Complex extract liposome containing evodia rutaecarpa bentham-broccoli complex extract and manufacturing method thereof

Abstract

The present invention relates to a complex extract liposome containing an Evodia rutaecarpa Bentham-broccoli complex extract and a manufacturing method thereof and, more specifically, to a complex extract liposome containing an Evodia rutaecarpa Bentham-broccoli complex extract and a manufacturing method thereof, wherein the complex extract liposome is manufactured by highly concentrating the Evodia rutaecarpa Bentham-broccoli complex extract into liposomes, thereby exhibiting excellent anti-inflammatory effects in hair root cells and thus exhibiting hair loss alleviation effects.

Description

Composite extract liposome containing a complex extract of sesame oil and broccoli and a method for preparing the same

[0001] The present invention relates to a complex extract liposome containing a complex extract of Osuyu-broccoli and a method for producing the same, and more particularly, to a complex extract liposome containing an Osuyu-broccoli complex extract which exhibits excellent anti-inflammatory effects in hair follicle cells and thus exhibits hair loss improvement effects by producing a complex extract liposome by highly concentrating an Osuyu-broccoli complex extract into liposomes, and a method for producing the same.

[0002] In general, the causes of hair loss include genetics, male hormones, stress, anemia, asthma, high blood pressure, arteriosclerosis, diabetes, and diseases such as modern society's eating habits and environmental pollution.

[0003] The most common cause of alopecia areata, caused by immune system disorders, is stress. Recently, women and children with alopecia areata complaining of this symptom account for approximately 40% of the overall hair loss population, making a solution urgently needed. Furthermore, the physical changes caused by hair loss also come with the side effect of exacerbating stress, which exacerbates symptoms.

[0004] As such, the number of people suffering from hair loss continues to increase regardless of gender or age due to environmental pollution, side effects from drug abuse, and social and environmental stress, and the need for the development of products using harmless natural substances that can prevent hair loss and promote hair growth is emerging.

[0005] The purpose of the present invention is to provide a complex extract liposome containing a complex extract of Osuyu-broccoli, which exhibits excellent anti-inflammatory effects in hair follicle cells and thus exhibits hair loss improvement effects by highly concentrating the complex extract of Osuyu-broccoli into liposomes to produce a complex extract liposome, and a method for producing the same.

[0006] A method for producing a complex extract liposome containing a complex extract of osuyu-broccoli according to an embodiment of the present invention comprises the steps of: (a) preparing a complex extract of osuyu and a broccoli extract; (b) mixing and stirring the complex extract of osuyu and a broccoli extract to form a complex extract of osuyu-broccoli; (c) diluting the complex extract of osuyu-broccoli with purified water to form a complex extract dilution; and (d) mixing a liposome base with the complex extract dilution of osuyu-broccoli and stirring to form a complex extract liposome of osuyu-broccoli.

[0007] In the step (a), the method comprises the steps of: washing the dried sorghum and then drying it with hot air; crushing the dried sorghum to obtain dried sorghum powder; adding the dried sorghum powder to a solvent and performing a first extraction to form a dried sorghum extract; and filtering the first extracted dried sorghum extract and then performing a second extraction using a rotary concentrate to form a dried sorghum extract.

[0008] In the step (a), the broccoli extract comprises: a step of washing broccoli and then drying it with hot air; a step of crushing the hot air-dried broccoli to obtain broccoli powder, a step of adding the broccoli powder to a solvent and performing a first extraction to form a broccoli extract; and a step of filtering the first-extracted broccoli extract and then performing a second extraction using a rotary concentrate to form a broccoli extract.

[0009] The above solvent is selected from water, methanol, ethanol, phenoxyethanol, propylene glycol, and dipropylene glycol, or a mixture of two or more thereof.

[0010] In the above step (b), the oil extract and broccoli extract are mixed in a weight ratio of 3:1 to 1:3.

[0011] In the above step (c), the diluted solution of the oil-soluble-broccoli complex extract is prepared by diluting the oil-soluble-broccoli complex extract in purified water at a weight ratio of 1:8 to 1:12.

[0012] In the above step (d), the liposome base is mixed in an amount of 50 to 150 parts by weight per 100 parts by weight of the diluted extract of the oil-broccoli complex.

[0013] In the step (d), the liposome base contains 70 to 90 wt% of a moisturizer, 10 to 30 wt% of an emulsifier, and 0.1 to 0.5 wt% of an oil.

[0014]

[0015] The complex extract liposome containing the complex extract of Osuyu-broccoli according to an embodiment of the present invention is characterized in that it exhibits an anti-inflammatory effect in hair follicle cells and thus has a hair loss improvement effect by being manufactured by concentrating the complex extract of Osuyu-broccoli into liposomes.

[0016] The complex extract liposome containing the complex extract of Osuyu-broccoli according to the present invention and the method for preparing the same were prepared by diluting the complex extract of Osuyu-broccoli in purified water, adding it to a liposome base, and stirring at room temperature.

