0094] This comparative example provides a Tibetan medicine composition prepared from Terminalia chebula and Terminalia chebula in a mass ratio of 7:1. The preparation method is the same as in Example 1.

[0095] Comparative Example 7

[0096] This comparative example provides a Tibetan medicine composition prepared from Phyllanthus emblica, Terminalia chebula, and Terminalia chebula in a mass ratio of 1:1:1 (a common ratio in foreign three-fruit soup products). The preparation method is the same as in Example 1.

[0097] Comparative Example 8

[0098] This comparative example provides a Tibetan medicine composition prepared from Phyllanthus emblica, Terminalia chebula, and Terminalia chebula in a mass ratio of 1.5:1:1.2 (the raw material ratio of the Three Fruit Decoction in the Tibetan Medicine Ministry Standard). The preparation method is the same as in Example 1.

[0099] Comparative Example 9

[0100] This comparative example provides a Tibetan medicine composition prepared from Phyllanthus emblica, Terminalia chebula, and Terminalia chebula in a mass ratio of 7:7:1. The preparation method is basically the same as in Example 1, except that sodium hydroxide solution was not used to adjust the pH.

[0101] Comparative Example 10

[0102] This comparative example provides a Tibetan medicine composition prepared from Phyllanthus emblica, Terminalia chebula, and Terminalia chebula in a mass ratio of 7:7:1. The preparation method is basically the same as in Example 1, except that hydrogen peroxide solution is not used.

[0103] Comparative Example 11

[0104] This comparative example provides a Tibetan medicine composition prepared from Phyllanthus emblica, Terminalia chebula, and Terminalia chebula in a mass ratio of 7:7:1. The preparation method is basically the same as in Example 1, except that activated carbon is not used.

[0105] Comparative Example 12

[0106] This comparative example provides a Tibetan medicine composition prepared from Phyllanthus emblica, Terminalia chebula, and Terminalia chebula in a mass ratio of 7:7:1. The preparation method is basically the same as in Example 1, except that 50% v / v ethanol is used instead of deionized water when extracting the powdered raw materials.

[0107] Example 1

[0108] This example demonstrates the antibacterial effect of the Tibetan medicine compositions prepared in Example 1 and Comparative Examples 1-12 of this invention.

[0109] 1. Experimental Methods

[0110] 1.1 Determination of inhibition zone

[0111] The diameter of the inhibition zone for each sample was determined using the agar perforation method. On a clean bench, sterilized tryptic soy agar (TSA) medium was cooled to approximately 60°C. 20 ml of the medium was poured into a petri dish and allowed to stand horizontally until completely solidified. An Oxford cup (6 mm inner diameter, 10 mm height) was placed on the surface of the agar medium, and a hole was gently punched under pressure. The medium was removed from the agar well using a sterile needle, and 0.1 ml of the test bacterial suspension was transferred to a plate and spread evenly using a spreader to prepare a bacterial plate. The prepared samples were then injected sequentially (80 μl per well). The petri dishes were then incubated at 37°C for 24 hours. The diameter of the inhibition zone was measured using calipers. A blank control group was prepared using 0.9% sterile sodium chloride solution.

[0112] 1.2 Method for determining the minimum inhibitory concentration (MIC)

[0113] The minimum inhibitory concentration (MIC) of each sample was determined using a two-fold serial dilution method. Solutions of different samples were serially diluted with TSB liquid medium to prepare solutions of specific concentrations. Indicator bacteria in the logarithmic growth phase were serially diluted with TSB liquid medium to 10⁻⁶. 5 For each CFU / mL bacterial suspension, 100 μL of the bacterial culture was added to wells 2-12 of a 96-well plate. An additional 100 μL of serial dilution was added to each well. The first well contained 100 μL of TSB liquid medium and 100 μL of bacterial suspension as a blank control. Each strain was tested three times. After mixing, the 96-well plate was incubated at 37°C for the specified time. Bacterial growth was observed. Each experiment was repeated three times, and the lowest concentration at which no colonies were observed in three trials was defined as the MIC.

