A hair loss prevention and blackening hair liquid for turning white hair black and its preparation method

Abstract

The present invention relates to the technical field of hair treatment drugs, and specifically discloses an anti-hair loss and blackening hair liquid for turning white hair into black hair and a preparation method thereof. The raw materials of the above-mentioned anti-hair loss and blackening hair liquid for turning white hair into black hair include, by mass: 5-10 parts of kaempferol, 1-5 parts of geranic acid, 1-2 parts of lecithin, 1-2 parts of surfactant, 40-60 parts of vegetable oil, 1-3 parts of copper tripeptide, 1-2 parts of dopamine hydrochloride, 0.01-0.1 part of 1-ethyl-(3-dimethylaminopropyl)carbodiimide, and 1-2 parts of traditional Chinese medicine extract. The present invention simultaneously loads hydrophilic and lipophilic active components, effectively promotes the repair and development of melanocytes, and can strengthen the connection between the dermal papilla and blood vessels, with significant hair growth effect, small irritation, capable of deeply nourishing hair follicles, and not generating dependence, being suitable for large-scale popularization and use.

Description

Technical Field

[0001] The present invention relates to the technical field of hair treatment drugs, and particularly relates to an anti - hair - loss and black - hair - promoting liquid for white hair and its preparation method. Background Art

[0002] Hair not only plays an important role in human appearance, but is also one of the important indicators of human health. The color of hair depends on the number, size, distribution of pigment granules in the hair shaft and the nature of the contained pigments. White hair refers to hair that is completely or partially white. The appearance of white hair is due to the reduction or filling of air in the melanin granules in the hair medulla and cortex.

[0003] Normally shed hair is hair in the telogen and exogen phases. The hair in the telogen and the newly entered anagen phase are constantly in a dynamic balance. Pathological white - hair and hair - loss is a common skin disease, referring to abnormal whitening or excessive shedding of hair. Affected by factors such as a busy life, work pressure, and environmental pollution, more and more modern people have problems such as white hair.

[0004] Currently, there are many cases of itchy scalp, severe hair loss, and a sharp increase in white hair. There are many treatment methods. Existing hair transplantation surgeries have side effects such as infection, damage to superficial sensory nerves, and scar formation. Currently, for hair dyes and anti - hair - loss shampoos for the head, their actual efficacy is unstable. Some not only have no protective effect but instead have a harmful effect, bringing inconvenience to patients, so that baldness is becoming younger and younger, and white - haired teenagers are increasing.

[0005] Although there are some drugs for treating white - hair and hair - loss on the market currently, most of them are traditional Chinese medicine compound prescriptions, with unclear medicinal ingredients, weak targeting, and it is difficult to achieve effective treatment effects. At the same time, it is easy to relapse after stopping the drug, and the effect is not ideal. Summary of the Invention

[0006] The purpose of the present invention is to solve the deficiencies in the prior art and propose an anti - hair - loss and black - hair - promoting liquid for white hair and its preparation method.

[0007] An anti - hair - loss and black - hair - promoting liquid for white hair, the raw materials of which include, by mass: 5 - 10 parts of kaempferol, 1 - 5 parts of geranic acid, 1 - 2 parts of lecithin, 1 - 2 parts of surfactant, 40 - 60 parts of vegetable oil, 1 - 3 parts of copper - peptide, 1 - 2 parts of dopamine hydrochloride, 0.01 - 0.1 part of 1 - ethyl - (3 - dimethylaminopropyl) carbodiimide, and 1 - 2 parts of traditional Chinese medicine extract.

[0008] Preferably, the surfactant is at least one of polyoxyethylene sorbitan fatty acid ester, sorbitan fatty acid ester, fatty acid glyceride, and cocamidopropyl betaine.

[0009] Preferably, the vegetable oil is at least one of peanut oil, rapeseed oil, soybean oil, corn oil, camellia oil, walnut oil, olive oil, palm oil, grape seed oil, and sunflower oil.

[0010] Preferably, the traditional Chinese medicine extract is at least one of mulberry fruit extract, licorice root extract, fleece-flower root extract, rehmannia root extract, biota orientalis leaf extract, black sesame extract, eclipta prostrata extract, dandelion extract, mulberry leaf extract, ginseng root extract, chuanxiong root extract, and magnolia bark extract.

[0011] The fleece-flower root extract and black sesame extract are obtained by extracting with an ethanol aqueous solution having a system fraction of 70%; while the mulberry fruit extract, licorice root extract, rehmannia root extract, biota orientalis leaf extract, eclipta prostrata extract, dandelion extract, mulberry leaf extract, ginseng root extract, chuanxiong root extract, and magnolia bark extract are water extracts.