[0017] In this way, the complex extract liposome containing the Osuyu-broccoli complex extract according to the present invention and the method for producing the same were prepared by highly concentrating the Osuyu-broccoli complex extract into liposomes, thereby confirming a concentration that does not show toxicity and confirming that β-catenin is expressed and that inflammatory cytokines are expressed.

[0018] As a result, the complex extract liposome containing the Osuyu-broccoli complex extract according to the present invention and the method for producing the same can exhibit excellent anti-inflammatory effects in hair follicle cells and thus exhibit hair loss improvement effects by producing the complex extract liposome by highly concentrating the Osuyu-broccoli complex extract into liposomes.

[0019] Figure 1 is a process flow diagram showing a method for manufacturing a complex extract liposome containing a complex extract of sesame oil and broccoli according to an embodiment of the present invention.

[0020] Figure 2 is a graph showing the results of absorbance measurements at 490 nm for broccoli, osuyu, and broccoli + osuyu.

[0021] Figure 3 is a graph showing the results of absorbance measurements at 490 nm for liposomes and broccoli+fructose liposomes.

[0022] Figure 4 shows the results of confirming β-catenin by confirming the band using ImageQuant 800 for broccoli, osuyu, broccoli + osuyu, liposome, and broccoli + osuyu liposome.

[0023] Figure 5 shows the results of confirming the JAK / STAT pathway by confirming the bands using ImageQuant 800 for broccoli, osuyu, broccoli + osuyu, liposome, and broccoli + osuyu liposome.

[0024] Figure 6 shows the results of confirming the MAPK pathway by confirming the bands using ImageQuant 800 for broccoli, osuyu, broccoli + osuyu, liposome, and broccoli + osuyu liposome.

[0025] Figure 7 is a graph showing the results of measuring the level of cytokines using an ELISA kit for broccoli, osuyu, broccoli + osuyu, liposome, and broccoli + osuyu liposome.

[0026] Figure 8 shows the results of a quality test on the total aerobic bacterial count for a scalp care hair shampoo composition containing a liposome containing a complex extract of sesame oil and broccoli.

[0027] Figure 9 shows the results of a quality test for non-detection of lead / arsenic heavy metals for a scalp care hair shampoo composition containing a liposome of a complex extract of Osuyu and broccoli.

[0028] Figure 10 shows the results of a pH quality test for a scalp care hair shampoo composition containing a liposome of a complex extract of sesame oil and broccoli.

[0029] Figure 11 shows the results of a quality test for the absence of E. coli for a scalp care hair shampoo composition containing a liposome of a complex extract of Osuyu and broccoli.

[0030] Figure 12 shows the results of a quality test for the non-detection of Staphylococcus aureus for a scalp care hair shampoo composition containing a liposome of a complex extract of Osuyu and broccoli.

[0031] Figure 13 shows the results of a quality test for non-detection of formaldehyde for a scalp care hair shampoo composition containing a liposome of a complex extract of sesame oil and broccoli.

[0032] Hereinafter, preferred embodiments of the present invention and the properties of each component will be described in detail. However, this is intended to be a detailed description to enable a person having ordinary knowledge in the technical field to which the present invention pertains to easily carry out the invention, and does not mean that the technical idea and scope of the present invention are limited thereby.

[0033]

[0034] A complex extract liposome containing a complex extract of Osuyu-broccoli according to an embodiment of the present invention was prepared by diluting the complex extract of Osuyu-broccoli in purified water, adding it to a liposome base, and stirring at room temperature.

[0035] In this way, the complex extract liposome containing the Osuyu-broccoli complex extract according to the embodiment of the present invention was manufactured by highly concentrating the Osuyu-broccoli complex extract into liposomes, and it was confirmed that the concentration was not toxic, and that β-catenin was expressed and inflammatory cytokines were expressed.

[0036] As a result, the complex extract liposome containing the Osuyu-broccoli complex extract according to the embodiment of the present invention can exhibit excellent anti-inflammatory efficacy in hair follicle cells and thus exhibit hair loss improvement efficacy by producing the complex extract liposome by highly concentrating the Osuyu-broccoli complex extract into liposomes.

[0037]

[0038] This will be described in more detail through a method for producing a complex extract liposome containing a complex extract of sesame oil and broccoli according to an embodiment of the present invention.

[0039] Figure 1 is a process flow diagram showing a method for manufacturing a complex extract liposome containing a complex extract of sesame oil and broccoli according to an embodiment of the present invention.

[0040] As shown in FIG. 1, the method for preparing a complex extract liposome containing a complex extract of osuyu-broccoli according to an embodiment of the present invention includes a step of preparing an osuyu extract and a broccoli extract (S110), a step of forming an osuyu-broccoli complex extract (S120), a step of diluting and stirring (S130), and a step of forming an osuyu-broccoli complex extract liposome (S140).

