CN116019734A - Anti-dandruff application of dihydroartemisinin - Google Patents

Abstract

The present invention provides the anti-dandruff application of dihydroartemisinin, wherein the concentration of dihydroartemisinin is 0.01-10% by weight. The present invention also relates to the application of dihydroartemisinin in the preparation of personal care products with anti-dandruff effect, the personal care products are selected from: anti-dandruff shampoo, anti-dandruff scalp care milk, anti-dandruff scalp care essence, scalp care hair mask.

Description

Application of dihydroartemisinin in the treatment of dandruff

Technical Field

The invention relates to the fields of natural drug chemistry and cosmetics, and in particular to the anti-dandruff application of dihydroartemisinin.

Background Art

Dandruff is one of the main head problems facing consumers today. In 2007, the China Health Association released the "Survey Report on the Health Status of Chinese Residents' Scalp". The survey results showed that among Chinese consumers aged 20-55, 83% are suffering from dandruff to varying degrees, and 56% believe that the dandruff problem has affected their normal work and life. In 2009, China's first white paper on dandruff trouble index pointed out that among the tens of thousands of valid questionnaires in 16 cities surveyed, about 67%, or two-thirds of the respondents, had dandruff problems, and nearly 95% of the respondents had recurrences, and the proportion of patients with dandruff recurrence within 5 days was as high as 90%. In 2014, the survey results of "Chinese Scalp Health Status" released by the Chinese Medical Doctor Association showed that 9 out of 10 Chinese people are in a "scalp overwork state"; more than 60% of Chinese people are troubled by the problem of "not being able to suppress dandruff for a long time". The 2021 National Scalp Health White Paper pointed out that dandruff ranks first among consumers' hair/scalp problems with 43%. It can be seen that dandruff problems are prevalent in consumers' daily lives and are one of the main scalp health problems that need to be solved urgently. It also has broad market potential.

Dandruff is caused by the disruption of the balance between epidermal cell shedding and cell regeneration, which causes keratinized cells to aggregate into visible slough. Under normal circumstances, scalp keratin will naturally metabolize and shed periodically, which is generally invisible to the naked eye and can be solved by washing your hair frequently. When the scalp microecology is unbalanced, it may lead to the proliferation of lipophilic Malassezia, which in turn may cause an imbalance in the secretion of scalp sebum, produce a large amount of unsaturated free fatty acids, irritate the scalp, and cause dandruff. In addition, when the scalp barrier function is unbalanced, it may lead to changes in the pH value and water content of the scalp, which may cause the proliferation of Malassezia, cause an imbalance in sebum secretion, and lead to itchy scalp, dandruff and other phenomena (Chen Dongzhi, Xu Mingli, Huo Yanli, Overview of Research on Mild Antidandruff Agent Piroctone Ethanolamine Salt [J], Daily Chemicals Science, 2021, 44(5): 40-44). A large number of studies have shown that the abnormal proliferation of Malassezia is closely related to the occurrence and severity of dandruff (Wang Ran, Lai Wei, Zhang Yuqing, et al., The role of Malassezia quantitative culture method in the evaluation of anti-dandruff efficacy [C], Proceedings of the Third Central and Southern Dermatology and Venereology Academic Conference, 2006: 245-247; Liu Di, Development of plant-based anti-dandruff agents and research on the mechanism of inhibiting Malassezia [D], Guangdong: Guangdong University of Technology, 2020). Therefore, inhibiting Malassezia has become one of the main targets of daily anti-dandruff products.