[0012] Preferably, the traditional Chinese medicine extract includes: mulberry fruit extract, fleece-flower root extract, rehmannia root extract, mulberry leaf extract, and ginseng root extract; the mass ratio of the mulberry fruit extract, fleece-flower root extract, rehmannia root extract, mulberry leaf extract, and ginseng root extract is 1-2:4-6:3-5:1-2:2-4.

[0013] The preparation method of the anti-hair loss and blackening hair liquid for gray hair includes the following steps:

[0014] S1. Add kaempferol to an ethanol aqueous solution, reflux and stir at 40-60 °C for 10-20 min, add geranic acid to it and continue stirring for 1-3 h, perform rotary evaporation, and vacuum dry to obtain a kaempferol composite.

[0015] S2. Add the kaempferol composite, lecithin, and surfactant to the vegetable oil and stir for 10-30 min, adjust the system temperature to 40-60 °C, and perform ultrasonic treatment for 10-20 min to obtain an oil phase.

[0016] S3. Add copper tripeptide to an acetic acid aqueous solution and stir evenly, add dopamine hydrochloride and 1-ethyl-(3-dimethylaminopropyl)carbodiimide to it, adjust the system pH value to 5.5-6, perform ultrasonic treatment for 1-2 h, dialyze for 5-10 h, freeze-dry, then mix with the traditional Chinese medicine extract, add it to water and stir evenly, and drop the oil phase into it under stirring and stir for 5-15 min.

[0017] Preferably, in S2, the ultrasonic frequency is 10-15 kHz.

[0018] Preferably, in S3, the concentration of the acetic acid aqueous solution is 0.01-0.05 mol / L.

[0019] Preferably, in S3, the ultrasonic frequency is 12 - 16 kHz.

[0020] Preferably, in S3, dialysis is performed for 5 - 10 h with an aqueous acetic acid solution having a concentration of 0.01 - 0.05 mol / L. Beneficial effects

[0021] The present invention uses kaempferol as an active ingredient, which can effectively reduce the inflammation of human dermal papilla cells and surrounding cells, and has a significant anti - hair - loss effect. It combines with geranic acid to form a synergistic effect, comprehensively improving the microenvironment around the hair follicles, and has a significant hair - blackening effect. When combined with lecithin, it can effectively penetrate into the scalp and hair follicles through interaction, effectively activate tyrosinase, repair damaged melanocytes, and has good hair - blackening persistence. Moreover, long - term medication has almost no side effects, and the hair - growth and hair - blackening effects are remarkable.

[0022] The present invention binds dopamine to the surface of blue copper peptide. It can not only be effectively dispersed at the oil - water interface, thus having a large interfacial area and small mass transfer resistance, which is more conducive to the transdermal absorption of active ingredients and has a significant anti - inflammatory effect, but also can effectively reduce sebum secretion, promote blood circulation, effectively improve the body's immunity, activate hair follicles to promote hair regrowth, and can well promote scalp absorption, achieving a good therapeutic effect.

[0023] The inventors of the present invention found through experiments that if only a simple mixture of blue copper peptide and dopamine is used as an active ingredient, not only is the hair - growth effect poor, but also the persistence is poor. However, through the organic combination of blue copper peptide and dopamine in the present invention, it not only has good biocompatibility, can effectively help active ingredients penetrate through the skin barrier, efficiently deliver active ingredients to the hair follicles, significantly promote hair growth, not only can extend the residence time, but also can comprehensively improve the microenvironment around the hair follicles, with good effects, and at the same time is not easy to relapse, and has a good therapeutic effect.

[0024] The present invention simultaneously loads hydrophilic and lipophilic active components, effectively promotes the repair and development of melanocytes, strengthens the connection between dermal papilla and blood vessels, has a significant hair - growth effect, little irritation, can deeply nourish hair follicles, does not produce dependence, and is suitable for large - scale popularization and use. Brief description of the drawings

[0025] Figure 1 It is a comparison chart of intracellular melanin content and tyrosinase activation rate for Example 5 and Comparative Example 1 - 2 groups.

[0026] Figure 2 It is a comparison chart of the number of apoptotic follicle cells and the number of melanin - containing follicles for the NC group, PC group, S5 group, D1 group, and D2 group.

[0027] Figure 3Comparison chart of the contents of inflammatory factors TNF-α and IL-6 secreted by macrophages RAW264.7 in Example 5, Comparative Example 1, Comparative Example 2, positive control, and negative control. Detailed implementation mode

[0028] The present invention will be further explained below with reference to specific embodiments.