[0041]

[0042] Preparation of osuyu extract and broccoli extract

[0043] In the step of preparing the oilseed oil extract and broccoli extract (S110), the oilseed oil extract and the broccoli extract are prepared.

[0044] Here, Evodiae Fructus is the dried ripe fruit of the Rutaceae family. In oriental medicine, the fruit is called Evodiae Fructus and has stomachic and analgesic properties. In addition, Evodiae Fructus contains effective ingredients such as evodiamine, which has anti-cancer properties, and dehydroevodiamine, evodiamine, and rutaecarpine, which have anti-inflammatory properties.

[0045] Broccoli also contains glucoraphanin and sulforaphane as its main components. Broccoli has also been reported to exhibit antibacterial activity against Helicobacter pylori, the cause of gastric ulcers, and to reduce the risk of cancer. Broccoli also contains twice as much vitamin C as lemons, and is rich in beta-carotene, vitamin B1, vitamin B2, vitamin E, potassium, and calcium.

[0046] In this step, the extract of the oily water may include a process of washing the oily water and then drying it with hot air, a process of grinding the dried oily water to obtain oily water powder, a process of adding the oily water powder to a solvent and performing a first extraction to form an oily water extract, a process of filtering the first extracted oily water extract, and a process of performing a second extraction using a rotary concentrate to form an oily water extract.

[0047] In addition, in this step, the broccoli extract may include a process of washing broccoli and then drying it with hot air, a process of crushing the hot air-dried broccoli to obtain broccoli powder, a process of adding the broccoli powder to a solvent and performing a first extraction to form a broccoli extract, a process of filtering the first extracted broccoli extract, and a process of performing a second extraction using a rotary concentrate to form a broccoli extract.

[0048] Here, washing of the milk and broccoli can be done by using water washing, ultrasonic washing, vibration washing, etc., but is not limited thereto.

[0049] In addition, it is desirable to perform hot air drying at 30 to 50℃ for 12 to 60 hours.

[0050] If the hot air drying temperature is below 30℃ or the hot air drying time is less than 12 hours, there is a risk that the osuyu or broccoli may not be completely dried. If the hot air drying temperature exceeds 50℃ or the hot air drying time exceeds 60 hours, it may only increase the manufacturing cost and time without any further increase in effectiveness, so it is not economical.

[0051] In addition, the grinding may be performed using one or more methods selected from a jet mill, a ball mill, a roller mill, etc., but is not limited thereto.

[0052] It is preferable that the osuyu powder and broccoli powder each have an average diameter of 100 to 1,000 nm, and more preferably an average diameter of 300 to 500 nm. If the average diameter of the osuyu powder or broccoli powder is less than 100 nm, the process cost for grinding into fine particles may be excessively incurred, and thus it is not preferable. In addition, if the average diameter of the osuyu powder or broccoli powder exceeds 1,000 nm, it may aggregate when mixed with a solvent, and thus it is not preferable.

[0053] In addition, the solvent may be one selected from among water, methanol, ethanol, phenoxyethanol, propylene glycol, and dipropylene glycol, or a mixture of two or more thereof, and among these, it is more preferable to use ethanol.

[0054] It is desirable to carry out this primary extraction at room temperature of 1 to 40°C for 12 to 48 hours.

[0055] In addition, it is preferable to perform filtration using a 200 to 600 mesh filter for the sesame oil extract or broccoli extract, respectively.

[0056] In this way, through the first and second extractions, highly concentrated sesame oil extract and broccoli extract can be obtained, respectively.

[0057]

[0058] Formation of a complex extract of sesame oil and broccoli

[0059] In the step of forming a sesame oil-broccoli complex extract (S120), the sesame oil extract and the broccoli extract are mixed and stirred to form a sesame oil-broccoli complex extract.

[0060] In this step, it is preferable to mix the osuyu extract and the broccoli extract in a weight ratio of 3:1 to 1:3, and more preferably, it is preferable to mix them in a weight ratio of 2:1 to 1:2. In this way, by mixing the osuyu extract and the broccoli extract in a weight ratio of 1 to 1:3, the combined effects of osuyu and broccoli can be exhibited.

[0061]

[0062] Dilution and stirring

[0063] In the dilution and stirring step (S130), the oil-soluble fiber-broccoli complex extract is diluted in purified water to form a oil-soluble fiber-broccoli complex extract dilution solution.

[0064] Here, purified water is used to dilute the fermented soybean-broccoli complex extract, and may be water obtained through one or more distillations or purified through a filter.

[0065] It is preferable to use a diluted solution of the Osuyu-broccoli complex extract by diluting the Osuyu-broccoli complex extract with purified water in a weight ratio of 1:8 to 1:12.

[0066]

[0067] Liposome formation of Osuyu-broccoli complex extract

[0068] In the step of forming a liposome of the oil-broccoli complex extract (S140), a liposome base is mixed with the oil-broccoli complex extract dilution and stirred to form an oil-broccoli complex extract liposome.