At present, common antidandruff agents on the market include ZPT (zinc pyrithione), climbazole (climbazole), HP100 (hexymetidine di(hydroxyethyl sulfonate) salt), Runanti HS (bispyrithione), salicylic acid, SL-900 (undecylenamide MEA disodium sulfosuccinate), etc. However, salicylic acid only has a bactericidal effect, and its antidandruff effect is poor, and it is highly irritating and can easily damage the scalp; zinc pyrithione has a good antibacterial effect, but its own toxicity and safety of use cannot be ignored; climbazole has an inhibitory effect on Bacillus ovale or Pityriasis ovale, Candida albicans, and Trichophyton, which cause dandruff, but it is harmful to aquatic organisms and can pollute the aquatic environment. In November 2021, the European Commission officially released the Regulation (EU) 2021/1902 regulatory bulletin, listing the anti-dandruff agent zinc pyrithione (ZPT) in the banned list because it is classified as reproductive toxicity 1B by GHS. The above-mentioned anti-dandruff agents have disadvantages such as high toxicity, high concentration, poor solubility or poor anti-dandruff effect, and their application in hair care products has varying degrees of limitations. At present, the temporarily safe anti-dandruff ingredient is piroctone ethanolamine (Octopirox, OCT), which also achieves the purpose of dandruff removal by killing fungi and is effective at low concentrations (0.1%-0.75%).

Therefore, the development of new natural sources of safe and effective anti-dandruff agents that inhibit Malassezia has very important practical significance and broad market application value.

Artemisia annua L. is a Chinese specialty plant. It is the dried aerial part of Artemisia annua L., a plant of the Asteraceae family. It is cold in nature and bitter in taste. It has the effects of clearing away heat from the body, removing bone steaming, relieving summer heat, stopping malaria, and relieving jaundice. Since Professor Tu Youyou of the China Academy of Chinese Medical Sciences first discovered artemisinin from Artemisia annua in the 1970s and applied it to the treatment of human antimalarial diseases, it has gradually become known to the world. Since the beginning of the 21st century, especially after Tu Youyou won the Nobel Prize, research on artemisinin and its derivatives and the plant Artemisia annua has become increasingly popular, and its research and application fields have gradually expanded from the field of oral medicine to the field of skin topical use.

Jiang Lan et al. reviewed the research status of artemisinin and its derivatives in the treatment of skin diseases in "Research Overview of Artemisia annua and Its Derivatives in the Treatment of Skin Diseases", Chinese Journal of Traditional Chinese Medicine, 2018, 33(1): 232-235. The results showed that artemisinin and its derivatives have anti-inflammatory, anti-allergic and antibacterial effects and can be used to treat dermatitis, skin allergies, acne, infectious diseases and other diseases.

Xue Xue et al., "Dihydroartemisinin reduces atopic dermatitis in mice by regulating mast cell infiltration", Journal of Southern Medical University, 2020, 40(10): 1480-1487. The study found that skin application of dihydroartemisinin has a certain therapeutic effect on the DNCB-induced AD mouse model. The skin lesion scores, scratching behavior, histopathological changes, and mast cell infiltration of mice were significantly improved, and the therapeutic effect of the 125 mg/kg DHA treatment group on AD mice was better than that of the commonly used clinical drug dexamethasone. It may be through inhibiting mast cell infiltration, regulating the immune function of the lesion site, and improving AD symptoms.

Wei Qiang et al., "Dihydroartemisinin improves psoriasis-like skin inflammation in mice by inhibiting the proliferation of keratinocytes and the production of proinflammatory factors", Chinese Journal of Immunology, 2020(5): 543-548. The study found that dihydroartemisinin can improve imiquimod-induced psoriasis-like skin inflammation in mice. The possible mechanism is to inhibit the excessive proliferation of keratinocytes and the cytokines they secrete through the MAPK/NF-κB signaling pathway.

Chinese patent CN109939105A discloses the use of artemisinin and its derivatives in the preparation of medicines and cosmetics for improving the barrier function of the skin stratum corneum and preventing and treating skin inflammation.

However, to date, there has been no report on the inhibitory effect of dihydroartemisinin on Malassezia, nor is there any precedent for the use of dihydroartemisinin as an anti-dandruff agent in daily chemical products.

Summary of the invention

In one aspect, the present invention provides the use of dihydroartemisinin for removing dandruff, wherein the dihydroartemisinin is used at a concentration of 0.01-10% by weight. In a preferred embodiment, the dihydroartemisinin is used at a concentration of 0.1-5% by weight.

In a preferred embodiment, the anti-dandruff effect is achieved by inhibiting Malassezia. In a preferred embodiment, the minimum inhibitory concentration of dihydroartemisinin against Malassezia is 1.0 mg/mL.