[0029] The following polygonum multiflorum root extract, platycladus orientalis leaf extract, ligusticum chuanxiong root extract, and mulberry fruit extract are purchased from Shaanxi Fenglinggu Medical Biotechnology Co., Ltd., and the following rehmannia glutinosa extract, mulberry leaf extract, and ginseng root extract are purchased from Shaanxi Bolin Biotechnology Co., Ltd. Example 1

[0030] An anti-hair loss and hair blackening liquid for white hair to turn black hair, the raw materials of which include: 5 g of kaempferol, 1 g of angelic acid, 1 g of lecithin, 1 g of cocamidopropyl betaine, 40 g of camellia oil, 1 g of copper peptide, 1 g of dopamine hydrochloride, 0.01 g of 1-ethyl-(3-dimethylaminopropyl)carbodiimide, 0.3 g of rehmannia glutinosa extract, and 0.7 g of platycladus orientalis leaf extract.

[0031] The preparation method of the above anti-hair loss and hair blackening liquid for white hair to turn black hair includes the following steps:

[0032] S1. Add kaempferol to 30 g of an ethanol aqueous solution with a mass fraction of 40%, reflux and stir at a temperature of 40°C for 10 min, add angelic acid to it, continue to stir for 1 h, rotary evaporate, and vacuum dry to obtain a kaempferol composite;

[0033] S2. Add the kaempferol composite, lecithin, and cocamidopropyl betaine to camellia oil, stir at a speed of 100 r / min for 10 min, adjust the system temperature to 40°C, and perform ultrasonic treatment for 10 min with an ultrasonic frequency of 10 kHz to obtain an oil phase;

[0034] S3. Add copper peptide to 30 g of acetic acid aqueous solution with a concentration of 0.01 mol / L and stir evenly, add dopamine hydrochloride and 1-ethyl-(3-dimethylaminopropyl)carbodiimide to it, adjust the system pH value to 5.5 - 6, perform ultrasonic treatment for 1 h with an ultrasonic frequency of 12 kHz, dialyze with acetic acid aqueous solution with a concentration of 0.01 mol / L for 5 h, freeze-dry, then mix with rehmannia glutinosa extract and platycladus orientalis leaf extract, and then add it to 100 g of water and stir evenly. While stirring, drop the oil phase into it and stir at a speed of 1000 r / min for 5 min. Example 2

[0035] A hair loss prevention and hair darkening liquid for turning white hair black, the raw materials of which include: 10 g of kaempferol, 5 g of ethyl angelate, 2 g of lecithin, 2 g of glyceryl stearate, 60 g of grape seed oil, 3 g of copper peptide, 2 g of dopamine hydrochloride, 0.1 g of 1-ethyl-(3-dimethylaminopropyl) carbodiimide, 1.5 g of ginseng root extract, and 0.5 g of chuanxiong root extract.

[0036] The preparation method of the above-mentioned hair loss prevention and hair darkening liquid for turning white hair black includes the following steps:

[0037] S1. Add kaempferol to 60 g of an ethanol aqueous solution with a mass fraction of 60%, reflux and stir at a temperature of 60 °C for 20 min, add ethyl angelate thereto, continue stirring for 3 h, perform rotary evaporation, and vacuum dry to obtain a kaempferol composite material;

[0038] S2. Add the kaempferol composite material, lecithin, and glyceryl stearate to grape seed oil, stir at a speed of 400 r / min for 30 min, adjust the system temperature to 60 °C, and perform ultrasonic treatment for 20 min with an ultrasonic frequency of 15 kHz to obtain an oil phase;

[0039] S3. Add copper peptide to 60 g of an acetic acid aqueous solution with a concentration of 0.05 mol / L and stir evenly, add dopamine hydrochloride and 1-ethyl-(3-dimethylaminopropyl) carbodiimide thereto, adjust the system pH value to 5.5 - 6, perform ultrasonic treatment for 2 h with an ultrasonic frequency of 16 kHz, dialyze with an acetic acid aqueous solution with a concentration of 0.05 mol / L for 10 h, perform freeze-drying, then mix with ginseng root extract and chuanxiong root extract, and then add to 200 g of water and stir evenly. While stirring, dropwise add the oil phase and stir at a speed of 2000 r / min for 15 min. Example Three

[0040] A hair loss prevention and hair darkening liquid for turning white hair black, the raw materials of which include: 7 g of kaempferol, 4 g of ethyl angelate, 1.3 g of lecithin, 1.8 g of Tween 60, 45 g of palm oil, 2.5 g of copper peptide, 1.3 g of dopamine hydrochloride, 0.08 g of 1-ethyl-(3-dimethylaminopropyl) carbodiimide, and 1.3 g of traditional Chinese medicine extract.