[0069] In this step, it is preferable to mix the liposome base in an amount of 50 to 150 parts by weight per 100 parts by weight of the diluted oil-broccoli complex extract. If the liposome base is mixed in an amount of less than 50 parts by weight per 100 parts by weight of the diluted oil-broccoli complex extract, it may be difficult to control the oil-broccoli complex extract to be stably included inside the spheres, and if it is mixed in an amount exceeding 150 parts by weight, the content of the oil-broccoli complex extract may relatively decrease, which may prevent it from being highly concentrated, and thus this is not preferable.

[0070] In this way, in the present invention, by strictly controlling the mixing ratio of the liposome base and the diluted solution of the Osuyu-broccoli complex extract, the Osuyu-broccoli complex extract is highly concentrated into liposomes to produce a complex extract liposome, thereby exhibiting excellent anti-inflammatory effects in hair follicle cells and thus exhibiting hair loss improvement effects.

[0071] Here, the liposome base comprises 70 to 90 wt% of a moisturizer, 10 to 30 wt% of an emulsifier, and 0.1 to 0.5 wt% of an oil.

[0072] Humectants provide moisturizing benefits. These humectants include at least one selected from the following: butylene glycol, polyethylene glycol, propylene glycol, dipropylene glycol, 1,3-butylene glycol, glycerin, diglycerin, propanediol, 1,2-hexanediol, and sodium hyaluronate. Among these, propanediol is more preferred as a moisturizer.

[0073] If the amount of such moisturizer is less than 70% by weight, the amount may be insufficient to provide adequate moisturizing effects. Conversely, if the amount exceeds 90% by weight, the viscosity may increase, resulting in stickiness, which is not desirable.

[0074] In addition, emulsifiers play a role in maintaining high usability and emulsion stability.

[0075] These emulsifiers include Glyceryl Stearate, Ceteareth-20, Ceteareth-12, Cetearyl Alcohol, Cetyl Palmitate, Glyceryl Stearate, PEG-100 Stearate, Polyhydroxystearic Acid, Lecithin, Ethylhexyl Palmitate, Isopropyl Myristate, Isostearic Acid, Polyglyceryl-3 Polyricinoleate, Lauryl PEG-9 Polydimethylsiloxyethyldimethicone. It may include at least one selected from polydimethylsiloxyethyl dimethicone and PEG-10 dimethicone. Among these, it is more preferable to use lecithin as an emulsifier.

[0076] Oil dissolves oil-based components and forms a strong interfacial film to improve spreadability, flowability, and absorbency. Any oil commonly used in the relevant technical field can be used without restriction. For example, various oils such as hydrocarbon oil, ester oil, silicone oil, and animal or vegetable oil can be used, and various other oils can also be applied. For example, the oil may be at least one selected from CEH (Cetyl 2-ethyl hexanoate), MCT (Medium Chain Triglyceride), BASF (Cetiol Ultimate), SF0015Z, SF9902E, jojoba oil, sunflower oil, Arlacel-165, Lecimul-LCR, stearic acid, and Erocol-6850. Among these, sunflower oil is preferably used as the oil.

[0077] If the amount of oil added is less than 0.1 wt%, it is difficult to properly exhibit the effects of improving spreadability, flowability, and absorbency, and if it exceeds 0.5 wt%, there is a concern that the content of other effective ingredients may be relatively low, so it is not desirable.

[0078] In addition, in the step of forming liposomes of the Osuyu-broccoli complex extract (S140), it is preferable to perform stirring at a room temperature of 1 to 40°C and a speed of 500 to 1,500 rpm for 1 to 3 hours.

[0079] If the stirring speed is less than 500 rpm or the stirring time is less than 1 hour, there is a risk that the liposome base may not be uniformly mixed in the diluted extract of the sesame oil-broccoli complex. If the stirring speed exceeds 1,500 rpm or the stirring time exceeds 3 hours, it may only increase the manufacturing cost without any further effect, making it uneconomical.

[0080] In addition, it is more preferable to perform ultrasonic treatment at the same time with a frequency of 10 to 25 KHz and an output voltage of 3 to 10 W during stirring, in order to further improve the dispersibility between the diluted extract of the Osuyu-broccoli complex and the liposome base.

[0081] When applying such ultrasound, if the frequency is less than 10 KHz or the output voltage is less than 3 W, the cavitation effect due to the application of ultrasound is minimal, making it difficult to properly exhibit the effect of improving dispersibility. In addition, if the frequency of the ultrasound exceeds 25 KHz or the output voltage exceeds 10 W, there is a risk that the liposome base may be damaged due to excessive application of ultrasound, which is not desirable.

[0082]

[0083] According to the above, the method for preparing a complex extract liposome containing a complex extract of Osuyu-broccoli according to an embodiment of the present invention was prepared by diluting the complex extract of Osuyu-broccoli in purified water, adding it to a liposome base, and stirring at room temperature.