On the other hand, the present invention relates to the use of dihydroartemisinin in the preparation of personal care products with anti-dandruff effects, wherein the dihydroartemisinin is used at a concentration of 0.01-10% by weight. In a preferred embodiment, the dihydroartemisinin is used at a concentration of 0.1-5% by weight.

In a preferred embodiment, the personal care product is selected from: anti-dandruff shampoo, anti-dandruff scalp care lotion, anti-dandruff scalp care essence, and scalp care hair mask.

BRIEF DESCRIPTION OF THE DRAWINGS

Figure 1 shows the results of the inhibition zone test of Example 1, wherein A represents the result of 10 mg/ml Artemisia annua extract, B represents the result of 100 mM artemisinin, C represents the result of 5 mM dihydroartemisinin, and D represents the result of 10 mM dihydroartemisinin.

DETAILED DESCRIPTION

The present invention unexpectedly found that dihydroartemisinin has an excellent effect of inhibiting Malassezia and can effectively remove dandruff. Moreover, dihydroartemisinin can also be used as an effective additive to prepare personal care products, especially hair products, to achieve the effect of removing dandruff.

The main purpose of the present invention is to provide a natural, safe and efficient Malassezia-inhibiting ingredient, which has the effect of removing dandruff and can be used as an anti-dandruff agent in daily chemical products used on the head, such as shampoo, hair conditioner, hair mask, etc.

In order to provide a more concise description, some quantitative expressions given herein are not modified by the term "about". It should be understood that, whether or not the term "about" is explicitly used, each quantity given herein is intended to refer to the actual given value, and is also intended to refer to the approximate values of these given values that can be reasonably inferred by ordinary technicians in the field, including the approximate values of these given values caused by experimental and/or measurement conditions.

To provide a more concise description, some quantitative expressions herein are described as a range of about X amount to about Y amount. It should be understood that when describing a range, the range is not limited to the upper and lower limits described, but should include the entire range of about X amount to about Y amount or any amount therebetween.

Dihydroartemisinin

The Malassezia inhibiting component of the present invention is dihydroartemisinin, C ₁₅ H ₂₄ O ₅ , CAS No. 71939-50-9, which is an important derivative of artemisinin. Dihydroartemisinin is prepared by reducing artemisinin from Artemisia annua L. as a precursor with sodium borohydride and other reducing agents, and has better antimalarial efficacy than artemisinin, and is a new generation of antimalarial therapeutic drug, which is widely used in the medical field. However, it is rarely used in the field of daily chemicals, especially as an anti-dandruff agent, and no application reports have been found.

The specific results of dihydroartemisinin are as follows:

The present application discovers for the first time that dihydroartemisinin can inhibit Malassezia, and therefore can be used as an effective additive with anti-dandruff function in personal care products (especially hair products) to achieve effective dandruff removal.

First, the present invention adopts the inhibition ring method to study and compare the inhibitory effects of dihydroartemisinin, artemisinin, and artemisia annua extract on Malassezia. The results show that artemisinin and artemisia annua extract have no inhibitory effect on Malassezia at the corresponding concentrations, while dihydroartemisinin has obvious inhibition rings at 5mM and 10mM concentrations, and the inhibition ring of 10mM is greater than that of the positive control OCT, indicating that dihydroartemisinin has a good inhibitory effect on Malassezia.

Secondly, the present invention uses a minimum inhibitory concentration (MIC) test to further study the minimum inhibitory concentration of dihydroartemisinin against Malassezia. The results show that the minimum inhibitory concentration MIC of dihydroartemisinin against Malassezia is 3.5mM (mass concentration 1.0mg/mL).

In some embodiments, dihydroartemisinin is used at a concentration of 0.01-10% by weight, preferably 0.01-5% by weight, more preferably 0.1-5% by weight. In some embodiments, dihydroartemisinin is used at a concentration of 1-5% by weight.

In some embodiments, the weight percentage of dihydroartemisinin in the personal care product can be 0.0001%-20% (w/w), preferably 0.001%-10% (w/w), more preferably 0.001%-5% (w/w), and most preferably 0.01%-5% (w/w).