[0041] The traditional Chinese medicine extract includes: 2 g of mulberry fruit extract, 4 g of polygonum multiflorum root extract, 5 g of rehmannia root extract, 1 g of mulberry leaf extract, and 4 g of ginseng root extract.

[0042] The preparation method of the above-mentioned hair loss prevention and hair darkening liquid for turning white hair black includes the following steps:

[0043] S1. Add kaempferol to 40 g of an ethanol aqueous solution with a mass fraction of 55%, reflux and stir at 45 °C for 18 min, add geranic acid to it, continue to stir for 1.5 h, perform rotary evaporation, and vacuum dry to obtain the kaempferol composite;

[0044] S2. Add the kaempferol composite, lecithin, and Tween 60 to palm oil, stir at a speed of 300 r / min for 15 min, adjust the system temperature to 55 °C, and perform ultrasonic treatment for 13 min with an ultrasonic frequency of 13 kHz to obtain the oil phase;

[0045] S3. Add copper tripeptide to 40 g of an acetic acid aqueous solution with a concentration of 0.04 mol / L and stir evenly, add dopamine hydrochloride and 1-ethyl-(3-dimethylaminopropyl)carbodiimide to it, adjust the system pH value to 5.5 - 6, perform ultrasonic treatment for 80 min with an ultrasonic frequency of 15.5 kHz, dialyze with an acetic acid aqueous solution with a concentration of 0.02 mol / L for 9 h, perform freeze-drying, then mix with the traditional Chinese medicine extract, add it to 120 g of water and stir evenly, and dropwise add the oil phase to it while stirring, and stir at a speed of 1700 r / min for 8 min. Example 4

[0046] An anti-hair loss and hair blackening liquid for white hair to turn black, the raw materials of which include: 9 g of kaempferol, 2 g of geranic acid, 1.7 g of lecithin, 1.2 g of Span 80, 55 g of palm oil, 1.5 g of copper tripeptide, 1.7 g of dopamine hydrochloride, 0.02 g of 1-ethyl-(3-dimethylaminopropyl)carbodiimide, and 1.7 g of the traditional Chinese medicine extract.

[0047] The traditional Chinese medicine extract includes: 1 g of mulberry fruit extract, 6 g of polygonum multiflorum root extract, 3 g of rehmannia root extract, 2 g of mulberry leaf extract, and 2 g of ginseng root extract.

[0048] The preparation method of the above anti-hair loss and hair blackening liquid for white hair to turn black includes the following steps:

[0049] S1. Add kaempferol to 50 g of an ethanol aqueous solution with a mass fraction of 45%, reflux and stir at 55 °C for 12 min, add geranic acid to it, continue to stir for 2.5 h, perform rotary evaporation, and vacuum dry to obtain the kaempferol composite;

[0050] S2. Add the kaempferol composite, lecithin, and Span 80 to palm oil, stir at a speed of 200 r / min for 25 min, adjust the system temperature to 45 °C, and perform ultrasonic treatment for 17 min with an ultrasonic frequency of 11 kHz to obtain the oil phase;

[0051] S3. Add the GHK-Cu peptide to 50 g of acetic acid aqueous solution with a concentration of 0.02 mol / L and stir evenly. Then add dopamine hydrochloride and 1-ethyl-(3-dimethylaminopropyl)carbodiimide to it, adjust the pH value of the system to 5.5 - 6, perform ultrasonic treatment for 100 min with an ultrasonic frequency of 13.5 kHz, dialyze with acetic acid aqueous solution with a concentration of 0.04 mol / L for 7 h, freeze-dry, then mix with the traditional Chinese medicine extract, and then add it to 180 g of water and stir evenly. While stirring, dropwise add the oil phase and stir at a speed of 1300 r / min for 12 min. Example 5

[0052] A hair loss prevention and black hair growth liquid for turning white hair black, the raw materials of which include: 8 g of kaempferol, 3 g of angelic acid, 1.5 g of lecithin, 1.5 g of Tween 80, 50 g of palm oil, 2 g of GHK-Cu peptide, 1.5 g of dopamine hydrochloride, 0.05 g of 1-ethyl-(3-dimethylaminopropyl)carbodiimide, and 1.5 g of traditional Chinese medicine extract.

[0053] The traditional Chinese medicine extract includes: 1.5 g of mulberry fruit extract, 5 g of polygonum multiflorum root extract, 4 g of rehmannia root extract, 1.5 g of mulberry leaf extract, and 3 g of ginseng root extract.

