KR102596741B1 - Hybrid liposome conjugated extracellular vesicles derived callus of plant and coenzyme q10 liposome and the use thereof - Google Patents

Abstract

The present invention relates to a hybrid liposome in which plant callus-derived extracellular endoplasmic reticulum and Coenzyme Q10 liposome are fused and its use. Specifically, the hybrid liposome according to the present invention is a fusion of plant callus-derived extracellular vesicles and Coenzyme Q10 liposomes, and exhibits significantly increased antioxidant activity compared to the case of treatment with Centella asiatica callus-derived extracellular vesicles and Coenzyme Q10 liposomes, respectively. It can be usefully used as a material for various functional cosmetics based on its antioxidant activity.

Description

Hybrid liposomes fused with plant callus-derived extracellular vesicles and Coenzyme Q10 liposomes and their uses {HYBRID LIPOSOME CONJUGATED EXTRACELLULAR VESICLES DERIVED CALLUS OF PLANT AND COENZYME Q10 LIPOSOME AND THE USE THEREOF}

The present invention relates to a hybrid liposome in which plant callus-derived extracellular endoplasmic reticulum and Coenzyme Q10 liposome are fused and its use.

The skin is largely composed of three layers: the epidermis, dermis, and subcutaneous fat tissue, and serves to protect the human body from the external environment. Among them, the epidermis is divided into the stratum corneum, stratum granulosum, stratum spinosum, and stratum basale. The stratum corneum, the outermost layer, has a brick and mortar structure. Keratinocytes act as bricks, and intercellular lipids form a skin barrier by acting as a mortar that holds the brick-like keratinocytes firmly, suppressing moisture evaporation to the outside of the body and external physical or chemical stimulation. protects the human body from

In addition, the stratum corneum contains a hydrophilic natural moisturizing factor with the ability to retain water, contributing to skin moisturization. Appropriate oil formed from sebum from sebaceous glands and sweat secreted from sweat glands minimizes moisture loss from the skin and moisturizes the skin. maintain

However, even if the skin is well-moisturized and healthy, the skin can dry out due to various factors such as stress caused by environmental pollution in modern society, external environment such as wind, extreme cold, and ultraviolet rays, natural aging, and skin stress caused by makeup. Phenomenon such as becoming dull, dull, and rough appear. Accordingly, research is being actively conducted to realize healthy skin by improving skin tightness, increasing moisturizing power, and increasing dermal density of the skin.

In this regard, Republic of Korea Patent Publication No. 10-2020-0138142 shows that daffodil, lily, and lily of the valley bulb extracts improve the skin's moisture retention ability, improve keratin, reduce skin irritation, increase skin elasticity, and improve wrinkles. , discloses that it can be used as a composition for improving skin.

Meanwhile, Centella asiatica is a perennial plant belonging to the dicotyledonous Apiaceae family and grows naturally in temperate and tropical wetlands in many regions of the world. Centella asiatica grows sideways, roots grow from the nodes, and has palm-shaped leaves. The top leaf emerges from the scale-like axil, has a long stalk, is 2 to 5 cm in diameter, has an oval shape, and has blunt sawtooth edges.

Republic of Korea Patent Publication No. 10-2020-0138142

The purpose of the present invention is to provide a hybrid liposome in which plant callus-derived extracellular endoplasmic reticulum and Coenzyme Q10 liposome are fused together, and a use thereof.

In order to achieve the above object, the present invention provides a hybrid liposome in which plant callus-derived extracellular vesicles and Coenzyme Q10 liposomes are fused.

Additionally, the present invention provides a cosmetic composition containing the hybrid liposome.

Additionally, the present invention provides an external skin preparation containing the hybrid liposome.

Furthermore, the present invention provides a functional composition containing hybrid liposomes.