Personal care products containing dihydroartemisinin

Dihydroartemisinin can be used as an effective additive in personal care products. The main purpose of the present invention is to provide a natural, safe and efficient Malassezia-inhibiting ingredient with an anti-dandruff function, which can be used as an anti-dandruff agent in daily chemical products used on the head, such as shampoo, hair conditioner, hair mask, etc.

Optionally, the personal care product may also contain from about 0.01% to about 1.0%, preferably from about 0.01% to about 0.5%, more preferably from about 0.01% to about 0.2% of at least one conditioning agent, based on the total weight. Examples of suitable cationic conditioning agents include, but are not limited to, cationic cellulose derivatives; cationic guar gum derivatives; diallyldimethylammonium chloride and mixtures thereof.

The cationic cellulose derivative is preferably a polymeric quaternary ammonium salt derived from the reaction of hydroxyethyl cellulose and a trimethylammonium substituted epoxide. The material known as Polyquaternium-10, commercially available as "Polymer JR-400" from Amerchol Corporation of Edison, New Jersey, is particularly useful in this regard.

The cationic guar gum derivative is preferably guar hydroxypropyltrimonium chloride, commercially available from Rhodia of Cranbury, New Jersey under the trade name "Jaguar-17".

Other cationic conditioning polymers are those derived from the monomer diallyldimethylammonium chloride. The homopolymer of this monomer is Polyquaternium-6, commercially available from Allied Colloids of Suffolk, Virginia, under the trade name "Salcare SC30". The copolymer of diallyldimethylammonium chloride and acrylamide is known as Polyquaternium-7, also available from Allied Colloids under the trade name "Salcare SC10".

In one embodiment, the conditioning agent portion comprises from about 0.01% to about 0.5%, such as from about 0.01% to about 0.2%, of a cationic guar gum derivative and from about 0.01% to about 0.5%, such as from about 0.01% to about 0.2%, of a homopolymer or copolymer of diallyldimethylammonium chloride, based on the total weight of the product.

Personal care products may also include one or more optional ingredients, which include but are not limited to pearlescent agents or opacifiers, thickeners, auxiliary conditioning agents, moisturizers, chelating agents, and additives that can improve their appearance, feel and fragrance, such as colorants, fragrances, preservatives, pH adjusters, etc. For example, the pH of a shampoo product is preferably maintained at about 5 to about 7.5, more preferably about 5.5 to about 7.0.

Commercially available pearlescent agents or opacifiers that can suspend water-insoluble additives such as silicones and/or make the user aware that the product is a conditioning shampoo are suitable for use in the present invention. The amount of pearlescent agent or opacifier used is about 1% to about 10%, preferably about 1.5% to about 7%, and more preferably about 2% to about 5%, based on the total weight of the product. Examples of suitable pearlescent or opacifying agents include, but are not limited to, mono- or diesters of (a) fatty acids having from about 16 to about 22 carbon atoms and (b) ethylene glycol or propylene glycol; mono- or diesters of (a) fatty acids having from about 16 to about 22 carbon atoms and (b) polyalkylene glycols of the formula: HO-(JO) _a -H wherein J is an alkylene group having from about 2 to about 3 carbon atoms and a is 2 or 3; fatty alcohols having from about 16 to about 22 carbon atoms; fatty acid esters of the formula: _KCOOCH2L wherein K and L each have from about 15 to about 21 carbon atoms; inorganic solids insoluble in shampoo products, and mixtures thereof.

In a preferred embodiment, the pearlescent or opacifying agent is added to the shampoo product in the form of a preformed, stable aqueous dispersion, such as is commercially available from Henkel Corporation of Hoboken, New Jersey under the trade name "Euperlan PK-3000". This material is a combination of glycol distearate (a diester of ethylene glycol and stearic acid), laureth- ₄ ( _CH3 ( _CH2 ) _10CH2 ( _OCH2CH2 ) _4OH ), and cocamidopropyl betaine, preferably in an amount of about 25 to about 30 weight percent, about 3 to about ₁₅ weight percent, and about 20 to about 25 weight percent, respectively.