[0054] The preparation method of the above-mentioned hair loss prevention and black hair growth liquid for turning white hair black includes the following steps:

[0055] S1. Add kaempferol to 45 g of ethanol aqueous solution with a mass fraction of 50%, reflux and stir at a temperature of 50 °C for 15 min, add angelic acid to it, continue to stir for 2 h, perform rotary evaporation, and vacuum dry to obtain the kaempferol composite.

[0056] S2. Add the kaempferol composite, lecithin, and Tween 80 to palm oil, stir at a speed of 250 r / min for 20 min, adjust the system temperature to 50 °C, perform ultrasonic treatment for 15 min with an ultrasonic frequency of 12 kHz to obtain the oil phase.

[0057] S3. Add the GHK-Cu peptide to 45 g of acetic acid aqueous solution with a concentration of 0.03 mol / L and stir evenly. Then add dopamine hydrochloride and 1-ethyl-(3-dimethylaminopropyl)carbodiimide to it, adjust the pH value of the system to 5.5 - 6, perform ultrasonic treatment for 90 min with an ultrasonic frequency of 15 kHz, dialyze with acetic acid aqueous solution with a concentration of 0.03 mol / L for 8 h, freeze-dry, then mix with the traditional Chinese medicine extract, and then add it to 150 g of water and stir evenly. While stirring, dropwise add the oil phase and stir at a speed of 1500 r / min for 10 min.

[0058] Comparative Example 1

[0059] A hair loss prevention and hair blackening liquid for gray hair turning black, the raw materials of which include: 8 g of kaempferol, 3 g of ethyl angelate, 1.5 g of lecithin, 1.5 g of tween 80, 50 g of palm oil, 2 g of copper peptide, 1.5 g of dopamine hydrochloride, and 1.5 g of Chinese herbal medicine extract.

[0060] The Chinese herbal medicine extract includes: 1.5 g of mulberry fruit extract, 5 g of fleece-flower root extract, 4 g of rehmannia root extract, 1.5 g of mulberry leaf extract, and 3 g of ginseng root extract.

[0061] The preparation method of the above-mentioned hair loss prevention and hair blackening liquid for gray hair turning black includes the following steps:

[0062] S1. Add kaempferol to 45 g of an ethanol aqueous solution with a mass fraction of 50%, reflux and stir at a temperature of 50 °C for 15 min, add ethyl angelate to it, continue to stir for 2 h, perform rotary evaporation, and vacuum dry to obtain a kaempferol composite;

[0063] S2. Add the kaempferol composite, lecithin, and tween 80 to palm oil, stir at a speed of 250 r / min for 20 min, adjust the system temperature to 50 °C, and perform ultrasonic treatment for 15 min with an ultrasonic frequency of 12 kHz to obtain an oil phase;

[0064] S3. Mix copper peptide, dopamine hydrochloride with the Chinese herbal medicine extract, then add it to 150 g of water and stir evenly, and dropwise add the oil phase to it while stirring, and stir at a speed of 1500 r / min for 10 min.

[0065] Comparative Example 2

[0066] A hair loss prevention and hair blackening liquid for gray hair turning black, the raw materials of which include: 8 g of kaempferol, 3 g of ethyl angelate, 1.5 g of lecithin, 1.5 g of tween 80, 50 g of palm oil, 2 g of copper peptide, 1.5 g of dopamine hydrochloride, 0.05 g of 1-ethyl-(3-dimethylaminopropyl)carbodiimide, and 1.5 g of Chinese herbal medicine extract.

[0067] The Chinese herbal medicine extract includes: 1.5 g of mulberry fruit extract, 5 g of fleece-flower root extract, 4 g of rehmannia root extract, 1.5 g of mulberry leaf extract, and 3 g of ginseng root extract.

[0068] The preparation method of the above-mentioned hair loss prevention and hair blackening liquid for gray hair turning black includes the following steps:

[0069] S1. Add kaempferol, ethyl angelate, lecithin, and tween 80 to palm oil, stir at a speed of 250 r / min for 20 min, adjust the system temperature to 50 °C, and perform ultrasonic treatment for 15 min with an ultrasonic frequency of 12 kHz to obtain an oil phase;

[0070] S2. Add the copper tripeptide to 45 g of acetic acid aqueous solution with a concentration of 0.03 mol / L and stir evenly. Add dopamine hydrochloride and 1-ethyl-(3-dimethylaminopropyl) carbodiimide thereto, adjust the pH value of the system to 5.5 - 6, perform ultrasonic treatment for 90 min with an ultrasonic frequency of 15 kHz, dialyze with acetic acid aqueous solution with a concentration of 0.03 mol / L for 8 h, freeze-dry, then mix with the traditional Chinese medicine extract, and then add it to 150 g of water and stir evenly. Dropwise add the oil phase thereto under stirring, and stir at a speed of 1500 r / min for 10 min.