The hybrid liposome according to the present invention is a fusion of plant callus-derived extracellular vesicles and Coenzyme Q10 liposomes, and exhibits significantly increased antioxidant activity compared to the case of treating Centella asiatica callus-derived extracellular vesicles and Coenzyme Q10 liposomes separately, thereby increasing the antioxidant activity. It can be usefully used as a material for various functional cosmetics.

Figure 1 is a photograph showing the results of photographing a Centella asiatica leaf (A) before inducing callus and a Centella asiatica leaf with callus induced (B) in one embodiment of the present invention.
Figure 2 is a graph showing the results of nanoparticle tracking analysis using Centella callus-derived extracellular vesicles (A) and hybrid liposomes of Centella callus-derived extracellular vesicles and Coenzyme Q10 liposomes (B) prepared in an example of the present invention. am.
Figure 3 is a graph showing the results of confirming the antioxidant activity of hybrid liposomes of extracellular vesicles derived from Centella asiatica callus in vitro Coenzyme Q10 liposomes in one embodiment of the present invention.

Hereinafter, the present invention will be described in detail.

The present invention provides a hybrid liposome in which plant callus-derived extracellular vesicles and Coenzyme Q10 liposomes are fused.

As used herein, the term “extracellular vesicles (EV)” refers to substances secreted from cells and refers to vesicles with a size of 30 to 1,000 nm surrounded by a phospholipid bilayer. Typically, the extracellular vesicles include apoptotic bodies, microvesicles, and exosomes. The extracellular vesicles are secreted by most eukaryotic cells and contain various substances such as RNA, DNA fragments, and proteins, and are known to serve as a delivery vehicle to regulate the microenvironment.

The plants may include all types of plants known in the art. In one embodiment of the present invention, the plant may be Centella asiatica. That is, the plant-derived extracellular vesicles according to the present invention may be centella asiatica-derived extracellular vesicles. In one embodiment of the present invention, the extracellular vesicles may be derived from centella asiatica leaves.

The callus can be cultured by methods known in the art, and the culture conditions can be appropriately modified by those skilled in the art. For example, the callus may be cultured in one or more medium selected from the group consisting of MS medium and B5 medium.

Meanwhile, the term “coenzyme Q10” used in this specification is a compound represented by the following formula (1) and refers to one of various coenzymes present in the human body. The Coenzyme Q10 is also referred to as GoQ10 or Ubiquinone, and can be synthesized in the body. The coenzyme Q10 has a similar effect to vitamin E and is known to have fatigue recovery and antioxidant activity.

[Formula 1]

In addition, the term "liposome" used herein refers to a small sac-shaped material formed of a phospholipid bilayer, which exhibits amphiphilic properties through an appropriate combination of hydrophilic and hydrophobic substances, and is mainly used as a carrier for delivering active substances in the body. It is used. The liposome may be a conventional spherical particle having a phospholipid bilayer and an empty space inside, and there are no particular restrictions on the type or manufacturing method of the phospholipid forming the phospholipid bilayer.

For example, the phospholipid may include one or more selected from the group consisting of soy lecithin, egg yolk lecithin, hydrogenated lecithin, hydrogenated lysolecithin, and hydrogenated lecithin. Additionally, the phospholipid may further include pegylated phospholipid or cholesterol, if necessary. In one embodiment of the present invention, the phospholipid may be hydrogenated lecithin.

The Coenzyme Q10 liposome can be prepared according to methods well known in the art. At this time, Coenzyme Q10 can be collected after manufacturing liposomes, or can be manufactured by adding it during the process of manufacturing liposomes. In one embodiment of the present invention, the liposome may be a liposome encapsulating Coenzyme Q10 therein. In addition, the Coenzyme Q10 liposome may further include components having the same or similar activity in addition to Coenzyme Q10. At this time, the additional ingredient may include hydroxydecylubiquinone.

The hybrid liposome according to the present invention may be prepared by fusing the plant-derived extracellular vesicle and Coenzyme Q10 liposome. At this time, the fusion can be performed under normal conditions and can be appropriately adjusted by a person skilled in the art as needed. The hybrid liposome may be a fusion of a phospholipid bilayer constituting a plant-derived extracellular vesicle and a Coenzyme Q10 liposome.