Commercially available thickeners that provide a suitable viscosity for conditioning shampoos are suitable for use herein. If used, the thickener should be present in the shampoo product in an amount sufficient to increase the Brookfield viscosity of the product to about 500 to about 10,000 centipoise. Examples of suitable thickeners include, but are not limited to: 1) _{polyethylene glycols of the formula: HO-(CH2CH2O ) zH} and 2) mono- or diesters of fatty acids containing from about 16 to about 22 carbon atoms, wherein z is an integer from about 3 to about 200; ethoxylated polyol fatty acid esters; ethoxylated derivatives of mono- or diesters of fatty acids and glycerol; hydroxyalkyl celluloses; alkyl celluloses; hydroxyalkyl alkyl celluloses and mixtures thereof. Preferred thickeners include polyethylene glycol esters, more preferably PEG-150 distearate, which is commercially available from Stepan Company of Northfield, Illinois or from Comiel, SpA of Bologna, Italy, under the trade name "PEG 6000DS".

Commercially available auxiliary conditioning agents such as volatile silicones that can impart other properties to the hair, such as shine, are suitable for use in the present invention. If a volatile silicone conditioning agent is used, it is preferably one that has a boiling point at atmospheric pressure of less than about 220°C. The amount of volatile silicone conditioning agent is from about 0% to about 3%, such as from about 0.25% to about 2.5% or from about 0.5% to about 1.0%, based on the total weight of the product. Examples of suitable volatile silicones include, but are not limited to: polydimethylsiloxane, polydimethylcyclosiloxane, hexamethyldisiloxane, cyclomethicone fluids such as polydimethylcyclosiloxane commercially available from Dow Corning Corporation of Midland, Michigan, under the trade name "DC-345", and mixtures thereof.

Commercially available humectants that provide moisturizing and conditioning properties to personal care products are suitable for use in the present invention. The amount of humectant is from about 0% to about 10%, preferably from about 0.5% to about 5%, and more preferably from about 0.5% to about 3%, based on the total weight of the product. Examples of suitable humectants include, but are not limited to: 1) a water-soluble liquid polyol selected from glycerol, propylene glycol, hexylene glycol, butylene glycol, dipropylene glycol, and mixtures thereof; 2) a polyalkylene glycol of the formula: HO-(R"O) _b -H, wherein R" is an alkylene group having from about 2 to about 3 carbon atoms and b is an integer from about 2 to about 10; 3) a polyethylene glycol ether of methyl glucose of the formula: _CH3 - _C6H10O5- ( _{OCH2CH2 ) c} - _OH , wherein _c is an integer from about 5 to about 25; 4) urea; and 5) mixtures thereof, glycerol being the preferred humectant.

Examples of suitable chelating agents include those that are capable of protecting and preserving the products of the present invention. A preferred chelating agent is EDTA, more preferably tetrasodium EDTA available from Dow Chemical Company of Midland, Michigan under the trade name "Versene 100XL", in an amount of about 0% to about 0.5%, preferably about 0.05% to about 0.25%, based on the total weight of the product.

Suitable preservatives include Quaternium-15 available as "Dowicil 200" from Dow Chemical Corporation of Midland, Michigan, in amounts ranging from about 0% to about 0.2%, preferably from about 0.05% to about 0.10%, based on the total weight of the product.

In some embodiments of the present invention, the amount of dihydroartemisinin in the personal care product is 0.001%-20% (w/w), preferably 0.01%-20% (w/w), more preferably 0.01%-10% (w/w), and most preferably 0.1%-5% (w/w).

Example

The present invention is further described below in conjunction with specific examples. It is necessary to point out that the examples are only used to further illustrate the present invention and cannot be construed as limiting the scope of the present invention. Those skilled in the art can make some non-essential improvements and adjustments based on the content of the present invention described above. The test methods in the following examples that do not specify specific conditions are usually based on conventional conditions or the conditions recommended by the manufacturer. Unless otherwise stated, all percentages and parts are by weight.