[0071] Use the anti-hair loss and hair blackening liquids obtained in Example 5 and Comparative Examples 1 - 2 to culture B16 melanoma cells, and detect the intracellular melanin content in each group after culture. Specifically as follows:

[0072] Adjust the density of the B16 melanoma cell suspension to 1×10 5 cells / mL, inoculate 1 mL per well in a 12-well plate to make the cell density 5×10 4 cells / well, and place it in an incubator (37℃, 5% CO2) for culture for 24 h. Take out the 12-well plate, aspirate the culture medium, add 1 mL of culture medium containing each group of samples (concentration is 1 mg / mL), and add 1 mL of blank culture medium to the blank control group and the zero adjustment group. Continue to place it in the incubator for culture for 12 h, and then aspirate the culture medium;

[0073] Add 1 mL of 1 μM α-MSH solution prepared with RPMI1640 culture medium to each well in the experimental group and the zero adjustment group, add 1 mL of culture medium without α-MSH to the blank group, and place it in the incubator for culture for 36 h. Wash twice with 1 mL of PBS respectively, add 100 μL of 1 M NaOH solution (containing 10% DMSO) to each well, and after water bath in a water bath at 80℃ for 1 h, transfer the cell lysates in each group in the 12-well plate to a 96-well plate, and measure the absorbance OD value of each well at 475 nm in an enzyme-linked immunosorbent assay instrument, and calculate the intracellular melanin content.

[0074] Intracellular melanin content = (OD1 - OD3) ÷ (OD2 - OD3) × 100%

[0075] Among them, OD1 is the absorbance value of the experimental group, OD2 is the absorbance value of the blank control group, and OD3 is the absorbance value of the zero adjustment group.

[0076] As Figure 1 shown, the intracellular melanin content in the group of Example 5 is the highest, which is better than that in the groups of Comparative Examples 1 - 2 (P < 0.05).

[0077] The tyrosinase activation rate of the anti - hair - loss and hair - blackening liquids obtained in Example 5 and Comparative Examples 1 - 2 was detected as follows: Using phosphate buffer as the solvent, 0.5 mg / mL L - tyrosine solution and 35 mg / mL tyrosinase solution were respectively prepared; using propylene glycol as the solvent, 1 mg / mL test sample (the anti - hair - loss and hair - blackening liquids obtained in Example 5 and Comparative Examples 1 - 2) solution was prepared.

[0078] Then, the following 4 groups of solutions were prepared:

[0079] A: 0.7 mL L - tyrosine solution + 0.7 mL pure propylene glycol + 0.6 mL phosphate buffer;

[0080] B: 0.7 mL pure propylene glycol + 1.3 mL phosphate buffer;

[0081] C: 0.7 mL L - tyrosine solution + 0.7 mL test sample solution + 0.6 mL phosphate buffer;

[0082] D: 0.7 mL test sample solution + 0.6 mL phosphate buffer.

[0083] After the above 4 groups of solutions were kept at a constant temperature in a water bath at 37 °C for 10 min, 0.3 mL of tyrosinase solution was added to each group. After reacting for 5 min, they were quickly transferred into cuvettes, and the absorbance value was measured at 475 nm, and the tyrosinase activation rate was calculated.

[0084]

[0085] Among them, (OD1 - OD2) represents the change in the OD value of the reaction solution under the condition of no sample, and (OD3 - OD4) represents the change in the OD value of the reaction solution under the condition of having a sample.

[0086] As Figure 1 shown, the tyrosinase activation rate of the group of Example 5 was the highest, superior to the groups of Comparative Examples 1 - 2 (P < 0.05).

[0087] A comparative test on the hair - growth and hair - blackening effects of the anti - hair - loss and hair - blackening liquids obtained in Example 5 and Comparative Examples 1 - 2 was carried out, and C57BL / 6 mice (half male and half female, clean - grade, 7 - 8 weeks old, weighing 20 - 22 g) were used as experimental subjects. The specific steps are as follows:

[0088] 1. Comparative test on hair - growth effect:

[0089] After all the mice were anesthetized, the hair on their backs was removed using paraffin - rosin, and the removal area was about 2 cm × 2 cm. They were randomly divided into 5 groups. Starting from the day after hair removal, the corresponding drugs were externally applied to the hair - removal area twice a day.

[0090] The NC group was smeared with distilled water, the PC group was smeared with minoxidil (2.0%, W / V), and the S5, D1, and D2 groups were smeared with the anti-hair loss liquid obtained in Example 5, the anti-hair loss liquid obtained in Comparative Example 1, and the anti-hair loss liquid obtained in Comparative Example 2, respectively.