[0084] In this way, the method for manufacturing a complex extract liposome containing a complex extract of Osuyu-broccoli according to an embodiment of the present invention was performed by highly concentrating the complex extract of Osuyu-broccoli into liposomes to manufacture a complex extract liposome, thereby confirming a concentration that does not show toxicity and confirming that β-catenin is expressed and that inflammatory cytokines are expressed.

[0085] As a result, the method for producing a complex extract liposome containing a complex extract of Osuyu-broccoli according to an embodiment of the present invention can exhibit excellent anti-inflammatory effects in hair follicle cells and thus exhibit hair loss improvement effects by producing a complex extract liposome by highly concentrating the complex extract of Osuyu-broccoli into liposomes.

[0086]

[0087] Example

[0088] Hereinafter, the structure and operation of the present invention will be described in more detail through preferred embodiments of the present invention. However, these are presented as preferred examples of the present invention and should not be construed as limiting the present invention in any way.

[0089] Anything not described here will be omitted as it is technically feasible for those skilled in this field to infer.

[0090]

[0091] 1. Sample preparation

[0092] Preparation of Osuyu extract

[0093] The waste milk was washed and the washed waste milk was dried with hot air at 40℃ for 48 hours.

[0094] Next, 100 g of dried sesame oil powder obtained by grinding hot-air-dried sesame oil was added to 1 L of 70% ethanol and primary extraction was performed at room temperature (20°C) for 24 hours to prepare an sesame oil extract.

[0095] Next, the prepared sesame oil extract was filtered through a 400-mesh polyethylene net and then extracted a second time using a rotary concentrator (N-1300, Sunil Aira Co., Ltd.) to prepare the sesame oil extract.

[0096]

[0097] Preparation of broccoli extract

[0098] Broccoli was washed and dried with hot air at 40°C for 48 hours.

[0099] Next, 100 g of broccoli powder obtained by crushing hot-air-dried broccoli was added to 1 L of 70% ethanol and subjected to a primary extraction at room temperature (20°C) for 24 hours to prepare a broccoli extract.

[0100] Next, the prepared broccoli extract was filtered through a 400-mesh polyethylene net and then subjected to secondary extraction using a rotary concentrator (N-1300, Sunil Aira Co., Ltd.) to prepare a broccoli extract.

[0101]

[0102] Preparation of liposomes containing complex extracts of sesame oil and broccoli extracts

[0103] The extract of Osuyu and the extract of Broccoli were mixed in a weight ratio of 1:1 and stirred to prepare a complex extract of Osuyu and Broccoli.

[0104] Next, the oil-soluble-broccoli complex extract was diluted with purified water at a weight ratio of 1:10 to prepare a diluted oil-soluble-broccoli complex extract solution, and then the oil-soluble-broccoli complex extract was mixed with a liposome base at a weight ratio of 4:6. At this time, the liposome base was used as a mixture of 79.8 wt% propanediol, 20 wt% lecithin, and 0.2 wt% sunflower oil.

[0105] Next, a mixture of the Osuyu-broccoli complex extract and liposome base was mixed and stirred at 1,000 rpm for 2 hours at room temperature (20°C) using a homodisper (Poonglim Tech Co., Ltd.) to prepare Osuyu-broccoli complex extract liposomes.

[0106]

[0107] 2. Confirmation of cytotoxicity and growth

[0108] Figure 2 is a graph showing the results of absorbance measurements at 490 nm for broccoli, oat milk, and broccoli + oat milk, and Figure 3 is a graph showing the results of absorbance measurements at 490 nm for liposomes and broccoli + oat milk liposomes.

[0109] <Experimental Method>

[0110] HDP cells (human dermal papilla cells) were seeded at 1.5 × 10 in a 96-well plate. 4 After dispensing cells / well, they were cultured for 24 hours.

[0111] Ethanol extracts were treated to cells at various concentrations (0, 5, 10, 50, 100, 300, 500, 1000 ㎍ / mL), and liposome extracts were also treated to cells at various concentrations (0, 1, 3, 5, 10, 30, 50, 100 ㎍ / mL) and cultured for 24 hours.

[0112] Cell toxicity and growth were confirmed using a proliferation kit.

[0113] After adding 20 μl of kit reagent to each well, the reaction was carried out in an incubator under the same conditions for 1 hour, and the absorbance was measured at 490 nm.

[0114]

[0115] As shown in Figures 2 and 3, the concentrations that did not show toxicity were confirmed to be 500 ㎍ / mL of broccoli (Broccoli E), 100 ㎍ / mL of oat milk (Oat milk E), 50 ㎍ / mL of broccoli + oat milk (Complex E), 100 ㎍ / mL of liposome (Liposome), and 100 ㎍ / mL of broccoli + oat milk liposome (Complex Liposome). ELISA and western blot experiments were performed at these concentrations in the future.