[0114] 2. Experimental Results

[0115] The results of the inhibition zone assay are shown in Table 1 and Figures 1-5 The MIC measurement results are shown in Table 2.

[0116] Table 1. Results of inhibition zone determination (n=3, )

[0117]

[0118]

[0119] Note: Compared with the sample in Example 1, P<0.05, P<0.01.

[0120] Table 6. MIC Measurement Results of the Verification Experiment (n=3)

[0121]

[0122] As can be seen from the results in Tables 1 and 2, the Tibetan medicine composition provided in Example 1 of this invention has a significant inhibitory effect on skin pathogens (Staphylococcus aureus, Pseudomonas aeruginosa, Escherichia coli, Candida albicans) and oral pathogens (Porphyromonas gingivalis), and can be used in skin and oral hygiene products; moreover, the inhibition zone and MIC test results of the Tibetan medicine composition provided in Example 1 of this invention are significantly better than those of the comparative examples.

[0123] Example 2

[0124] This example demonstrates the antioxidant effects of the Tibetan medicine compositions prepared in Example 1 and Comparative Examples 1-12 of this invention.

[0125] 1. Experimental Methods

[0126] 1.1 Effect on DPPH free radical scavenging

[0127] A certain amount of sample was weighed and dissolved in DMSO (analytical grade), and sonicated until fully dissolved. 80 μL of sample solutions of different concentrations and 80 μL of 1 mmol / L DPPH solution were added sequentially to a 96-well plate. After reacting at room temperature in the dark for 30 min, the absorbance was measured at 517 nm and recorded as A1. Three parallel measurements were performed, and the average value was taken. Vitamin C was used as a positive control.

[0128] The calculation formula is as follows:

[0129]

[0130] A2: Replace DPPH solution with the same volume of DMSO; A0: Replace the sample with the same volume of DMSO, and the procedure is the same as A1.

[0131] 1.2 ABTS + Effects of free radical scavenging

[0132] Accurately measure 1 mL each of 7.4 mmol / L ABTS solution and 2.6 mmol / L K₂S₂O₈ solution, mix well, and store at 4°C in the dark for 12–16 h to obtain ABTS stock solution. Then, mix this stock solution with distilled water at a specific ratio (to achieve an absorbance A = 0.700 ± 0.02) to obtain ABTS working solution. Add 10 μL of different concentrations of the sample and 200 μL of ABTS working solution sequentially to a 96-well plate. Incubate for 6 min at room temperature in the dark, and measure the absorbance at 734 nm, recording it as A1. Vitamin C was used as a positive control.

[0133] The calculation formula is as follows:

[0134]

[0135] A0: Replace the sample with the same volume of distilled water, and follow the same procedure as A1.

[0136] 1.3 Effect on TPTZ Reduction Capacity

[0137] The working solution is prepared by mixing 1 mL of 10 mmol / L TPTZ solution, 1 mL of 20 mmol / L FeCl3(6H2O), and 10 mL of 0.3 mol / L acetate buffer (pH 3.6) in a 1:1:10 ratio. Different concentrations of analytes are prepared using DMSO reagent in a given sample volume. 150 μL of the working solution is added to a 96-well plate and incubated at 37°C for 8 min. The absorbance measured at 593 nm is recorded as A0. Then, 20 μL of analytes at different concentrations are added to the working solution in the 96-well plate and incubated at 37°C for 8 min. The absorbance measured at 593 nm is recorded as A. Trial measurements are performed in triplicate, and the average value is taken. All reagents are prepared fresh before use.

[0138] The calculation formula is as follows:

[0139]

[0140] A max This refers to the maximum value measured in a particular experiment. This value can come from the sample or from the positive control. Vitamin C was used as the positive control.

[0141] 2. Data Processing

[0142] Based on the linear fitting curves of DPPH and ABTS radical scavenging of different sample solutions, regression analysis using IBM SPSS Statistics 26 was used to calculate the amount of sample solution added when the original mass concentrations of DPPH and ABTS were reduced to 50% (steady state), which is the half-inhibitory amount (IC50). 50 According to half-inhibition level (IC50) 50 The size of the IC50 value can determine the strength of the sample solution's ability to scavenge free radicals. 50 The smaller the value, the stronger the ability to scavenge free radicals.

