A hair care composition containing extract of ilex kudingcha and ilex latifolia leaf and preparation method thereof

Abstract

The present application relates to a kind of anti-shedding shampoo and conditioner containing kudingcha ilex extract and preparation method, including shampoo and conditioner, and shampoo and / or conditioner contain kudingcha ilex extract;Kudingcha ilex extract is added to shampoo or conditioner, can play the effect that kudingcha ilex extract has anti-hair loss when using;Kudingcha ilex extract is 0.05% to 10% by weight ratio.In shampoo and conditioner, adding kudingcha ilex extract, long-term use can produce the effect of promoting hair growth and inhibiting hair loss, and collocation can realize better effect.

Description

Technical Field

[0001] This invention belongs to the field of hair loss prevention technology, and in particular relates to a hair loss prevention shampoo and conditioner composition containing extract of bitter tea and holly leaves, and its preparation method. Background Technology

[0002] In contemporary society, with the fast pace of life, increased work pressure, and the influence of environmental factors, hair loss is increasingly troubling many people. The incidence of hair loss is rising rapidly and showing a trend towards affecting younger people. As a result, the demand for hair loss prevention and hair regrowth products is growing.

[0003] Shampoo and conditioner are essential products for daily scalp care and hair loss prevention, and their market demand continues to rise. However, existing hair loss prevention shampoos and conditioners still have many technical shortcomings that need to be addressed in practical applications, making it difficult to achieve a synergistic effect of cleansing efficacy, hair condition improvement, and hair loss prevention.

[0004] From a cleansing perspective, existing anti-hair loss shampoos generally suffer from insufficient cleansing power. The scalp and hair surface easily accumulate oil, dust, dead skin cells, and styling product residue. If not thoroughly cleaned, these residues can clog hair follicles and irritate the scalp, affecting scalp health and weakening the effectiveness of anti-hair loss ingredients. Simultaneously, existing anti-hair loss shampoos have a significant weakness in improving hair appearance and texture. Most products focus solely on adding anti-hair loss ingredients, neglecting the conditioning of the hair cuticle. Hair cuticles are prone to lifting and breaking after physical friction, chemical stimulation, or environmental damage, leading to frizz, tangles, and reduced shine. Existing shampoos lack effective conditioning mechanisms, failing to provide continuous protection for the hair after cleansing and failing to smooth the cuticle, leaving hair still frizzy and unruly after drying, affecting overall appearance. Summary of the Invention

[0005] To solve the above-mentioned technical problems, the present invention provides an anti-hair loss shampoo and conditioner composition containing kuding tea holly leaf extract and a preparation method thereof.

[0006] The technical solution adopted in this invention is: an anti-hair loss shampoo and conditioner composition containing holly leaf extract of bitter tea, including shampoo and conditioner, wherein the shampoo and / or conditioner contain 0.05%-10% by weight of holly leaf extract of bitter tea; the holly leaves of bitter tea are extracted by decoction, the filtrate obtained by decoction is concentrated and dried, and the resulting solid substance is holly leaf extract of bitter tea;

[0007] Preferably, the extract of holly leaves from bitter tea contains kuding saponins as active substances.

[0008] A shampoo containing holly leaf extract of bitter tea includes 0.05%-10% by weight of holly leaf extract of bitter tea; the holly leaves of bitter tea are extracted by decoction in water, the filtrate obtained by decoction is concentrated and dried, and the resulting solid substance is holly leaf extract of bitter tea.

[0009] Preferably, it further includes 10-20% surfactant, 1.5-3% thickener, 0.1-8% conditioning agent, 0.01-2% preservative and 0.1-0.5% chelating agent, with the balance being water.

[0010] Preferably, it also includes 0.01-1% of a nucleating agent.

[0011] Preferably, the surfactant is selected from one or more of ammonium lauryl ether sulfate, ammonium lauryl sulfate, sodium lauryl ether sulfate, and TEA salt of dodecylbenzenesulfonate;

[0012] The thickener is selected from one or more of cocamide MEA, cocamide methyl MEA, sodium chloride, and cetyl alcohol;

[0013] The conditioning agent is selected from one or more of polyquaternium-39, polyquaternium-10, guar hydroxypropyltrimethylammonium chloride, and emulsified silicone oil;

[0014] The nucleating agent is selected from one or more of ceramides, sphingolipids, cholesterol, and phytosphingosine.

[0015] A method for preparing a shampoo containing holly leaf extract of bitter tea involves weighing each component by weight percentage, mixing and stirring until emulsified, to obtain a shampoo containing holly leaf extract of bitter tea.

[0016] A hair conditioner containing holly leaf extract of bitter tea includes 0.05%-10% by weight of holly leaf extract of bitter tea; the holly leaves of bitter tea are extracted by decoction in water, the filtrate obtained by decoction is concentrated and dried, and the resulting solid substance is holly leaf extract of bitter tea.

[0017] Preferably, it further includes 3-11% surfactant, 3-5% thickener, 3.5-4% hair conditioner and 0.55-1% preservative, with the balance being water.

[0018] Preferably, the surfactant is behenyltrimethylammonium methyl sulfate and / or behenyltrimethylammonium chloride;

[0019] The thickener is cetyl alcohol and / or stearyl alcohol;

[0020] The hair conditioning agent is one or more of bis-aminopropylpolydimethylsiloxane, squalane, and sea buckthorn fruit oil;

[0021] The preservatives are one or more of the following: methyl ester, sodium benzoate, phenoxyethanol, ethylhexylglycerin, 1,2-hexanediol, and p-hydroxyacetophenone;

[0022] Preferably, it also includes a pH adjuster and a conditioning agent;

[0023] Preferably, the pH adjuster is citric acid and / or sodium citrate;

[0024] The conditioning agent is one or more of hydrolyzed keratin, glycerin, D-panthenol, and EDTA-2Na.

[0025] A method for preparing a hair conditioner containing holly leaf extract of bitter tea involves weighing each component by weight percentage, mixing and stirring until emulsified, to obtain a hair conditioner containing holly leaf extract of bitter tea.

[0026] The advantages and positive effects of this invention are: Kuding tea holly can prevent abnormal scalp lipid metabolism, oxidative stress damage and inflammation-induced hair loss; shampoos and / or conditioners containing Kuding tea holly leaf extract can promote hair growth and inhibit hair loss.

[0027] Shampoos containing bitter tea and holly leaf extract have significant anti-hair loss effects and hair repair properties. When nucleating agents are added and the shampoo components are properly proportioned, the shampoo's cleansing effect, stability, and ability to improve hair loss can be further enhanced. Attached Figure Description

[0028] Figure 1 Comparison of the effects of active substances from bitter tea and holly leaves on the proliferation activity of hair follicle stem cells;

[0029] Figure 2 Mass spectrometry analysis of total saponin composition in holly leaf extract of bitter tea;

[0030] Figure 3 Determination of total saponin content in extracts of bitter tea holly, large-leaf holly, or wolfberry;

[0031] Figure 4 Comparison of the proliferative activity of total saponins from bitter tea holly, large-leaf holly, or wolfberry on hair follicle stem cell proliferation.

[0032] Figure 5 Mass spectrometry analysis of holly saponins in *Ilex kudingcha*, *Ilex macrophylla*, or *Ilex thunbergii*; A: *Ilex kudingcha*; B: *Ilex macrophylla*; C: *Ilex thunbergii*;

[0033] Figure 6 A shampoo containing bitter tea and holly leaf extract shows its effect on preventing hair loss in mice with androgenetic alopecia.