[0116]

[0117] 3. Confirmation of β-catenin

[0118] Figure 4 shows the results of confirming β-catenin by confirming the band using ImageQuant 800 for broccoli, osuyu, broccoli + osuyu, liposome, and broccoli + osuyu liposome.

[0119] <Experimental Method>

[0120] HDP cells (human dermal papilla cells) were seeded at 2 × 10 in a 6-well plate. 5 The cells were dispensed per well and cultured in a CO2 incubator at 37°C for 24 hours.

[0121] Each extract, 500 ㎍ / mL of broccoli, 100 ㎍ / mL of oat milk, 50 ㎍ / mL of broccoli + oat milk, 100 ㎍ / mL of liposome, and 100 ㎍ / mL of broccoli + oat milk liposome, was treated to HDP cells and cultured for 24 hours.

[0122] After culturing, the cells were washed with PBS, 1 mL of PBS was dispensed per well, the cells were harvested using a scraper, transferred to a 1.5 mL tube, centrifuged at 15,000 rpm for 10 minutes in a 4°C centrifuge, the supernatant was removed, 200 μl of RIPA buffer was added, mixed, and centrifuged again at 15,000 rpm for 10 minutes in a 4°C centrifuge. Only the supernatant was harvested and quantified.

[0123] After preparing an 8% gel (gel_) according to the composition, electrophoresis was performed at 50°C for 30 minutes and at 100°C for 1 hour 30 minutes, and then transferred to a PVDF membrane for 1 hour 30 minutes.

[0124] Blocking was performed with 5% BSA for 1 hour at room temperature, and β-catenin primary antibody was reacted at a 1:1000 dilution for 1 hour at room temperature.

[0125] After washing with TBST and reacting with secondary antibody for 1 hour, the bands were confirmed with ImageQuant 800.

[0126]

[0127] As shown in Fig. 4, as a result of the experiment, it was confirmed that the expression of β-catenin increased by each extract: broccoli 500 ㎍ / mL (BrE), oil 100 ㎍ / mL (OE), broccoli + oil 50 ㎍ / mL (BoE), liposome 100 ㎍ / mL (L), and broccoli + oil liposome 100 ㎍ / mL (BoL).

[0128]

[0129] 4. Check the JAK / STAT path

[0130] Figure 5 shows the results of confirming the JAK / STAT pathway by confirming the bands using ImageQuant 800 for broccoli, osuyu, broccoli + osuyu, liposome, and broccoli + osuyu liposome.

[0131] <Experimental Method>

[0132] HDP cells were seeded at 2 × 10 in a 6-well plate. 5 The cells were dispensed per well and cultured in a CO2 incubator at 37°C for 24 hours.

[0133] Each extract, 500 ㎍ / mL of broccoli, 100 ㎍ / mL of oat milk, 50 ㎍ / mL of broccoli + oat milk, 100 ㎍ / mL of liposome, and 100 ㎍ / mL of broccoli + oat milk liposome, was treated to HDP cells and cultured for 24 hours.

[0134] After 24 hours, LPS was treated at 1 μg / mL for 1 hour.

[0135] After the reaction, the cells were washed with PBS, 1 mL of PBS was dispensed per well, the cells were harvested with a scraper, transferred to a 1.5 mL tube, centrifuged at 15,000 rpm for 10 minutes in a 4°C centrifuge, the supernatant was removed, 200 μl of RIPA buffer was added, mixed, and centrifuged again at 15,000 rpm for 10 minutes in a 4°C centrifuge. Only the supernatant was harvested and quantified.

[0136] After preparing an 8% gel according to the composition, electrophoresis was performed at 50°C for 30 minutes and at 100°C for 1 hour and 30 minutes, and then transferred to a PVDF membrane for 1 hour and 30 minutes.

[0137] Blocking was performed with 5% BSA for 1 hour at room temperature, and JAK1, p-JAK1, STAT3, and p-STAT3 primary antibodies were diluted 1:1000 and reacted for 1 hour at room temperature.

[0138] After washing with TBST and reacting with secondary antibody for 1 hour, the bands were confirmed with ImageQuant 800.

[0139]

[0140] As shown in Fig. 5, phosphorylation of JAK1 and STAT3 was increased by LPS, and phosphorylation was decreased by each extract: broccoli 500 ㎍ / mL (BrE), oil 100 ㎍ / mL (OE), broccoli + oil 50 ㎍ / mL (BoE), liposome 100 ㎍ / mL (L), and broccoli + oil liposome 100 ㎍ / mL (BoL).

[0141]

[0142] 5. Confirm the MAPK pathway

[0143] Figure 6 shows the results of confirming the MAPK pathway by confirming the bands using ImageQuant 800 for broccoli, osuyu, broccoli + osuyu, liposome, and broccoli + osuyu liposome.

[0144] <Experimental Method>

[0145] HDP cells were seeded at 2 × 10 in a 6-well plate. 5 The cells were dispensed per well and cultured in a CO2 incubator at 37°C for 24 hours.