Artemisinin (purity> 90%, from Artemisia annua, provided by the Institute of Chinese Materia Medica, China Academy of Chinese Medical Sciences);

Dihydroartemisinin (purity>90%, prepared with artemisinin as precursor, provided by the Institute of Chinese Materia Medica, China Academy of Chinese Medical Sciences).

Example 1: Inhibition ring experiment

Sample preparation :

Dihydroartemisinin 1mM: Take dihydroartemisinin, dissolve it in DMSO and prepare it to 1mM (0.284mg/ml).

Dihydroartemisinin 5mM: Take dihydroartemisinin, dissolve it in DMSO and prepare it to 5mM (1.42mg/ml).

Dihydroartemisinin 10 mM: Dissolve dihydroartemisinin in DMSO to prepare 10 mM (2.84 mg/ml).

Artemisinin 1mM: Take artemisinin, dissolve it in DMSO and prepare it to 1mM (0.282mg/ml).

Artemisinin 5mM: Take artemisinin, dissolve it in DMSO and prepare it to 5mM (1.41mg/ml).

Artemisinin 10mM: Take artemisinin, dissolve it in DMSO and prepare it to 10mM (2.82mg/ml).

Artemisinin 100 mM: Dissolve artemisinin in DMSO to prepare 100 mM (28.23 mg/ml).

Artemisia annua extract (10 mg/ml): Weigh Artemisia annua (Beijing-Anhui Medicinal Piece Factory, produced in Anhui), extract twice with 15 and 10 times deionized water, each time for 1 h, filter with sieve (Shanghai Yichang Instrument Screen Factory, 20 mesh), concentrate the filtrate to 3 times the amount of crude drug, add a certain amount of 95% ethanol for alcohol precipitation, filter with filter paper (Whatman Company), take the filtrate, concentrate and recover until there is no alcohol taste, prepare an aqueous solution of 10 mg/ml Artemisia annua extract, and set aside.

Positive control OCT (5 mg/ml, equivalent to): Take 0.5 g of piroctone olamine salt (OCT), dissolve it in 12% AES aqueous solution, and quantify it to 100 ml.

Solvent control 1 (DMSO): commercially available dimethyl sulfoxide, chemically pure;

Solvent control 2 (deionized water): homemade in the laboratory, conforming to the third-grade water specified in GB/T 6682;

Solvent control 3 (12% AES aqueous solution): 12 g of AES (trade name Texapon N70, supplier BASF) was weighed, dissolved in deionized water, and quantified to 100 g.

Testing process: Oxford cup double-layer plate method

Experimental bacteria: Malassezia furfur ATCC44344 (Shanghai Branch of Guangzhou Huayuexing Instrument Co., Ltd.).

The culture medium was homemade with the following ratios: 1% glucose, 1% peptone, 0.2% yeast extract, 0.8% ox bile salt, 2% olive oil, 0.5% Tween 60, 1% glycerol, 0.05% monostearate, 1.5% agar (all reagents used were purchased from Sinopharm Chemical Reagents), and the balance was water.

1. Preparation of Oxford cup double-layer plate:

(1) Pour 10 ml of melted sterile culture medium into a sterile culture dish and let it stand until it solidifies.

(2) Place several sterilized Oxford cups vertically and arrange them neatly in a certain order. The distance between the centers of the Oxford cups should be more than 25 mm, and the distance from the edge of the plate should be more than 15 mm. At least 3 or more Oxford cups should be placed on each plate (1 or more with the sample to be tested, 1 with sterile saline as a negative control, and 1 with 0.5% OCT solution as a positive control).

(3) Preparation of bacterial solution: Rinse the bacterial moss with sterile saline solution and place the rinse solution into a sterile test tube; adjust the concentration of the bacterial solution to 10 ^7-8 CFU/ml.

(4) Take an appropriate amount of bacterial solution and add it to a sterile culture medium cooled to 50°C and mix well to prepare a culture medium with a final concentration of 10 ⁵ to 10 ⁶ CFU/ml of indicator bacterial solution.

(5) Pipette 15 ml of the above culture medium containing bacterial solution into the culture dish, and avoid the formation of bubbles. After it is fully solidified, use sterile tweezers to remove the Oxford cup.