[0034] Figure 7Microscopic image of a shampoo product without added nucleating agents; magnification: 200x;

[0035] Figure 8 Microscopic images of shampoo products containing nucleating agents; magnification: 200x;

[0036] Figure 9 The repairing effects of different bitter tea and holly shampoos on hair cuticles;

[0037] Figure 10 The repairing effects of different bitter tea and wintergreen hair conditioners on hair cuticles;

[0038] Figure 11 A hair conditioner containing bitter tea and holly leaf extract shows its effect on preventing hair loss in mice with androgenetic alopecia. Detailed Implementation

[0039] The embodiments of the present invention will now be described with reference to the accompanying drawings.

[0040] This invention relates to an anti-hair loss shampoo and conditioner composition containing holly leaf extract of bitter tea and its preparation method. The anti-hair loss shampoo and conditioner composition includes shampoo and conditioner, wherein the shampoo and / or conditioner contain plant extracts, the plant extracts being holly leaf extract of bitter tea or plant compositions with holly leaf of bitter tea as the main component.

[0041] Kuding tea, made from the holly genus of the Aquifoliaceae family, is a tree primarily produced in Hainan and Guangxi provinces. Its dried leaves are mainly used medicinally and as food. They are cool in nature, bitter and slightly sweet in taste, and are believed to have effects such as dispelling wind and heat, clearing the head and eyes, relieving irritability and quenching thirst. Kuding tea has a long history in my country, with the large-leaved holly being a representative variety. Ilex latifolia Thunb.), bitter tea holly ( Ilex kudingcha CJ Tseng) and holly ( Ilex cornuta Lindl. et Paxt.). The main component of Kuding tea holly is saponins, and it also contains flavonoids, polyphenols, polysaccharides, and volatile oils. The extract obtained from the effective components of Kuding tea holly leaves can be used to prepare topical anti-hair loss preparations.

[0042] Kuding tea holly leaf extract is obtained by decoction of the leaves. The solution obtained after decoction can be further concentrated and dried, and the resulting Kuding tea holly leaf extract can be used as a raw material to prepare other products. In some embodiments of the present invention, the Kuding tea holly leaf extract contains effective components including total polysaccharides, total phenolic acids, total saponins, and total flavonoids. The total polysaccharides, total phenolic acids, total saponins, and total flavonoids in the aqueous extract of Kuding tea holly leaves can be separated using ethanol of different concentrations. The separated total polysaccharides, total phenolic acids, total saponins, and total flavonoids are then mixed and compounded to obtain a new type of Kuding tea holly leaf extract. In other embodiments of the present invention, Kuding tea holly leaf extract specifically refers to the saponin-like active substances extracted.

[0043] Boil holly leaves of Kuding tea in water, then add distilled water at a w / v ratio of 1:10-15 to extract the active ingredients. The extraction time is 0.5-3 hours. The filtered solution is concentrated and dried to obtain the holly leaf extract of Kuding tea. Alternatively, boil holly leaves of Kuding tea in water to extract the active ingredients. After concentrating the filtrate, add 70% ethanol and collect the precipitate as total polysaccharides. The supernatant is extracted and loaded onto an adsorption resin column. Eluenting with 30% ethanol yields total phenolic acids, 50% ethanol yields total saponins, and 80% ethanol yields total flavonoids. The total polysaccharides, total phenolic acids, total saponins, and total flavonoids are mixed and dried to obtain the holly leaf extract of Kuding tea. Alternatively, the total saponin components prepared by the aforementioned method can be used as the extract of Ilex kudingcha leaves; 70% ethanol is added to the aqueous extract of Ilex kudingcha leaves to remove total polysaccharides, the separated liquid portion is loaded onto an adsorption resin column, and the total phenolic acids are removed by elution with 30% ethanol. The eluted components with 50% ethanol are then collected, concentrated, and dried to obtain the total saponins of Ilex kudingcha leaves.

[0044] Experimental results showed that *Ilex kudingcha* leaf extract significantly inhibited the levels of triglycerides, total cholesterol, and 5α-reductase in a mouse model of androgenetic alopecia; it also inhibited excessively high levels of ROS or MDA or abnormal expression of inflammatory factors TNF-α, IL-1β, IL-6, and INF-γ in mouse hair follicle stem cells induced by ultraviolet irradiation or hydrogen peroxide. *Ilex kudingcha* leaf extract can be used to improve hair loss caused by various mechanisms, such as inhibiting hair loss caused by abnormally high sebum synthesis and secretion in the scalp or excessively high androgen levels, clearing hair loss caused by excessively high levels of reactive oxygen species (ROS) induced by physical damage or chemical stimulation, and inhibiting hair loss caused by abnormal expression of inflammatory factors induced by physical, chemical, or biological factors.

[0045] Adding holly leaf extract from Kuding tea to shampoo or conditioner can enhance its effects on promoting hair growth and inhibiting hair loss. The weight percentage of holly leaf extract is 0.05%-10%. This extract is obtained by concentrating the solution obtained by decocting holly leaves in water and then filtering it. Holly leaf extract contains holly saponins and other active substances.

[0046] A shampoo containing holly leaf extract of bitter tea, comprising, by weight percentage, 0.05%-10% holly leaf extract of bitter tea, 10-20% surfactant, 1.5-3% thickener, 0.01-1% nucleating agent, 0.1-8% conditioning agent, 0.01-2% preservative and 0.1-0.5% chelating agent, with the balance being water.

[0047] The surfactant is selected from one or more of ammonium lauryl ether sulfate, ammonium lauryl ether sulfate, sodium lauryl ether sulfate, and TEA salt of dodecylbenzenesulfonate; the thickener is selected from one or more of cocamide MEA, cocamide methyl MEA, sodium chloride, and cetyl alcohol; the nucleating agent is selected from one or more of ceramides (NP, AP, NG, NS), sphingolipids, cholesterol, and phytosphingosine; the conditioning agent is selected from one or more of polyquaternium-39, polyquaternium-10, guar hydroxypropyltrimethylammonium chloride, and emulsified silicone oil; the preservative is selected from one or more of methyl ester, phenoxyethanol, ethylhexylglycerin, and sodium benzoate; and the chelating agent is selected from one or more of disodium EDTA, citric acid, and sodium citrate.

[0048] In some embodiments of the present invention, the surfactant comprises ammonium lauryl ether sulfate and ammonium lauryl sulfate in a mass ratio of 1:0.8-1.25, specifically 1:1, 1.2:1 or 1.2:1.5; the thickener comprises cocamidomethyl MEA, sodium chloride and cetyl alcohol in a mass ratio of 1:0.5-1:0.6-1, specifically 1:0.6:0.8, 1:0.6:0.7 or 0.8:1:0.8.