Additionally, the hybrid liposome can be prepared by mixing plant-derived extracellular vesicles and Coenzyme Q10 liposomes in an appropriate ratio. For example, the hybrid liposome contains 1 to 30% (v/v), 1 to 25% (v/v), 1 to 20% (v/v), 1 to 20% (v/v) of plant-derived extracellular vesicles based on the total of the hybrid liposome. 15% (v/v), 1 to 12% (v/v), 5 to 25% (v/v), 5 to 20% (v/v), 5 to 15% (v/v), 5 to 12% (v/v), 8 to 25% (v/v), 8 to 20% (v/v), 8 to 15% (v/v) or 8 to 12% (v/v). You can.

The hybrid liposome according to the present invention may further include polyol. The polyol may include all types of polyols known in the art, and may be included in an appropriate amount by those skilled in the art. For example, the polyol is butylene glycol, glycerin, glycerol, propanol, 1,2-hexanediol, 1,3-butylene glycol, propylene glycol, dipropylene glycol, isoprene glycol, sorbitol, xylitol. , erythol, fructose, lactose, mannitol, glucose, sugar, maltose, and malditol. In one embodiment of the present invention, the polyol may be butylene glycol and glycerin.

The hybrid liposome can exhibit antioxidant activity.

Additionally, the present invention provides a cosmetic composition containing the hybrid liposome.

Hybrid liposomes included in the cosmetic composition according to the present invention may have the characteristics described above.

Additionally, the cosmetic composition may exhibit antioxidant activity. The term "antioxidant activity" refers to the action of suppressing oxidation, removing reactive oxygen species, which are by-products produced through oxygen activity and reaction processes, and removing damaged DNA, proteins, fats, etc. from oxidation reactions. It means protective activity. For example, the cosmetic composition may exhibit one or more activities selected from the group consisting of whitening, wrinkle improvement, anti-aging, and elasticity promotion.

The cosmetic composition of the present invention may contain 0.1 to 50% by weight of hybrid liposomes, specifically 1 to 10% by weight. The cosmetic composition can be applied directly to the skin for the purpose of antioxidant activity.

The cosmetic composition may be formulated into a commonly manufactured cosmetic formulation. The cosmetic composition may be formulated as a solution, suspension, emulsion, paste, gel, lotion, cream, powder, soap, surfactant-containing cleansing, oil, powder foundation, emulsion foundation, wax foundation and spray. Specifically, it may be softening lotion, nourishing lotion, nourishing cream, massage cream, essence, eye cream, cleansing cream, cleansing foam, cleansing water, pack, spray, or powder.

When the formulation of the cosmetic composition of the present invention is a paste, cream or gel, animal oil, vegetable oil, wax, paraffin, starch, tragacanth, cellulose derivative, polyethylene glycol, silicone, bentonite, silica, talc, zinc oxide or It may include mixtures thereof. Additionally, when the formulation of the cosmetic composition is powder or spray, it may contain lactose, talc, silica, aluminum hydroxide, calcium silicate, polyamide powder, or mixtures thereof. In particular, in the case of a spray, it may further include chlorofluorohydrocarbon, propane/butane, or dimethyl ether.

When the formulation of the cosmetic composition of the present invention is a solution or emulsion, it may include a solvent, solvating agent, emulsifying agent, or a mixture thereof as a carrier. Examples thereof include water, ethanol, isopropanol, ethyl carbonate, ethyl acetate, benzyl alcohol, benzyl benzoate, propylene glycol, 1,3-butyl glycol oil, glycerol aliphatic ester, polyethylene glycol, sorbitan fatty acid ester, etc.