[0146] Each extract, 500 ㎍ / mL of broccoli, 100 ㎍ / mL of oat milk, 50 ㎍ / mL of broccoli + oat milk, 100 ㎍ / mL of liposome, and 100 ㎍ / mL of broccoli + oat milk liposome, was treated to HDP cells and cultured for 24 hours.

[0147] After 24 hours, LPS was treated at 1 μg / mL for 1 hour.

[0148] After the reaction, the cells were washed with PBS, 1 mL of PBS was dispensed per well, the cells were harvested with a scraper, transferred to a 1.5 mL tube, centrifuged at 15,000 rpm for 10 minutes in a 4°C centrifuge, the supernatant was removed, 200 μl of RIPA buffer was added, mixed, and centrifuged again at 15,000 rpm for 10 minutes in a 4°C centrifuge. Only the supernatant was harvested and quantified.

[0149] After preparing an 8% gel according to the composition, electrophoresis was performed at 50°C for 30 minutes and at 100°C for 1 hour and 30 minutes, and then transferred to a PVDF membrane for 1 hour and 30 minutes.

[0150] Blocking was performed with 5% BSA for 1 hour at room temperature, and then ERK, p-ERK, JNK, and p-JNK primary antibodies were reacted at a 1:1000 dilution for 1 hour at room temperature.

[0151] After washing with TBST and reacting with the secondary antibody for 1 hour, the bands were confirmed using ImageQuant 800.

[0152]

[0153] As shown in Fig. 6, phosphorylation of ERK and JNK increased by LPS, and phosphorylation decreased by each extract: broccoli 500 ㎍ / mL (BrE), oil 100 ㎍ / mL (OE), broccoli + oil 50 ㎍ / mL (BokE), liposome 100 ㎍ / mL (L), and broccoli + oil liposome 100 ㎍ / mL (BokL).

[0154]

[0155] 6. Confirmation of inflammatory cytokines

[0156] Figure 7 is a graph showing the results of measuring cytokine levels using an ELISA kit for broccoli, oat milk, broccoli + oat milk, liposomes, and broccoli + oat milk liposomes. In this case, Figure 7 (a) shows the results of measuring the level of TNF-α, and Figure 7 (b) shows the results of measuring the level of IL-1β.

[0157] <Experimental Method>

[0158] HDP cells were seeded at 1.5 × 10 in a 96-well plate. 4 Cells were dispensed per well and cultured for 24 hours.

[0159] After pretreatment for 24 hours with the concentrations of each extract (broccoli 500 μg / mL, oat milk 100 μg / mL, broccoli + oat milk 50 μg / mL, control liposome 100 μg / mL, broccoli + oat milk liposome 100 μg / mL), LPS (1 μg / mL) was treated for 1 hour.

[0160] After the reaction was completed, only the culture medium was collected, centrifuged, and only the supernatant was collected. Then, the cytokine levels were evaluated using the TNF-α and IL-1β ELISA kits.

[0161]

[0162] As shown in (a) and (b) of Figure 7, the levels of cTNF-α and IL-1β increased by LPS were reduced by each extract: broccoli 500 ㎍ / mL (BrE), osuyu 100 ㎍ / mL (OE), broccoli + osuyu 50 ㎍ / mL (BokE), liposome 100 ㎍ / mL (L), and broccoli + osuyu liposome 100 ㎍ / mL (BokL).

[0163]

[0164] 7. Quality Test Evaluation

[0165] Figure 8 shows the results of a quality test on the total aerobic bacterial count for a scalp care hair shampoo composition containing a liposome containing a complex extract of sesame oil and broccoli.

[0166] As shown in Fig. 8, the results of the total aerobic bacterial count quality test on the scalp care hair shampoo composition containing the liposome of the Osuyu-broccoli complex extract commissioned to the Korea Testing & Research Institute for Chemical Industry showed that there were no unusual issues.

[0167]

[0168] Figure 9 shows the results of a quality test for non-detection of lead / arsenic heavy metals for a scalp care hair shampoo composition containing a liposome of a complex extract of Osuyu and broccoli.

[0169] As shown in Fig. 9, the results of a quality test for non-detection of heavy metals such as lead / arsenic on a scalp care hair shampoo composition containing a liposome of a complex extract of oosuyu and broccoli commissioned to the Korea Testing & Research Institute for Chemical Industry confirmed that no heavy metals such as lead / arsenic were detected.

[0170]

[0171] Figure 10 shows the results of a pH quality test for a scalp care hair shampoo composition containing a liposome of a complex extract of sesame oil and broccoli.

[0172] As shown in Fig. 10, the results of the PH quality test on the scalp care hair shampoo composition containing the liposome of the Osuyu-broccoli complex extract commissioned to the Korea Testing & Research Institute for Chemical Industry showed that there were no unusual issues.

[0173]

[0174] Figure 11 shows the results of a quality test for the absence of E. coli for a scalp care hair shampoo composition containing a liposome of a complex extract of Osuyu and broccoli.