[0049] In preparing shampoo, each component is weighed according to its weight percentage, mixed and stirred until emulsified. The components can be added sequentially according to the characteristics of each additive and existing technical methods to finally obtain the shampoo product. In some embodiments of the present invention, the shampoo preparation method is as follows:

[0050] (1) Weigh out the following ingredients by weight percentage: surfactant, thickener, cationic conditioning agent, chelating agent, and preservative;

[0051] (2) Add purified water, bitter tea holly leaf extract, and chelating agent to an emulsifying pot, turn on the stirrer, and stir at 25-35 rpm for 10-15 min; stop stirring, add the first conditioning agent, and stir at 25-35 rpm for 10-15 min. Add the first surfactant, and stir at 20-30 rpm for 10-15 min, then stop stirring;

[0052] (3) Add the second surfactant and the second conditioner to the emulsifying pot, turn on the stirring, turn on the homogenizer at 25-35 rpm, and turn on the homogenizer at 2000-3000 rpm to make the second surfactant completely dispersed to help dissolve, and then add the first thickener;

[0053] (4) Heat to 80℃ and continue stirring for 20-30 minutes; keep the aqueous phase warm and stir for 20-40 minutes to defoam; add the second thickener sodium chloride, stir well and cool down.

[0054] Adding bitter tea and holly leaf extract to the shampoo can promote hair growth and inhibit hair loss with long-term use. The addition of cationic conditioning agents, without amphoteric conditioning agents, can shorten the hair cuticle repair time. Furthermore, the shampoo incorporates ceramides, scientifically formulated with surfactants and thickeners, allowing the surfactants and thickeners to form crystalline particles from a conventional emulsification system. During cleansing, these particles only exert their cleaning power, leaving no residue on the hair surface, maximizing quick rinsing and minimal residue. Simultaneously, the emulsion droplets form a uniform protective film on the hair surface upon contact with water, smoothing the cuticles, reducing frizz, and maintaining volume.

[0055] A hair conditioner containing holly leaf extract of bitter tea, comprising, by weight percentage, 0.05%-10% holly leaf extract of bitter tea, 3-11% surfactant, 3-5% thickener, 3.5-4% hair conditioning agent, 0.55-1% preservative, pH adjuster and conditioning agent, with the balance being water.

[0056] The surfactant is behenyltrimethylammonium methyl sulfate and / or behenyltrimethylammonium chloride; the thickener is cetyl alcohol and / or stearyl alcohol; the hair conditioning agent is one or more of bis-aminopropyl polydimethylsiloxane, squalane and sea buckthorn fruit oil; the preservative is one or more of methyl ester, sodium benzoate, phenoxyethanol, ethylhexylglycerin, 1,2-hexanediol and p-hydroxyacetophenone; the pH adjuster is citric acid and / or sodium citrate; and the conditioning agent is one or more of hydrolyzed keratin, glycerin, D-panthenol and EDTA-2Na.

[0057] When preparing a hair conditioner containing bitter tea and holly leaf extract, weigh all the raw material components and add them to water, stirring until emulsified. To improve emulsification efficiency, the temperature can be raised to 75-90℃, and emulsification can be carried out while stirring. For raw materials that may have their properties damaged by overheating, they can be added to the stirred reaction system during the cooling process.

[0058] In some embodiments of the present invention, the conditioner is prepared as follows:

[0059] (1) Add purified water to the water pot, turn on the stirrer, stir at 25-500 rpm for 10-15 minutes, and heat to 80℃;

[0060] (2) Add surfactants (behenryl trimethylammonium methyl sulfate and / or behenryl trimethylammonium chloride), thickeners (cetyl alcohol and / or stearyl alcohol), hair conditioning agents (squalane and / or sea buckthorn fruit oil), preservatives, conditioning agents, etc. into the emulsifying pot, turn on the stirring, and heat to 80°C at 25-35 rpm to homogenize appropriately to help disperse;

[0061] (3) Pump the aqueous phase into the main pot, add preservative, control the temperature at about 80℃, keep warm and stir for 5-15 minutes, homogenize for 5-8 minutes at a speed of 2600-3000 rpm, and keep warm for 15-20 minutes.

[0062] (4) Cool down to 65-70℃ and keep warm. Add the hair conditioner (bis-aminopropyl polydimethylsiloxane) and keep warm and stir evenly for 5-20 minutes. Adjust the pH value to 3.0-5.0 using a pH adjuster.

[0063] (5) Continue to cool down to below 40°C and remove from the pot.

[0064] Adding bitter tea holly leaf extract to hair conditioner can promote hair growth and inhibit hair loss with long-term use. For even better results, use it in conjunction with a shampoo containing bitter tea holly leaf extract.

[0065] The present invention will now be described with reference to the accompanying drawings. Experimental methods not specifically described in terms of operation steps are performed in accordance with the corresponding product manuals. Unless otherwise specified, the instruments, reagents, and consumables used in the embodiments can be purchased from commercial companies.

[0066] Example 1: Preparation and characterization of Ilex kudingcha leaf extract

[0067] 1.1 Preparation of Ilex kudingcha leaf extract

[0068] The active ingredients of Kuding tea holly leaves can be extracted by crushing dried Kuding tea holly leaves and then boiling them in water.

[0069] Weigh 1000g of dried bitter tea holly leaves, pulverize them, add distilled water at a ratio of w / v = 1:15 and heat to boil for 2 hours to extract; filter while hot using a 100-mesh stainless steel sieve, concentrate the filtrate using a flash evaporator (JMF-320, Xi'an), dry the concentrated product, and the resulting solid substance is the bitter tea holly leaf extract.

[0070] 1.2 Component Analysis of Ilex kudingcha Leaf Extract

[0071] To further analyze the key active components in the holly leaves of Kuding tea, different types of active components in the holly leaf extract of Kuding tea were separated and purified. Total polysaccharides, total phenolic acids, total saponins and total flavonoids were collected, and the functions of each type of active component were analyzed and verified.

[0072] 1000g of dried Kuding tea holly leaves were weighed, pulverized, and extracted by boiling with distilled water at a w / v ratio of 1:15 for 2 hours. The extract was then filtered while hot using a 100-mesh stainless steel sieve. The filtrate was concentrated to a suspension with a density of 1.1 (approximately 2.5 L) using a flash evaporator (JMF-320, Xi'an). Ethanol was added to the Kuding tea holly leaf extract suspension to adjust the ethanol content to 70%. After mixing, the solution was allowed to stand overnight. The precipitate was washed three times with anhydrous ethanol. The main component obtained was the total polysaccharide, totaling 35.4 g. The supernatant was further concentrated to prepare a solution with a solid content of approximately 1%. The solution was loaded onto an AB-8 macroporous adsorption resin column (diameter: 5.5 cm; column height: 35 cm; column volume: approximately 850 mL) at a flow rate of 2 mL / min. The column was equilibrated with 5% ethanol solution to elute water-soluble impurities. Gradient elution was then performed with ethanol solutions of different concentrations at a rate of 1 mL / min. The components obtained from the elution with 30% ethanol solution were mainly total phenolic acids, totaling 18.7 g; the components obtained from the elution with 50% ethanol solution were mainly total saponins, totaling 21.8 g; and the components obtained from the elution with 80% ethanol solution were mainly total flavonoids, totaling 28.4 g.