When the formulation of the cosmetic composition of the present invention is a suspension, the carrier may include water, a liquid diluent such as ethanol or propylene glycol, a suspending agent such as ethoxylated isostearyl alcohol, polyoxyethylene sorbitol ester, and polyoxyethylene sorbitan ester, It may include microcrystalline cellulose, aluminum metahydroxide, bentonite, agar, tragacanth, or mixtures thereof. In addition, when the formulation of the cosmetic composition is a cleansing surfactant, the carrier may include aliphatic alcohol sulfate, aliphatic alcohol ether sulfate, sulfosuccinic acid monoester, isethionate, imidazolinium derivative, methyl taurate, sarcosinate, fatty acid. It may include amide ethersulfate, alkalamidobetaine, fatty alcohol, fatty acid glyceride, fatty acid diethanolamide, vegetable oil, lanolin derivative, ethoxylated glycerol fatty acid ester, or mixtures thereof.

In addition to the carrier component, the cosmetic composition of the present invention may include antioxidants, stabilizers, solubilizers, moisturizers, pigments, fragrances, sunscreens, coloring agents, surfactants, or mixtures thereof as auxiliaries. The auxiliary agent can be any material commonly used in the production of cosmetic compositions.

Additionally, the present invention provides an external skin preparation containing the hybrid liposome.

Hybrid liposomes included in the external skin preparation according to the present invention may have the characteristics described above.

The external skin preparation of the present invention may contain a pharmaceutically acceptable carrier and excipient. The carrier and excipients may include preservatives, stabilizers, wetting agents, emulsification accelerators, and buffering agents. Specifically, the excipient may be lactose, dextrin, starch, mannitol, sorbitol, glucose, saccharose, microcrystalline cellulose, gelatin, polyvinylpyrrolidone, or mixtures thereof. The skin external preparation can be appropriately prepared according to methods well known in the art. The skin external preparations can be manufactured in the form of powder, gel, ointment, cream, liquid, and aerosol.

Furthermore, the present invention provides a functional composition comprising a hybrid liposome having the characteristics described above.

The functional composition may have antioxidant activity and can be used as various functional materials based on antioxidant activity.

Hereinafter, the present invention will be described in detail through the following examples. However, the following examples are only for illustrating the present invention and are not intended to limit the present invention thereto. Anything that has substantially the same structure as the technical idea described in the claims of the present invention and achieves the same operation and effect is included in the technical scope of the present invention.

Example 1. Preparation of callus derived from Centella asiatica

Callus was cultured from Centella asiatica leaves in the following manner.

First, centella asiatica leaves were cut to a certain size, mixed with the composition shown in Table 1 below, and stirred at 1,000 rpm for 20 minutes to prepare a medium mixture. The prepared medium mixture was sterilized under high pressure at 121°C for 15 minutes and cooled to prepare MS solid medium (pH 5.7-5.8). Centella asiatica leaves were added to the prepared MS medium and cultured at 28°C for 28 days. When callus was formed in the cultured Centella asiatica leaves, only the callus was isolated and subcultured.

ingredient content(%) Purified water 91.1624032 myo-inositol One thiamine hydrochloride One Pyridoxine hydrochloride One nicotinic acid One α-naphthaleneacetic acid 0.54 6-benzylaminopurine 0.22 sucrose 3 phyto agar 0.65 ammonium nitrate 0.165 boric acid 0.0062 Calcium chloride, anhydrous 0.03 cobalt chloride 6H _{2 O} 0.0000025 Cupric sulfate 5H _{2 O} 0.0000025 Ferric sodium EDTA 3H _{2 O} 0.004211 Magnesium sulfate, anhydrous 0.018 Manganese sulfate H _{2 O} 0.00169 Molybdic acid (sodium salt) 2H ₂ O (molybdic acid (sodium salt) 2H ₂ O) 0.000025 Potassium iodide 0.000083 potassium nitrate 0.19 Potassium phosphate (monobasic) 0.017 Zinc sulfate 7H _{2 O} 0.00086 glycine 0.0002

As a result, photos of the Centella asiatica leaves before callus induction and the callus induced from the Centella asiatica leaves are shown in Figure 1.