2. Sample addition: Use a disposable pipette to absorb 100-200ul of sample and add it to the round hole of the test plate with sterile operation. Cover the plate and place it in a constant temperature incubator for 48 hours.

3. Measurement of the diameter of the inhibition ring: Take out each plate after culture, measure the diameter of the inhibition ring (including the pore size) and record it.

4. Determination of antibacterial effect:

(1) If the diameter of the antibacterial ring is greater than 9 mm, it is judged to have antibacterial effect.

(2) No inhibition zone should be produced in the negative control group, otherwise the experiment will be invalid.

Test results:

Table 1

The results showed that the diameter of the inhibition zone of 10 mg/ml Artemisia annua extract was 7 mm, which was equivalent to the solvent blank (deionized water), indicating that the Artemisia annua extract had no inhibitory effect on Malassezia.

When artemisinin was at a concentration of 1-100 mM, the diameter of the inhibition zone was 7 mm, which was equivalent to the solvent blank (DMSO), indicating that artemisinin had no inhibitory effect on Malassezia.

When dihydroartemisinin was at a concentration of 1mM, the diameter of the inhibition zone was 7mm, which was equivalent to the solvent blank (DMSO), indicating that dihydroartemisinin had no inhibitory effect on Malassezia at this concentration; but at 5mM and 10mM concentrations, the diameters of the inhibition zones were 18.95±0.33mm and 23.63±0.35mm, respectively, which were significantly higher than the 7mm of the solvent blank (DMSO), and the 10mM inhibition zone was larger than the positive (OCT) inhibition zone of 21.59±0.24mm, indicating that dihydroartemisinin had a good inhibitory effect on Malassezia. In addition, the inhibition zone of dihydroartemisinin at a concentration of 10mM was larger than that of 5mM, indicating that the inhibitory effect of dihydroartemisinin on Malassezia increased with increasing concentration, with a dose-dependent effect.

Example 2: Minimum Inhibitory Concentration (MIC) Determination

Sample preparation :

Solvent control (10% AES aqueous solution): 10 g of AES (trade name Texapon N70, supplier BASF) was weighed, dissolved in deionized water, and quantified to 100 g.

Positive control OCT (10 mM, ie, 2.98 mg/mL): piroctone olamine salt (OCT, molecular weight: 298.42) was dissolved in 10% AES aqueous solution and quantified to 10 mM.

Dihydroartemisinin (10 mM, ie, 2.84 mg/mL): dihydroartemisinin (molecular weight: 284.35) was dissolved in 10% AES aqueous solution and quantified to 10 mM.

Testing process :

Experimental reagents and instruments:

1. Experimental strain: Malassezia furfur ATCC44344. (Guangzhou Huayuexing Instrument Co., Ltd. Shanghai Branch)

2. Micropipettes, test tubes, vortex mixers, culture media, etc.

The culture medium was homemade with the following ratios: 1% glucose, 1% peptone, 0.2% yeast extract, 0.8% ox bile salt, 2% olive oil, 0.5% Tween 60, 1% glycerol, 0.05% monostearate, 1.5% agar (all reagents used were purchased from Sinopharm Chemical Reagents), and the balance was water.

Specific method:

(1) Prepare a double concentration of ox bile salt liquid culture medium, sterilize it with pressure steam at 121°C for 15 min, and place it at room temperature for later use. This culture medium will be used to dilute the antibacterial (inhibitory) solution/suspension.

(2) Preparation of test solutions of different concentrations: dilute the antibacterial (inhibitory) solution/suspension with sterile deionized water in equal-fold series to produce test solutions of different concentrations (or dilute the antibacterial (inhibitory) solution/suspension with sterile deionized water to produce test solutions of corresponding concentrations) and place at room temperature for later use.

(3) Preparation of ox bile salt liquid culture medium containing antibacterial (inhibitory) agents: Take 2.5 ml of the test solution of each dilution and add it to a test tube containing 2.5 ml of double-concentration ox bile salt liquid culture medium and mix well.