[0073] 1.3 Effects of active components from bitter tea and holly leaves on the proliferation activity of hair follicle stem cells

[0074] The proliferative activity of rat hair follicle stem cells (HFSCs) was tested using the isolated total polysaccharides, total phenolic acids, total saponins, and total flavonoids. Rat hair follicle stem cells (HFSCs) were purchased from Wuhan Pronosei Life Science Technology Co., Ltd. HFSCs were placed in a special culture medium and continuously cultured in a 37 ℃, 5% CO2 incubator, with the medium changed every other day. When the cells reached 70%–80% confluence, they were passaged. HFSCs in the logarithmic growth phase were harvested and cultured at a ratio of 1×10⁻⁶. 4100 μL of HFSCs (cells / well) were seeded into 96-well plates, with four groups: control group, bitter tea holly group, total polysaccharide group, total phenolic acid group, total saponin group, and total flavonoid group. The control group used basal medium, while the other groups used basal medium solutions containing 0.1% of the corresponding components. HFSCs were seeded into the control and experimental groups, respectively, and cultured for 24 h. 10 μL of CCK-8 reagent was added to each group, and the plates were returned to the incubator for another 1 h. Absorbance was measured at 450 nm. Each group was divided into three replicates to ensure accuracy. The increase in cell proliferation in each experimental group relative to the control group was compared, and the cell proliferation enhancement rate was calculated to measure the promoting effect of different experimental groups on cell proliferation.

[0075] Cell proliferation enhancement rate % = (experimental group absorbance value - control group absorbance value) / control group absorbance value × 100%.

[0076] Depend on Figure 1 It was found that the extract of Ilex kudingcha leaves significantly promoted the proliferation of hair follicle stem cells (HFSCs), with an average cell proliferation rate increase of 37.1% compared to the control group. Its main components (total polysaccharides, total phenolic acids, total saponins, and total flavonoids) all exhibited certain proliferative activities, with the total saponin group showing the most significant promoting effect (proliferation rate increased by 58.4%). This indicates that Ilex kudingcha has the effect of enhancing the activity of hair follicle stem cells, suggesting its potential application value in preventing hair loss, with total saponins being the core active component.

[0077] 1.4 Analysis and Performance Comparison of Total Saponins in Ilex kudingcha Leaf Extract

[0078] Further analysis was conducted on the key active components of total saponins in holly leaves of bitter tea, and the components were separated and identified by UPLC-Q-TOF / MS.

[0079] Chromatographic column: CORTECS T3 (2.1 mm × 150 mm, 1.6 μm); chromatographic gradient elution conditions: mobile phase A was acetonitrile, and mobile phase B was 0.1% formic acid aqueous solution; flow rate: 0.2 mL / min; gradient elution (0–5 min: 5% A, 95% B; 5–35 min: 25%, 75% B; 35–40 min: 40% A, 60% B; 40–45 min: 75% A, 25% B; 45–50 min: 95% A, 5% B). Results are as follows. Figure 2The key saponin components are shown in the figure. Table 1 illustrates the most representative saponin components identified through secondary mass spectrometry fragmentation analysis. The main saponin components are kudinoside G, kudinoside C, kudinoside A, and kudinoside D. Fractions were collected based on peak times, and the four saponin components were enriched, concentrated, and dried. The fraction from 36.5 min to 40.41 min was also collected, concentrated, and dried. The activity of the four kudinoside saponins in promoting hair follicle stem cell proliferation was determined according to the method described in section 1.3, and the results are shown in Table 1. It can be seen that all four saponin components at a concentration of 10 μg / mL have activity in promoting hair follicle stem cell proliferation, but their activity is lower than that of the fraction from 36.5 min to 40.41 min (total kudinoside saponins) at the same concentration. Comparison shows that a mixture of different types of kudinoside saponins (total kudinoside saponins) has a better effect.

[0080] Table 1. Total saponin composition and activity analysis in holly leaf extract of bitter tea.

[0081]

[0082] Example 2: Determination and performance analysis of total saponin content in bitter tea holly, large-leaf holly and wolfberry

[0083] 2.1 Comparison of total saponin content in bitter tea holly, large-leaf holly, and wolfberry

[0084] A comparison was made between representative Kuding tea leaves: Ilex kudingcha, Ilex macrocarpa, and Ilex chinensis. The leaves of Ilex kudingcha, Ilex macrocarpa, and Ilex chinensis were purchased from medicinal herb markets in Guangxi Zhuang Autonomous Region, Zhejiang Province, and Guizhou Province, respectively. The total saponin content of the three raw materials was tested.

[0085] The content of total saponins in three batches of Kuding tea (Ilex kudingcha, Ilex macrocarpa, and Ilex cornuta) was determined by ultraviolet-visible spectrophotometry, using ginsenoside Re (purchased from Shanghai Yuanye Biotechnology Co., Ltd., catalog number A10036) as a reference standard and vanillin-glacial acetic acid method for color development. Specifically, 100 μL of the test sample or reference solution was accurately pipetted into a 10 mL stoppered test tube, the solvent was evaporated in a 60 °C water bath, 0.2 mL of 10% vanillin-glacial acetic acid solution and 0.8 mL of perchloric acid were added, and the mixture was incubated in a 60 °C water bath for 15 min. After that, the tube was placed in an ice-water bath for 5 min, 5 mL of glacial acetic acid was added, and the mixture was stirred well. The absorbance was measured at 550 nm.

[0086] Preparation of reference solution: Accurately weigh 10 mg of ginsenoside Re reference standard, place it in a 10 mL volumetric flask, add methanol to make up to volume and shake well to obtain a 1 mg / mL ginsenoside Re reference solution.

[0087] Accurately pipette 0, 10, 20, 40, 60, 80, and 100 μL (1 mg / mL) of ginsenoside Re standard solution and measure the absorbance (A) using the method described above. Plot a standard curve with A concentration on the x-axis (X) and A value on the y-axis (Y). The obtained linear regression equation is y = 0.0204x + 0.0017, R0 2 =0.9995.

[0088] Preparation of the test solution: Weigh 100 g each of dried bitter tea holly leaves, large-leaf holly leaves, and wolfberry leaves, add 10 times the amount of water, soak for 30 minutes, heat to boiling and extract for 1 hour, filter, concentrate the filtrate to a relative density of 1.1-1.2, dry under reduced pressure, and pulverize to obtain each extract. Accurately weigh 10 mg of each extract powder, place it in a 10 mL volumetric flask, add methanol to make up to volume and shake well to obtain the test solution.

[0089] The total saponin content in the test samples was determined according to the above method, and the results are shown in Table 2. The highest total saponin content was found in *Ilex kudingcha*, with an average content of 7.43%; the total saponin content in *Ilex macrocarpa* was 5.18%; and the total saponin content in *Ilex cornuta* was 4.96%. Figure 3 As shown, the holly variety of Kuding tea has a higher content of total saponins, an active ingredient, compared to the other two types.

[0090] Table 2. Total saponin content of three extracts (Ilex kudingcha, Ilex macrocarpa, and Ilex cornuta) from different batches.

[0091]

[0092] 2.2 Comparison of the effects of total saponins from bitter tea holly, large-leaf holly, and wolfberry on the proliferative activity of hair follicle stem cells

[0093] Following the method for preparing total saponins in Example 1, total saponins from three species—Ilex kudingcha, Ilex macrocarpa, and Ilex cornuta—were separated, and the effects of the three total saponins on the proliferation activity of hair follicle stem cells were determined at a concentration of 0.1%.

[0094] When HFSCs reach a confluence of 70%–80%, passage them and collect HFSCs in the logarithmic growth phase at a ratio of 1 × 10⁻⁶. 4100 μL of reagent (per well) was inoculated into 96-well plates, with four groups: control group, Kuding tea group, Kuding tea holly group, large-leaf holly group, and holly group. The control group used basal medium, while the other groups used basal medium solutions containing 0.1% of the corresponding total saponins. The Kuding tea group consisted of one-third each of Kuding tea holly, large-leaf holly, and holly (0.033% of each). The plates were incubated for 24 h, then 10 μL of CCK-8 reagent was added, and the plates were returned to the incubator for another h. The absorbance was measured at 450 nm. Each group was tested in triplicate to ensure accuracy.