[0091] 15 days after depilation, all mice were killed by cervical dislocation. The skin of the same part of the depilated area was fixed with 10% formaldehyde, dehydrated, embedded in paraffin, sliced, and spread on poly-lysine slides. In situ hybridization technology and terminal labeling were used to detect the growth / apoptosis of cells in the hair follicles.

[0092] 2. Comparative test of black hair effect:

[0093] After all mice were anesthetized, paraffin-rosin was used to remove the hair on their backs, and the removal area was about 2cm×2cm. 12h after depilation, mice without wounds on the back were selected, and a 6% hydrogen peroxide solution was applied to the depilated area twice a day for 7 days. The successful modeling criteria were: the mice had no abnormal performance, white hair grew on the surface of the back skin, and there was no black hair on the back, no black skin and wounds.

[0094] The mice were randomly divided into 5 groups. The corresponding drugs were applied to the light-colored hair growth area from the next day after the model was successfully established, twice a day. The NC group was applied with distilled water, the PC group was applied with minoxidil (2.0%, W / V), and the S5, D1, and D2 groups were applied with the anti-hair loss liquid obtained in Example 5, the anti-hair loss liquid obtained in Comparative Example 1, and the anti-hair loss liquid obtained in Comparative Example 2, respectively.

[0095] 15 days after successful modeling, all mice were killed by cervical dislocation. The skin of the same part of the depilated area was fixed with 10% formaldehyde, dehydrated, and embedded in paraffin. Two longitudinal and cross-sections of the skin were taken for each slice, and HE staining was performed respectively. The distribution of melanin in each group in the longitudinal section was observed under a microscope.

[0096] like Figure 2 As shown, the number of apoptotic hair follicle cells in group S5 was the least, while the number of melanin hair follicles was the largest, which was better than that in groups D1 and D2 (P < 0.05) and also better than that in group PC (P < 0.05).

[0097] The anti-hair loss and blackening liquid obtained in Example 5 and Comparative Examples 1-2 were used to conduct anti-inflammatory experiments, as follows:

[0098] Mouse macrophage RAW264.7 cells were cultured at 2×10 5The cell concentration per well was subcultured in a 24-well plate and cultured in DMEM medium at 37°C and 5% CO2 for 24 h. Then, the anti-hair loss and hair blackening liquids obtained in Example 5 and Comparative Examples 1-2 were added at a concentration of 1 mg / mL, and 100 μg / mL dexamethasone was added to the positive control group. 30 min after adding the samples, LPS was added to a final concentration of 1 μg / mL, and the cells were cultured at 37°C for 20 h. The culture medium was collected, and the secreted IL-6 and TNF-α were quantified using a mouse IL-6 ELISA kit and a mouse TNF-α ELISA kit, respectively.

[0099] As Figure 3 shown, compared with the negative control, the secretion levels of the inflammatory factors IL-6 and TNF-α in macrophages RAW264.7 were significantly decreased under 100 μg / mL dexamethasone in the positive control (P < 0.05), indicating that the positive control test was effective in this study. The anti-hair loss and hair blackening liquid obtained in Example 5 had similar secretion levels of the inflammatory factors IL-6 and TNF-α in macrophages RAW264.7 to those in the positive control (P > 0.05), and was superior to Comparative Examples 1-2 (P < 0.05), confirming that the anti-hair loss and hair blackening liquid obtained in Example 5 could inhibit the secretion of the inflammatory factors TNF-α and IL-6 in macrophages RAW264.7.

[0100] The applicant believes that the reason for the above results is that the present invention uses kaempferol as the active ingredient, which can effectively reduce the inflammation of human dermal papilla cells and surrounding cells, and has a significant anti-hair loss effect. It combines with geranic acid to form a synergistic effect, comprehensively improving the microenvironment around the hair follicles, and has a significant hair blackening effect. With the cooperation of lecithin, it can effectively penetrate into the scalp and hair follicles through interaction, effectively activate tyrosinase, repair damaged melanocytes, and has good hair blackening persistence. The present invention also binds dopamine to the surface of the blue copper peptide, which can not only be effectively dispersed at the oil-water interface, thus having a large interfacial area and a small mass transfer resistance, good biocompatibility, effectively helping the active ingredients to penetrate through the skin barrier, being more conducive to the transdermal absorption of the active ingredients, enabling the active ingredients to be efficiently delivered to the hair follicles, having a significant anti-inflammatory effect, but also effectively reducing sebum secretion, promoting blood circulation, effectively improving the body's immunity, activating hair follicles to promote hair regrowth, and being able to well promote scalp absorption, achieving a good therapeutic effect.