Example 2. Isolation of extracellular vesicles derived from Centella asiatica callus

Extracellular vesicles were isolated using the Centella asiatica callus obtained above as follows.

First, the callus induced in Example 1 was collected and pulverized using a homomixer. At this time, pulverization was performed for 30 minutes under conditions of 1,500 watt and 20 kHz, and the process of turning on the pulse for 5 seconds and stopping the pulse for 10 seconds was repeated. The pulverized material was centrifuged at 7,000 rpm for 30 minutes to recover the supernatant, and the recovered supernatant was centrifuged again at 12,000 rpm for 30 minutes. After centrifugation, only the supernatant was recovered and filtered through filter paper with a pore size of 0.2 ㎛ to finally obtain centella callus-derived extracellular vesicles.

Example 3. Preparation of hybrid liposomes of extracellular vesicles derived from Centella asiatica callus and Coenzyme Q10 liposomes

Hybrid liposomes were prepared by mixing the obtained extracellular vesicles derived from Centella asiatica callus with Coenzyme Q10 liposomes.

First, the ingredients as listed in Table 2 below were mixed and stirred at 700 rpm for 30 minutes, and the resulting mixture was homogenized with a homomixer under the conditions described above.

ingredient content(%) glycerin 40.75 butylene glycol 20 Purified water 18 Hydrogenated lecithin 9 Hydroxydecyl ubiquinone 2 Ubiquinone (coenzyme Q10) 0.25

Here, 10% of the extracellular vesicles obtained in Example 2 were added and homogenized by using a homomixer under the same conditions to prepare hybrid liposomes of Centella callus-derived extracellular vesicles and Coenzyme Q10 liposomes.

Example 4. Analysis of hybrid exosomes

The size, size distribution, and number of the hybrid liposomes prepared above were confirmed by performing nanoparticle tracking analysis (NTA) using a conventional method. At this time, extracellular vesicles derived from Centella asiatica callus were used as a control, and the resulting graph is shown in Figure 2.

As shown in Figure 2, the hybrid liposome according to the present invention showed a size of about 100 to 250 nm, and the particle size of the prepared hybrid liposome increased compared to the extracellular vesicles derived from Centella callus.

Experimental Example 1. Antioxidant activity of hybrid liposomes

The antioxidant activity of the hybrid liposome prepared above was confirmed by analyzing the DPPH radical scavenging ability. The experiment was performed in a conventional manner, and L-ascorbic acid was used as a positive control, and extracellular vesicles and Coenzyme Q10 liposomes derived from Centella asiatica callus were used as controls, respectively. As a result, the results calculated as relative values based on the DPPH radical scavenging ability of L-ascorbic acid are shown in Figure 3.

As shown in Figure 3, the hybrid liposome according to the present invention showed a significantly increased DPPH radical scavenging ability than when treated with Centella asiatica callus-derived extracellular vesicles and Coenzyme Q10, respectively.

Therefore, from the above results, the hybrid liposome according to the present invention can be usefully used in the production of functional cosmetics as a functional material based on antioxidant activity.

Claims (8)

It is a hybrid liposome that is a fusion of plant callus-derived extracellular vesicles and Coenzyme Q10 liposomes,
The extracellular vesicles contain 10% (v/v) based on the total hybrid liposome,
The size of the hybrid liposome is 100 to 250 nm,
The liposome is composed of hydrogenated lecithin,
A hybrid liposome in which hydroxydecylubiquinone is further entrapped inside the hybrid liposome.
The hybrid liposome according to claim 1, wherein the plant is Centella asiatica.
The hybrid liposome according to claim 1, wherein the Coenzyme Q10 is a compound represented by the following formula (1):
[Formula 1]
.
A cosmetic composition comprising the hybrid liposome of claim 1.
The cosmetic composition according to claim 4, wherein the cosmetic composition exhibits antioxidant activity.
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