[0095] The cell proliferation enhancement rate was calculated according to the method in Example 1, and the results are as follows: Figure 4 As shown, the total saponins in the Kuding tea group, Kuding tea holly group, large-leaf holly group, and wolfberry group all had significant proliferative effects on HFSCs. Compared with the control group, the average cell proliferation rate of total saponins in Kuding tea increased by 41.1%, in large-leaf holly by 36.8%, in wolfberry by 27.9%, and in Kuding tea holly by 57.9%. This indicates that at the same level, the activity of total saponins in Kuding tea holly is significantly higher than that in large-leaf holly or wolfberry, or a combination of the three (Kuding tea), suggesting that the potential anti-hair loss effect of Kuding tea holly is the best among various Kuding teas.

[0096] 2.3 Analysis of saponin components in Ilex kudingcha, Ilex macrocarpa, or Ilex thunbergii

[0097] Furthermore, the key active components of total saponins from bitter tea holly, large-leaf holly, or wolfberry were identified and compared, and the components were separated and identified by UPLC-Q-TOF / MS.

[0098] Following the mass spectrometry analysis conditions shown in Example 1, parallel measurements were performed on three samples at 10 mg / mL, and the results are as follows: Figure 5 As shown, the three substances exhibit similar component categories, while their individual component contents also show significant differences. Based on a comparison of relative peak areas, the calculated contents of kudinoside G, kudinoside C, kudinoside A, and kudinoside D in Kuding tea holly are 1.57 times that in large-leaf holly and 2.54 times that in holly. This indicates that the content of active holly saponins in Kuding tea holly is significantly higher than that in large-leaf holly or holly.

[0099] Simultaneously, the fraction from 36.5 min to 40.41 min was collected and concentrated. The activity of four types of Kuding tea holly saponins in promoting hair follicle stem cell proliferation was determined according to the method described in Example 1, and the results are shown in Table 3. It can be seen that the saponins from Kuding tea holly, large-leaf holly, or wolfberry holly all exhibited activity in promoting hair follicle stem cell proliferation at a concentration of 10 μg / mL, but the activities varied considerably. The holly saponins from Kuding tea holly showed the best activity, significantly superior to those from large-leaf holly or wolfberry holly. This indicates that different total saponin extracts exhibit different abilities to promote hair follicle stem cell proliferation due to differences in the ratio of saponin components.

[0100] Table 3. Proliferative Activity of Ilex kudingcha saponins from Ilex kudingcha, Ilex macrocarpa, or Ilex cornuta

[0101]

[0102] Example 3: Preparation of shampoo containing bitter tea and holly leaf extract

[0103] The bitter tea holly leaf extract prepared in Example 1 was added to the shampoo. In addition to the bitter tea holly leaf extract, the shampoo also contained surfactants, thickeners, conditioners, preservatives, chelating agents and water. The specific categories and amounts added are shown in Table 4.

[0104] Table 4

[0105]

[0106] During preparation, the surfactant, thickener, conditioning agent, chelating agent, and preservative are weighed out according to their weight percentages.

[0107] Add purified water, bitter tea holly leaf extract, and chelating agent to an emulsifying pot, start stirring, and stir at 25-35 rpm for 10-15 minutes; stop stirring, add conditioning agent (polyquaternium-39, guar hydroxypropyltrimethylammonium chloride), and stir at 25-35 rpm for 10-15 minutes. Add surfactant (ammonium lauryl sulfate), and stir at 20-30 rpm for 10-15 minutes, then stop stirring.

[0108] The surfactant (ammonium lauryl ether sulfate) and conditioning agent (emulsified silicone oil) were added to the emulsification pot. Stirring was started at 25-35 rpm, and homogenization was started at 2000-3000 rpm to ensure complete dispersion and dissolution of the surfactant. Thickener (cocamide MEA, cetyl alcohol) and preservative were added. The temperature was raised to 80℃, and stirring was continued for 20-30 minutes. The aqueous phase was kept warm and stirred for 20-40 minutes to remove foam. Thickener (sodium chloride) was added, stirred evenly, and cooled to prepare a shampoo containing holly leaf extract of bitter tea.

[0109] Example 4: The effect of the amount of bitter tea and holly leaf extract added on the anti-hair loss effect of shampoo

[0110] To investigate the effect of adding *Ilex kudingcha* leaf extract on the hair growth-promoting and hair loss-preventing effects of shampoo, shampoo products with added *Ilex kudingcha* leaf extract at mass percentages of 0, 0.05, 1, 3.35, and 5% were prepared according to the method in Example 3, and designated as Experiment 1-1, Experiment 1-2, Experiment 1-3, Experiment 1-4, and Experiment 1-5, respectively. These products were then used in androgenetic alopecia mouse models. Thirty-five male C57BL / 6J mice were randomly divided into seven groups: a blank group, a model group, and Experiment 1-1, Experiment 1-2, Experiment 1-3, Experiment 1-4, and Experiment 1-5. Except for the blank group, the other groups were treated daily with 200 μL of testosterone (5 mg / mL) for modeling. After complete absorption, 200 μL of the test substance (prepared in the experimental examples) shampoo was applied. The model group was treated with a blank matrix without active substances for 14 consecutive days. After anesthetizing the mice, the hair growth in the bald area on their backs was photographed. Then, a 1cm... 2 A pull-out test was conducted on the back test area using transparent tape. The number of hairs removed was counted to reflect the anti-hair loss effect of each group. Hair that grew out in the shaved area was then shaved off with a razor, collected on weighing paper, and weighed using a 1 / 100,000 electronic balance.

[0111] The effects of bitter tea and holly on the amount and weight of hair loss in mice with androgenetic alopecia are as follows: Figure 6 As shown in the results, Experiment 1-1 had no significant effect on preventing hair loss, and the hair of the mice remained consistent with that of the model group. The Kuding tea and holly leaf shampoos in Experiments 1-2 to 1-5 significantly inhibited hair loss in androgenetic alopecia mice, reduced the amount of hair loss, and increased the weight of hair in the bald areas. Furthermore, the treatment group in Experiment 1-3 (containing 1% Kuding tea and holly leaf extract) showed the best effects compared to the treatment groups in Experiment 1-4 (containing 3.35% Kuding tea and holly leaf extract) and Experiment 1-5 (containing 5% Kuding tea and holly leaf extract), all demonstrating significant anti-hair loss effects.

[0112] Example 5: The effect of surfactants on shampoo performance

[0113] To investigate the effects of different surfactants on shampoo performance, shampoos containing surfactants of different compositions were prepared. The mass percentage of bitter tea holly leaf extract in the shampoos was 1%. The surfactant addition is shown in Table 5, and the other components are the same as in Table 4.

[0114] Table 5

[0115]

[0116] Shampoo products of Experiment 2-1, Experiment 2-2 and Experiment 2-3 were prepared according to the method of Experiment 3; and the cleaning power, residue rate and stability of each group of shampoos were compared.

[0117] Cleaning power analysis. Thirty strands of human hair were selected and divided into three groups of ten strands each. Artificial hair oil was evenly applied to the surface of the hair, and the hair was washed with different shampoos. After drying, the average weight was measured. The results are shown in Table 6.