[0143] 3. Experimental Results

[0144] 3.1 In vitro antioxidant (DPPH, ABTS, TPTZ) performance analysis

[0145] The results are as follows Figures 6-8As shown, in scavenging DPPH free radicals, the sample of Example 1 achieved a DPPH scavenging rate of 40% at a concentration of 0.02 mg / mL, while the DPPH free radical scavenging rates in the comparative samples were significantly less than 40%. The scavenging rate reached its maximum at approximately 0.2 mg / mL, stabilizing at around 94%, which was higher than the positive control, Vitamin C. In scavenging ABTS free radicals, the inhibition rate of the sample of Example 1 at a concentration of approximately 0.2 mg / mL was stable at around 94%, higher than the positive control. In the TPTZ reduction reaction experiment, the FRAP value of the sample of Example 1 increased with increasing concentration, reaching a maximum of 4.68 mmol / L, significantly higher than other comparative samples. Other comparative samples also showed an increased antioxidant effect with increasing sample concentration when scavenging free radicals. However, the in vitro antioxidant effect of the comparative samples was not as good as that of the sample of Example 1.

[0146] 3.2 Analysis of in vitro antioxidant assay results

[0147] Table 3. Results of in vitro antioxidant activity assay for the samples (n=3, X±SD)

[0148]

[0149]

[0150] Note: Compared with the sample in Example 1, * P<0.05, ** P<0.01.

[0151] As shown in Table 3, the IC50 of the sample in Example 1 effectively scavenges DPPH and ABTS free radicals. 50 The values ​​were 0.026 mg / mL and 0.043 mg / mL, respectively, significantly lower than those of the comparative samples. However, in the TPTZ reducing power experiment, the maximum FRAP value of the Example 1 sample was 4.68 mmol / L, significantly higher than that of the comparative samples. Overall, the Example 1 sample significantly outperformed the other comparative samples, exhibiting better in vitro antioxidant effects. Within the set concentration range, the ability of the Example 1 sample to scavenge DPPH and ABTS free radicals showed a good concentration-effect relationship with the sample concentration. The ability of the Example 1 sample to scavenge DPPH and ABTS free radicals increased with increasing sample concentration. In conclusion, the antioxidant capacity of the Example 1 sample was significantly better than that of all comparative samples, which was statistically significant (P < 0.05 or 0.01).

[0152] Example 2

[0153] This example demonstrates the anti-allergic effect (evaluation of histamine inhibition effect) of the Tibetan medicine compositions prepared in Example 1 and Comparative Examples 1-12 of this invention.

[0154] 1. Experimental Methods

[0155] Mouse mast cell line MC / 9 was purchased from the American Type Organisms Collection (ATCC). MC / 9 cells were seeded in IMEM medium containing 10% FBS and 1% penicillin / streptomycin antibiotics. Cells were cultured at 37°C in a 5% CO2 incubator, and cells in the logarithmic growth phase were used for experiments. In the histamine production assay, cells were seeded in 24-well plates at a density of 1 × 10⁶ cells / well. 5 Cells were cultured at 100 μg / mL for 12 hours. Then, they were pre-treated and protected for 12 hours with samples from Example 1, Comparative Example 1, Comparative Example 2, Comparative Example 3, Comparative Example 4, Comparative Example 5, Comparative Example 6, Comparative Example 7, Comparative Example 8, Comparative Example 9, Comparative Example 10, Comparative Example 11, and Comparative Example 12 (100 μg / mL). MC / 9 cells were then induced for 24 hours with propylene glycol methyl ether acetate (PMA, 50 nM) and calcium iontophoresis agent A23187 (1 μM). After induction, the cell culture supernatant was collected, and histamine production was detected using an ELISA kit.

[0156] 2. Statistical methods

[0157] The experimental data were statistically analyzed using SPSS 22.0 software. The statistics were expressed as mean ± standard deviation (X ± SD), and one-way ANOVA was used for comparisons between groups.