[0175] As shown in Fig. 11, it was confirmed that no E. coli was detected in the quality test results for the scalp care hair shampoo composition containing the liposome of the Osuyu-broccoli complex extract commissioned to the Korea Testing & Research Institute for Chemical Industry.

[0176]

[0177] Figure 12 shows the results of a quality test for the non-detection of Staphylococcus aureus for a scalp care hair shampoo composition containing a liposome of a complex extract of Osuyu and broccoli.

[0178] As shown in Fig. 12, it was confirmed that Staphylococcus aureus was not detected in the quality test results for a scalp care hair shampoo composition containing a liposome of a complex extract of sesame oil and broccoli commissioned to the Korea Testing & Research Institute for Chemical Industry.

[0179]

[0180] Figure 13 shows the results of a quality test for non-detection of formaldehyde for a scalp care hair shampoo composition containing a liposome of a complex extract of sesame oil and broccoli.

[0181] As shown in Fig. 13, it was confirmed that no formaldehyde was detected in the formaldehyde-free quality test results for a scalp care hair shampoo composition containing a liposome of a complex extract of oosuyu and broccoli commissioned to the Korea Testing & Research Institute for Chemical Industry.

[0182]

[0183] Although the embodiments and comparative examples of the present invention have been described in detail as above, the scope of the present invention is not limited thereto, and the scope of the present invention extends to a range substantially equivalent to the embodiments of the present invention.

Claims

1. (a) Step of preparing a milk extract and a broccoli extract; (b) a step of mixing and stirring the above-mentioned oil extract and broccoli extract to form an oil-broccoli complex extract; (c) a step of diluting the above-mentioned Osuyu-broccoli complex extract in purified water to form a Osuyu-broccoli complex extract dilution solution; and (d) a step of mixing a liposome base into the diluted solution of the above-mentioned oil-broccoli complex extract and stirring to form an oil-broccoli complex extract liposome; A method for producing a complex extract liposome containing a complex extract of sesame oil and broccoli, characterized in that it comprises:

2. In paragraph 1, In step (a) above, The above oil extract Step of washing the sewage and then drying it with hot air; A step of grinding the above-mentioned hot-air-dried waste milk to obtain waste milk powder, and then putting the waste milk powder into a solvent and performing a first extraction to form a waste milk extract; and A step of filtering the first extracted sesame oil extract and then performing a second extraction using a rotary concentrate to form an sesame oil extract; A method for producing a complex extract liposome containing a complex extract of sesame oil and broccoli, characterized in that it comprises:

3. In paragraph 1, In step (a) above, The above broccoli extract Step 1: Wash and dry broccoli with hot air; A step of crushing the hot-air-dried broccoli to obtain broccoli powder, then adding the broccoli powder to a solvent and performing a first extraction to form a broccoli extract; and A step of filtering the first extracted broccoli extract and then performing a second extraction using rotary concentration to form a broccoli extract; A method for producing a complex extract liposome containing a complex extract of sesame oil and broccoli, characterized in that it comprises:

4. In paragraph 2 or 3, The above solvent is A method for producing a complex extract liposome containing a complex extract of osuyu-broccoli, characterized in that it uses one selected from water, methanol, ethanol, phenoxyethanol, propylene glycol and dipropylene glycol alone or a mixture of two or more thereof.

5. In paragraph 1, In step (b) above, The above oilseed oil extract and broccoli extract A method for producing a complex extract liposome containing a complex extract of osuyu and broccoli, characterized in that the complex extract is mixed in a weight ratio of 3:1 to 1:

3.

6. In paragraph 1, In step (c) above, The above diluted extract of Osuyu-broccoli complex is A method for producing a complex extract liposome containing a complex extract of osuyu-broccoli, characterized in that the complex extract of osuyu-broccoli is diluted in purified water in a weight ratio of 1:8 to 1:

12.

7. In paragraph 1, In step (d) above, The above liposome base is A method for producing a complex extract liposome containing a complex extract of osuyu-broccoli, characterized in that 50 to 150 parts by weight are mixed with respect to 100 parts by weight of the diluted solution of the complex extract of osuyu-broccoli.

8. In paragraph 1, In step (d) above, The above liposome base is A method for producing a complex extract liposome containing a complex extract of osuyu-broccoli, characterized in that it comprises 70 to 90 wt% of a moisturizer, 10 to 30 wt% of an emulsifier, and 0.1 to 0.5 wt% of an oil.

9. A complex extract liposome containing a complex extract of sesame oil and broccoli, manufactured by a method according to any one of claims 1 to 8, The above complex extract liposome is manufactured by concentrating the complex extract of Osuyu-broccoli into liposomes, thereby exhibiting anti-inflammatory effects in hair follicle cells and thus having a hair loss improvement effect. A complex extract liposome containing the complex extract of Osuyu-broccoli.

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