[0118] Table 6

[0119]

[0120] Residual rate analysis. Thirty strands of real human hair were selected and divided into three groups of 10 strands each. Each group was washed with 2g of different shampoos and then dried. The process was repeated 20 times, and the hair was weighed. The results are shown in Table 7.

[0121] Table 7

[0122]

[0123] Stability analysis. Different shampoos were selected and placed under different temperature and light conditions for 3 months to observe their stability and score them (5 points - stability OK; 4 points - stability slightly poor but acceptable; 3 points - minor flaws; below 3 points - unqualified). The scoring results are shown in Table 8.

[0124] Table 8

[0125]

[0126] Based on the comprehensive evaluation of the above experiments, Experiment 2-2 showed the strongest overall performance, followed by Experiment 2-1 and Experiment 2-3. When the surfactant components include equal amounts of ammonium lauryl ether sulfate and ammonium lauryl sulfate, it is more beneficial to improve the overall performance of the shampoo.

[0127] Example 6: The effect of thickeners on the anti-hair loss effect of shampoos

[0128] To investigate the effects of different thickeners on shampoo performance, shampoos containing thickeners of different compositions were prepared. The shampoos contained 1% by mass of bitter tea holly leaf extract, and 8% each of ammonium lauryl ether sulfate and ammonium lauryl sulfate as surfactants. The thickener addition is shown in Table 9, and other components are the same as in Table 4.

[0129] Table 9

[0130]

[0131] Shampoo products of Experimental Example 3-1, Experimental Example 3-2 and Experimental Example 3-3 were prepared according to the method of Example 3. The cleaning power, residue rate and stability of each group of shampoos were compared. The comparison and verification method is as shown in Example 5. The results of the cleaning power comparison are shown in Table 10, the results of the residue rate comparison are shown in Table 11, and the results of the stability comparison are shown in Table 12.

[0132] Table 10

[0133]

[0134] Table 11

[0135]

[0136] Table 12

[0137]

[0138] Based on the comprehensive evaluation of the above experiments, Experiment 3-2 showed the strongest overall performance, followed by Experiment 3-3 and Experiment 3-1. A mass ratio of cocoamide MEA, sodium chloride, and cetyl alcohol of 1:0.6:0.7 was more conducive to improving the overall performance of the shampoo.

[0139] Example 7: The effect of nucleating agents on the anti-hair loss effect of shampoo

[0140] Ammonium lauryl ether sulfate exists in ammonium lauryl sulfate solution in both micelle and crystal forms. It can interconvert between micelle and crystal forms and undergo reversible reactions. The content of micelles and crystals is greatly affected by temperature, salt content, etc. In order to promote the formation of crystal form, nucleating agents need to be added to provide a support for crystallization and make the crystals exist stably in the system, so as to achieve the shampoo's strong cleaning, low residue, and fast rinsing effect.

[0141] Nucleating agents were added to the shampoo product. The nucleating agent used was ceramide (in this example, Best-Carrier transparent composite ceramide was used, which contains NP, AP, NG and NS), with mass ratios of 0.01, 0.05, 0.1 and 0.5, respectively. Other components are shown in Table 13. The shampoo product was prepared according to the preparation method of Example 3. When the product cooled to 50-60°C, the nucleating agent was added and mixed to obtain the final shampoo product. The products were numbered as Experiment 4-1, Experiment 4-2, Experiment 4-3 and Experiment 4-4 according to the mass ratio of the nucleating agent.

[0142] Table 13

[0143]

[0144] Microscopic image (200x magnification) of shampoo product without nucleating agents (before crystallization). Figure 7 As shown, the microscopic images (magnification 200x) of the products from Experiments 4-1, 4-2, 4-3, and 4-4 after the addition of the nucleating agent are as follows. Figure 8 As shown in the figure, the comparison shows that an appropriate concentration of nucleating agent can effectively promote crystallization.

[0145] To analyze the stability of each group of shampoos, different shampoos were selected and placed under different temperature and light conditions for 3 months. Their stability was observed and scored (5 points - stability OK, 4 points - stability slightly poor but acceptable, 3 points - minor defects, below 3 points - unqualified). The results are shown in Table 14.

[0146] Table 14

[0147]

[0148] Overall, Experiment 4-2 showed the best stability, followed by Experiment 4-3, Experiment 4-4, and Experiment 4-1.

[0149] Example 8: The effect of the ratio of surfactant, thickener and nucleating agent on shampoo performance

[0150] Shampoos containing different proportions of surfactants, thickeners, and nucleating agents were prepared. The mass percentage of Kuding tea holly leaf extract was 1%. The proportions of surfactant (ammonium lauryl ether sulfate and ammonium lauryl sulfate in a mass ratio of 1:1), thickener (cocamide MEA, sodium chloride, and cetyl alcohol in a mass ratio of 1:0.6:0.7) and nucleating agent were 5:2:0.05, 15:2:0.05, 20:2:0.05, and 30:2:0.05, respectively. Other ingredients were the same as those in Table 13. The resulting shampoo products were Experimental Example 5-1, Experimental Example 5-2, Experimental Example 5-3, and Experimental Example 5-4, respectively.

[0151] To investigate the efficacy of the preferred shampoo formula in this patent on human trials, suitable volunteers were recruited to test the anti-hair loss and repair effects of the Kuding Tea and Holly Shampoo. Suitable volunteers were recruited for the human trials of the Kuding Tea and Holly Shampoo. Subjects were randomly assigned to groups of 6, aged 18-50 years, both male and female (stratified by gender), and were required to have mild to moderate hair loss, not have used hair growth medications in the past 3 months, and have no serious scalp diseases. One mL was applied once daily to the scalp and massaged until absorbed. The preparation method of the essence described in Experiment Examples 5-1-5-4 was followed for group testing.

[0152] During testing, the efficacy of cosmetic hair loss prevention was tested using a 60-comb test, and the shed hairs were collected and counted. Before use and 28 days after use, images of hair in designated areas were collected using a trichoscope. Results for representative volunteers are shown below. Figure 9 As shown in Table 15, Experimental Examples 5-1, 5-2, 5-3, and 5-4 all exhibited significant cuticle repair effects on hair loss. Before use, the hair showed dull luster, raised cuticle edges, and breakage. After using the bitter tea shampoo, the overall hair quality improved, hair shine increased, the texture became stronger, and the surface became smoother. Furthermore, the amount of hair loss was significantly reduced. Among these results, Experimental Example 5-2 (surfactant, thickener, and nucleating agent in a mass ratio of 15:2:0.05) showed the best hair loss prevention effect, with a hair loss improvement rate of 60.0%; Experimental Example 5-3 had a hair loss improvement rate of 50.0%; Experimental Example 5-4 had a hair loss improvement rate of 43.7%; and Experimental Example 5-1 had a hair loss improvement rate of 37.5%.

[0153] Table 15 Statistics on the Number of Hair Loss Cases

[0154]

[0155] In conclusion, shampoos containing holly leaf extract from bitter tea have significant effects in preventing hair loss and repairing hair. When nucleating agents are added and the shampoo components are properly proportioned, the shampoo's cleansing effect, stability, and ability to improve hair loss can be further enhanced.