[0158] 3. Experimental Results

[0159] Table 4. Results of experiments on inhibition of histamine production (n=3, X±SD)

[0160]

[0161] Note: Compared with the blank group, ## P<0.01. Compared with the PMA / A23187 group, & P<0.05, && P<0.01, compared with the sample in Example 1, * P<0.05, ** P<0.01.

[0162] Table 4 shows that PMA / A23187 stimulated histamine release in MC / 9 cells, with a significant increase in histamine content compared to the control group (P<0.01). After pretreatment of MC / 9 cells with each sample, histamine release decreased in all groups, but only the sample from Example 1 showed a highly significant difference compared to the PMA / A23187 group (P<0.01). Furthermore, the histamine release in the Example 1 sample group was significantly lower than that in each comparative sample group, showing a significant difference (P<0.05 or 0.01). When the body comes into contact with allergens, the large release of histamine is one of the protective reactions, but it can also lead to inflammatory reactions such as skin redness and swelling. Therefore, reducing histamine release is one of the main measures for the body to combat allergies. The above experimental results show that the Tibetan medicine composition provided in the embodiments of the present invention has a significant effect in reducing histamine release, and its anti-allergic effect is significantly better than that of the comparative samples. This indicates that the raw materials of the Tibetan medicine composition obtained by the present invention through scientific formulation have a significant synergistic effect, and the preparation method of the present invention plays an important role in improving the anti-allergic effect of the Tibetan medicine composition.

[0163] The above description is only a preferred embodiment of the present invention and is not intended to limit the present invention. Any modifications, equivalent substitutions or improvements made within the spirit and principles of the present invention should be included within the protection scope of the present invention.

Claims

1. A Tibetan medicine composition with antibacterial, antioxidant, and anti-allergic effects, characterized in that, The raw materials of the Tibetan medicine composition include Phyllanthus emblica, Terminalia chebula, and Terminalia chebula in a mass ratio of (6~8):(6~8):1; the preparation method specifically includes the following steps: S1. After crushing the raw material, soak it in water for extraction at a temperature of 40-80℃. Filter the resulting extract through a 0.22μm-10μm filter membrane and concentrate it to 0.1-1.0g crude drug / ml to obtain a concentrated solution. S2. Adjust the pH of the concentrated solution obtained in S1 to 7.0-7.5 with 1-50 g / L sodium hydroxide solution, then add hydrogen peroxide solution to the hydrogen peroxide concentration to 0.1%-3% v / v, and stir at 20-80℃ for 10-60 min. S3. Add 0.5%~5% m / m activated carbon to the solution obtained in S2, stir and treat at a temperature of 20~80℃ for 1~5 hours, and filter to remove the activated carbon.

2. The Tibetan medicine composition according to claim 1, characterized in that, The mass ratio of Phyllanthus emblica, Terminalia chebula and Terminalia chebula is (6.5~7.5):(6.5~7.5)1.

3. The Tibetan medicine composition according to claim 2, characterized in that, The mass ratio of Phyllanthus emblica, Terminalia chebula and Terminalia chebula is 7:7:

1.

4. The Tibetan medicine composition according to claim 1, characterized in that, The pulverization refers to pulverizing to a particle size ≤ 0.85 mm; and / or The mass of the water is 5 to 20 times the mass of the raw material; and / or The extraction is performed 1-2 times, each time for 2-4 hours; and / or The concentration temperature is 50~80℃.

5. The Tibetan medicine composition according to claim 1 or 4, characterized in that, The preparation method further includes drying the filtrate obtained by filtering S3 at a temperature not exceeding 80°C.

6. The use of the Tibetan medicine composition according to any one of claims 1 to 5 in the preparation of products with antibacterial, antioxidant and anti-allergic effects.

7. The application according to claim 6, characterized in that, The product in question is a skin product or an oral product.

8. The application according to claim 7, characterized in that, The skin products include topical skin medications and daily skin care products.

9. The application according to claim 7, characterized in that, The oral care products include, but are not limited to, dental medications and oral daily chemical products.

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