[0101] The above is only a preferred specific embodiment of the present invention, but the protection scope of the present invention is not limited thereto. Any person skilled in the art within the technical scope disclosed by the present invention, according to the technical solution and inventive concept of the present invention, makes equivalent substitutions or changes, and all should be covered within the protection scope of the present invention.

Claims

1. A hair loss prevention and blackening hair liquid for turning white hair black, characterized in that, Its raw materials by mass include: 5-10 parts of kaempferol, 1-5 parts of geranic acid, 1-2 parts of lecithin, 1-2 parts of surfactant, 40-60 parts of vegetable oil, 1-3 parts of copper tripeptide, 1-2 parts of dopamine hydrochloride, 0.01-0.1 part of 1-ethyl-(3-dimethylaminopropyl)carbodiimide, and 1-2 parts of traditional Chinese medicine extract; The traditional Chinese medicine extract includes: mulberry fruit extract, polygonum multiflorum root extract, rehmannia root extract, mulberry leaf extract, and ginseng root extract; It is prepared by the following steps: S1. Add kaempferol to an ethanol aqueous solution, reflux and stir at 40-60 °C for 10-20 min, add geranic acid to it and continue stirring for 1-3 h, rotary evaporate, and vacuum dry to obtain a kaempferol composite; S2. Add the kaempferol composite, lecithin, and surfactant to vegetable oil and stir for 10-30 min, adjust the system temperature to 40-60 °C, and ultrasonically treat for 10-20 min to obtain an oil phase; S3. Add copper tripeptide to an acetic acid aqueous solution and stir evenly, add dopamine hydrochloride and 1-ethyl-(3-dimethylaminopropyl)carbodiimide to it, adjust the system pH value to 5.5-6, ultrasonically treat for 1-2 h, dialyze for 5-10 h, freeze-dry, then mix with the traditional Chinese medicine extract, add it to water and stir evenly, and dropwise add the oil phase to it under stirring and stir for 5-15 min.

2. The anti-hair loss and blackening hair liquid for treating white hair to black hair according to claim 1, wherein The surfactant is at least one of polyoxyethylene sorbitan fatty acid ester, sorbitan fatty acid ester, fatty acid glyceride, and cocamidopropyl betaine.

3. The anti - hair - loss and black - hair - promoting liquid for gray hair turning black, according to claim 1, is characterized in that, The vegetable oil is at least one of peanut oil, rapeseed oil, soybean oil, corn oil, camellia oil, walnut oil, olive oil, palm oil, grape seed oil, and sunflower oil.

4. The anti-hair loss and blackening hair liquid for treating white hair to black hair according to claim 1, wherein The mass ratio of mulberry fruit extract, polygonum multiflorum root extract, rehmannia root extract, mulberry leaf extract, and ginseng root extract is 1-2:4-6:3-5:1-2:2-4.

5. A method for preparing an anti-hair loss and blackening hair liquid for turning white hair black as described in any one of claims 1-4, characterized in that, It includes the following steps: S1. Add kaempferol to an ethanol aqueous solution, reflux and stir at 40-60 °C for 10-20 min, add geranic acid to it and continue stirring for 1-3 h, rotary evaporate, and vacuum dry to obtain a kaempferol composite; S2. Add the kaempferol composite, lecithin, and surfactant to vegetable oil and stir for 10-30 min, adjust the system temperature to 40-60 °C, and ultrasonically treat for 10-20 min to obtain an oil phase; S3. Add copper tripeptide to an acetic acid aqueous solution and stir evenly, add dopamine hydrochloride and 1-ethyl-(3-dimethylaminopropyl)carbodiimide to it, adjust the system pH value to 5.5-6, ultrasonically treat for 1-2 h, dialyze for 5-10 h, freeze-dry, then mix with the traditional Chinese medicine extract, add it to water and stir evenly, and dropwise add the oil phase to it under stirring and stir for 5-15 min.

6. The preparation method of the anti - hair - loss and black - hair - growing liquid for white hair as claimed in claim 5, wherein, In S2, the ultrasonic frequency is 10-15 kHz.

7. The preparation method of the anti - hair - loss and black - hair - promoting liquid for gray hair to turn black, characterized in that, In S3, the concentration of the acetic acid aqueous solution is 0.01-0.05 mol / L.

8. The preparation method of the anti - hair - loss and black - hair - promoting liquid for gray hair as claimed in claim 5, characterized in that, In S3, the ultrasonic frequency is 12-16 kHz.

9. The preparation method of the anti - hair - loss and black - hair - promoting liquid for white hair according to claim 5, characterized in that, In S3, dialysis is carried out for 5-10 h using an acetic acid aqueous solution with a concentration of 0.01-0.05 mol / L.

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