[0156] Example 9: Hair conditioner containing bitter tea and holly leaf extract

[0157] By weight, the conditioner contains 10% bitter tea holly leaf extract, 6% behenyltrimethylammonium methyl sulfate, 5% behenyltrimethylammonium chloride, 3.5% stearyl alcohol, 4% bis-aminopropyl polydimethylsiloxane, 0.15% methyl methacrylate, 0.4% sodium benzoate, 0.01% citric acid, 0.01% hydrolyzed keratin, and 0.05% EDTA-2Na, with the balance being water.

[0158] The specific preparation method is as follows:

[0159] a. Add purified water, bitter tea holly leaf extract, methyl ester, sodium benzoate, EDTA-2 Na, and citric acid to an emulsifying pot, turn on the stirrer, stir at 25-50 rpm, and heat to 80 ℃ (appropriate homogenization can help with dispersion).

[0160] b. Add behenyltrimethylammonium methyl sulfate, behenyltrimethylammonium chloride, stearyl alcohol, etc. into the oil phase pot and heat to 80 ℃, then add to the emulsification pot. Homogenize at 2600-3000 rpm for 5-8 min and keep warm for 15-20 min.

[0161] c. Cool to 65-70℃ and keep warm. Add bis-aminopropylpolydimethylsiloxane, keep warm and stir evenly for 15-20 minutes. Continue to cool to below 50℃ and add hydrolyzed keratin. Adjust the pH value with citric acid.

[0162] d. Continue cooling to below 40 ℃ and remove from the pot to prepare a hair conditioner containing bitter tea holly leaf extract.

[0163] Example 10: Hair conditioner containing bitter tea and holly leaf extract

[0164] By weight, the conditioner contains 5% bitter tea holly leaf extract, 6% behenyltrimethylammonium methyl sulfate, 3% behenyltrimethylammonium chloride, 5% cetyl alcohol, 2.5% bis-aminopropyl polydimethylsiloxane, 1% squalane, 0.5% 1,2-hexanediol, 0.5% p-hydroxyacetophenone, and 0.1% EDTA-2Na, with the balance being water.

[0165] The specific preparation method is as follows:

[0166] a. Add purified water, bitter tea holly leaf extract, 1,2-hexanediol, p-hydroxyacetophenone, and EDTA-2Na to an emulsifying pot, turn on the stirrer, stir at 25-50 rpm, and heat to 80 ℃ (appropriate homogenization can help with dispersion).

[0167] b. Add behenyltrimethylammonium methyl sulfate, behenyltrimethylammonium chloride, cetyl alcohol, squalane, etc. into the oil phase pot and heat to 80 ℃, then add to the emulsification pot. Homogenize at 2600-3000 rpm for 5-8 min and keep warm for 15-20 min.

[0168] c. Cool to 65-70℃ and keep warm. Add bis-aminopropylpolydimethylsiloxane, keep warm and stir evenly for 15-20 minutes. Adjust the pH value with citric acid.

[0169] d. Continue cooling to below 40 ℃ and remove from the pot to prepare a hair conditioner containing bitter tea holly leaf extract.

[0170] Example 11: Hair conditioner containing bitter tea and holly leaf extract

[0171] By weight, the conditioner contains 3% bitter tea holly leaf extract, 2.5% behenyltrimethylammonium methyl sulfate, 0.5% behenyltrimethylammonium chloride, 3% stearyl alcohol, 3.5% bis-aminopropyl polydimethylsiloxane, 0.15% methyl methacrylate, 0.4% phenoxyethanol, 0.1% ethylhexylglycerin, 0.3% sodium citrate, and 1% D-panthenol, with the balance being water.

[0172] The specific preparation method is as follows:

[0173] a. Add purified water, bitter tea holly leaf extract, methyl ester, phenoxyethanol, ethylhexylglycerin, D-panthenol, and sodium citrate to an emulsifying pot, turn on the stirrer, stir at 25-50 rpm, and heat to 80 ℃ (appropriate homogenization can help with dispersion).

[0174] b. Add behenyltrimethylammonium methyl sulfate, behenyltrimethylammonium chloride, stearyl alcohol, etc. to the oil phase pot and heat to 80°C, then add to the emulsification pot. Homogenize at 2600-3000 rpm for 5-8 minutes, and hold at this temperature for 15-20 minutes.

[0175] c. Cool to 65-70℃ and keep warm. Add bis-aminopropylpolydimethylsiloxane, keep warm and stir evenly for 15-20 minutes. Adjust the pH value with citric acid.

[0176] d. Continue cooling to below 40 ℃ and remove from the pot to prepare a hair conditioner containing bitter tea holly leaf extract.

[0177] Example 12: Hair conditioner containing bitter tea and holly leaf extract

[0178] By weight, the conditioner contains 0.1% bitter tea holly leaf extract, 3% behenyltrimethylammonium methyl sulfate, 2% behenyltrimethylammonium chloride, 3% cetyl alcohol, 0.5% bis-aminopropylpolydimethylsiloxane, 3% squalane, 0.15% sea buckthorn fruit oil, 0.4% sodium benzoate, 0.3% phenoxyethanol, 0.1% ethylhexylglycerin, 0.5% citric acid, and 5% glycerin, with the balance being water.

[0179] The specific preparation method is as follows:

[0180] a. Add purified water, bitter tea holly leaf extract, sodium benzoate, phenoxyethanol, ethylhexylglycerin, glycerin, and citric acid to an emulsifying pot, turn on the stirrer, stir at 25-50 rpm, and heat to 80 ℃ (homogeneous mixing can be used to aid dispersion).

[0181] b. Add behenyltrimethylammonium methyl sulfate, behenyltrimethylammonium chloride, cetyl alcohol, squalane, sea buckthorn fruit oil, etc. into the oil phase pot, heat to 80 ℃ and then add to the emulsification pot. Homogenize at 2600-3000 rpm for 5-8 min and keep warm for 15-20 min.

[0182] c. Cool to 65-70℃ and keep warm. Add bis-aminopropylpolydimethylsiloxane and keep warm while stirring until homogeneous. Control the time for 15-20 minutes. Adjust the pH value with citric acid.

[0183] d. Continue cooling to below 40 ℃ and remove from the pot to prepare a hair conditioner containing bitter tea holly leaf extract.

[0184] Example 13: Analysis of the effects of hair conditioner containing bitter tea and holly leaf extract

[0185] The hair conditioners prepared in Examples 9-12 were tested and compared.

[0186] 13.1 Moisturizing performance test

[0187] The moisture permeation cup test method was adopted, referring to the group standard T / CSBME 068-2023. The hair conditioner samples to be tested (Examples 9-12) were added to the moisture permeation cup device, placed in a constant temperature and humidity test chamber for 24 hours, and the test was carried out. The moisturizing performance of each sample was calculated according to the following calculation formula.

[0188]

[0189] Note: In the formula: the weight unit is grams (g); A is the weight loss after 24 hours after sample addition; B is the weight loss of the sample itself after 24 hours; C is the weight loss before sample addition after 24 hours.

[0190] The results are shown in Table 16. It can be seen that the four hair conditioners have good moisturizing properties, among which the hair conditioner preparation of Example 9 has the best moisturizing properties, with a moisturizing rate of 66.26%.