(4) Preparation of bacterial suspension: Take the bacterial suspension of the test bacteria and adjust the concentration of the bacterial suspension to 10 ⁷ to 10 ⁸ cfu/ml with sterile saline.

(5) Take 0.1 ml of bacterial suspension containing 10 ⁷ to 10 ⁸ cfu/ml and inoculate it into a test tube containing ox bile salt liquid culture medium containing antibacterial (inhibitory) agents as the test group sample.

(6) Inoculate the bacterial suspension into a test tube containing a single-fold ox bile salt liquid culture medium without antibacterial (inhibitory) agents in the same manner as the positive control sample.

(7) Take two test tubes containing only single-dose ox bile salt liquid culture medium as negative control group samples.

(8) After placing the test group samples, positive control group samples, and negative control group samples in a 37°C shaker for 48 h, take 1 ml of the liquid culture medium in each test tube and transfer it to a plate. Pour the culture medium into a 37°C incubator and culture. Observe the results after 48-72 h.

(9) During the test, live bacterial culture counts should be performed on the test bacterial suspension at the same time, and the effective concentration should be 5×10 ⁵ cfu/ml to 5×10 ⁶ cfu/ml.

Determination of antibacterial effect:

The lowest concentration of antibacterial solution at which the growth of the colony is completely inhibited is the MIC of the sample against the tested bacteria. The growth of a single colony can be ignored.

Test results :

Analysis of results: The inhibition zone experiment showed that dihydroartemisinin had an inhibitory effect on Malassezia at concentrations of 5mM and 10mM, in order to further understand the minimum inhibitory concentration of dihydroartemisinin on Malassezia. The results showed that the minimum inhibitory concentration of the solvent control (10% AES solution) was >50%, indicating that the solvent control (10% AES solution) had no inhibitory effect after being half-diluted with distilled water, and had no interference with the antibacterial test of the sample. The minimum inhibitory concentration MIC of dihydroartemisinin on Malassezia was 3.5mM, which was close to the inhibitory effect of the positive control OCT. It has a very good inhibitory effect on Malassezia and has great market application potential.

Dihydroartemisinin can also be used as an effective additive in the preparation of personal care products, and the personal care products are preferably anti-dandruff shampoo, anti-dandruff scalp care lotion, anti-dandruff scalp care essence, and scalp care hair mask. The weight percentage of dihydroartemisinin in the personal care product can be 0.0001%-20% (w/w), preferably 0.001%-10% (w/w), more preferably 0.001%-5% (w/w), and most preferably 0.01%-5% (w/w).

The following are examples of specific applications of dihydroartemisinin in hair products, and the formulations and preparation methods of these dosage forms. In the following tables, "-" means no addition.

Application Example 1: Preparation of anti-dandruff shampoo

Application Example 2: Preparation of Anti-Dandruff Scalp Care Lotion

Application Example 3: Preparation of anti-dandruff scalp care essence

Application Example 4: Preparation of scalp care hair mask

Claims (9)

1. The anti-dandruff application of the dihydroartemisinin, wherein the use concentration of the dihydroartemisinin is 0.01-10 weight percent.

2. The use according to claim 1, wherein the dihydroartemisinin is used in a concentration of 0.1 to 5% by weight.

3. Use according to claim 1, wherein the anti-dandruff effect is achieved by inhibiting malassezia.

4. The use according to claim 3, wherein the minimum inhibitory concentration of dihydroartemisinin on malassezia is 1.0mg/mL.

5. Use of dihydroartemisinin for the preparation of a personal care product with anti-dandruff effect, wherein the dihydroartemisinin is used in a concentration of 0.01 to 10% by weight.

6. The use according to claim 5, wherein the dihydroartemisinin is used in a concentration of 0.1 to 5% by weight.

7. The use according to claim 5, wherein the anti-dandruff effect is achieved by inhibiting malassezia.

8. The use according to claim 7, wherein the minimum inhibitory concentration of dihydroartemisinin on malassezia is 1.0mg/mL.

9. The use according to claim 5, wherein the personal care product is selected from the group consisting of: anti-dandruff shampoo, anti-dandruff scalp care emulsion, anti-dandruff scalp care essence and scalp care hair mask.

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