[0191] Table 16

[0192]

[0193] 13.2 Repair Performance Test

[0194] The degree of hair damage was observed using a trichoscope to examine the microstructure of the hair surface. Healthy, undamaged hair showed neatly arranged and smooth cuticles; damaged hair showed raised and broken cuticles. After using the conditioner prepared in Examples 9-12 for a period of time (28 days), the repair of the hair cuticles was observed. If the cuticles were more neatly arranged and less raised, it indicated a repairing effect. Results are as follows... Figure 10 As shown, after using the conditioner products for 28 days, compared with before use, the conditioner products in each example all improved the hair damage to a certain extent. Before use, the hair cuticles were noticeably raised, with irregular edges, loosely arranged like roof tiles, and some were even missing or broken. The hair surface was rough and uneven, with poor reflectivity. The open cuticles caused light scattering, making the hair appear dull and lifeless. In Examples 9-12, the cuticle edges became smoother, the raised portion decreased, and the cuticles were more tightly closed. The hair surface smoothness improved, reflectivity increased, the feel became softer, and the cuticle flexibility improved, making it less prone to further peeling due to external force. Among Examples 9-12, Example 9 showed the best results.

[0195] 13.3 System Test of Hair Loss Prevention and Hair Growth Effect

[0196] An experiment was conducted to evaluate the efficacy of Kuding tea and wintergreen hair conditioner in a mouse model of androgenetic alopecia. Male C57BL / 6J mice, 7 weeks old and weighing approximately 22 g, were purchased from the Beijing Vital River Laboratory Animal Center. The C57BL / 6J mice were housed under normal conditions: temperature 25±2℃, humidity 50%–70%, alternating between light and dark for 12 hours each, with free access to food and water, and regular bedding changes. The experiment began after the mice had adapted to their enclosure. The androgenetic alopecia model was established using C57BL / 6J mice. Mice were anesthetized by intraperitoneal injection of 100 μL of 4% chloral hydrate. All hair on the back of the mice was shaved using a razor, and then treated with commercially available depilatory wax (area 2 cm × 2.5 cm). Thirty male C57BL / 6J mice were randomly divided into six groups: control group, model group, Example 9 group, Example 10 group, Example 11 group, and Example 12 group. Except for the control group, the other groups were treated daily with 200 μL of testosterone (5 mg / mL) for modeling. After complete absorption, 200 μL of hair conditioner sample was applied evenly for 14 consecutive days. Hair growth on the backs of the mice was recorded on day 14. The experimental results are as follows: Compared with the control group, the hair density in the model area of ​​the model group was significantly reduced, indicating successful modeling. Compared with the model group, Examples 9-12 all showed anti-hair loss effects, such as... Figure 11 As shown in the figure. Among them, the hair loss prevention effect of the hair conditioner in Example 9 is significantly better than that of the other three examples, while the effect of Example 11 is the worst among the four hair conditioners.

[0197] 13.4 Determination of the irritant properties of Kuding tea wintergreen hair conditioner using the zein method

[0198] Zein, due to its unique amino acid structure, is almost insoluble in water. When zein interacts with surfactants, the protein structure changes, increasing its solubility in water. The stronger the interaction, the more significant the increase in water solubility, reflecting the greater the skin irritation of the surfactant. Based on this, this method was used to compare the irritation of four hair conditioner products in Examples 9-12. Each hair conditioner product was diluted to a 10% aqueous solution, and the pH was adjusted to 4-6 with citric acid. 50 mL of the sample solution was taken, and 0.5 g of zein was added to each. After magnetic stirring at room temperature for 2 hours, the mixture was allowed to stand and centrifuged. The undissolved zein was placed in a 40°C oven and dried for 3 days. The mass of the dried zein was weighed, and the mass of dissolved zein and the zein value were calculated. A higher zein value indicates a stronger irritation.

[0199] The calculation formula is: Zein value (g / g) = (ab) / (c×V)

[0200] Where a is the mass of Zein before the experiment, b is the mass of Zein after drying, c is the concentration of the sample aqueous solution, and V is the sample volume / mass.

[0201] The results are shown in Table 17. It can be seen that the conditioners in the four examples have very low irritation and all fall into the category of non-irritating.

[0202] Table 17

[0203]

[0204] The embodiments of the present invention have been described in detail above, but the content described is only a preferred embodiment of the present invention and should not be considered as limiting the scope of the present invention. All equivalent changes and improvements made within the scope of the present invention should still fall within the patent coverage of the present invention.

Claims

1. A shampoo containing extracts of bitter tea and holly leaves, characterized in that: It includes 0.05%-10% by weight of Kuding tea holly leaf extract; the Kuding tea holly leaves are extracted by decoction, 70% ethanol is added to the aqueous extract of Kuding tea holly leaves to remove total polysaccharides, the separated liquid fraction is loaded onto an adsorption resin column, eluted with 30% ethanol to remove total phenolic acids, and then the eluted fraction with 50% ethanol is collected, concentrated and dried to obtain total saponins of Kuding tea holly leaves, and the obtained solid substance is Kuding tea holly leaf extract. It also includes 0.01-1% nucleating agent, 10-20% surfactant, 1.5-3% thickener, 0.1-8% conditioning agent, 0.01-2% preservative and 0.1-0.5% chelating agent, with the balance being water; wherein the nucleating agent is selected from one or more of ceramides, sphingolipids, cholesterol and phytosphingosine.

2. The shampoo containing bitter tea holly leaf extract according to claim 1, characterized in that: The surfactant is selected from one or more of ammonium lauryl ether sulfate, ammonium lauryl sulfate, sodium lauryl ether sulfate, and TEA salt of dodecylbenzene sulfonate; The thickener is selected from one or more of cocamide MEA, cocamide methyl MEA, sodium chloride, and cetyl alcohol; The conditioning agent is selected from one or more of polyquaternium-39, polyquaternium-10, guar hydroxypropyltrimethylammonium chloride, and emulsified silicone oil.

3. A method for preparing the shampoo containing holly leaf extract of bitter tea as described in claim 1 or 2, characterized in that: Weigh each component by weight percentage, mix and stir until emulsified to obtain a shampoo containing bitter tea holly leaf extract.

4. A hair conditioner containing extracts of bitter tea and holly leaves, characterized in that: It includes 0.05%-10% by weight of Kuding tea holly leaf extract; the Kuding tea holly leaves are extracted by decoction, 70% ethanol is added to the aqueous extract of Kuding tea holly leaves to remove total polysaccharides, the separated liquid fraction is loaded onto an adsorption resin column, eluted with 30% ethanol to remove total phenolic acids, and then the eluted fraction with 50% ethanol is collected, concentrated and dried to obtain total saponins of Kuding tea holly leaves, and the obtained solid substance is Kuding tea holly leaf extract. It also includes 3-11% surfactant, 3-5% thickener, 3.5-4% hair conditioner and 0.55-1% preservative, with the balance being water.

5. The hair conditioner containing holly leaf extract from bitter tea according to claim 4, characterized in that: The surfactant is behenyltrimethylammonium methyl sulfate and / or behenyltrimethylammonium chloride; The thickener is cetyl alcohol and / or stearyl alcohol; The hair conditioning agent is one or more of bis-aminopropylpolydimethylsiloxane, squalane, and sea buckthorn fruit oil; The preservative is one or more of the following: methyl ester, sodium benzoate, phenoxyethanol, ethylhexylglycerin, 1,2-hexanediol, and p-hydroxyacetophenone.

6. A method for preparing the hair conditioner containing holly leaf extract of bitter tea as described in claim 4 or 5, characterized in that: Weigh each component by weight percentage, mix and stir until emulsified to obtain a hair conditioner containing bitter tea holly leaf extract.

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