KR102448514B1 - Composition for preventing, ameliorating or treating arteriosclerosis containing Cannabis sativa stem extract as effective component - Google Patents

Abstract

The present invention relates to a composition for preventing, improving or treating arteriosclerosis comprising a hemp stem extract as an active ingredient. It was confirmed that the expression was decreased and the expression of the cholesterol excretion gene was increased. Therefore, it is expected that the hemp stem extract, which is the active ingredient of the present invention, can be usefully used as a composition for preventing, improving or treating arteriosclerosis.

Description

Composition for preventing, ameliorating or treating arteriosclerosis containing Cannabis sativa stem extract as effective component

The present invention relates to a composition for preventing, improving or treating arteriosclerosis containing a hemp stem extract as an active ingredient.

Recently, the incidence of adult diseases is increasing due to westernized eating habits and lack of exercise in modern people, and among them, the number of arteriosclerosis is increasing year by year. Atherosclerosis is a very important disease to treat and manage because it causes a problem that greatly reduces the quality of life of patients along with various complications.

Arteriosclerosis (or atherosclerosis) may appear in the coronary arteries, aorta, and peripheral arteries, and there is a problem in that the blood vessels gradually narrow as they occur on the inner wall of the blood vessels. Causes include hyperlipidemia, high blood pressure, diabetes, or smoking, and periodic observation of the individual's condition is required. Hypertensive disease, heart disease, and cerebrovascular disease account for 30% of the total, and the problem of arteriosclerosis is gradually developing into a dangerous disease. It occurs wherever there are blood vessels, and especially myocardial infarction, stroke, or aortic dissection, etc., can cause sudden death, so be careful.

To keep blood vessels healthy, it is most important to control blood pressure and prevent the deposition of foreign substances in blood vessels. Blood pressure is maintained by organically regulating the contraction and relaxation of vascular endothelial cells and muscle cells along with the beating of the heart. If the blood vessels do not relax properly when pressure is applied to the blood vessels or the blood flow increases, blood pressure rises, which leads to hypertension, arrhythmia, heart attack, arteriosclerosis, and the like. Nitric oxide (NO) generated by endothelial nitric oxide synthase (eNOS) of vascular endothelial cells is well known as a representative vasodilator factor, and it has an antiatherogenic effect. It is also known to induce.

In the course of atherosclerosis, macrophages migrate into the blood vessel wall and lose control over oxidized-LDL and modified LDL, resulting in accumulation of lipid peroxide and continuous accumulation of cholesterol in the macrophage. As a result, cholesterol and cholesterol esters It can cause the formation of fatty streaks and fibrous plaques, which are thought to be early lesions of atherosclerosis, by forming foam cells in which lipids, the main component of which are the back, are deposited.

Although many drugs for treating arteriosclerosis have already been developed, existing drugs have side effects, and some drugs have serious side effects and their use has been limited. Therefore, studies are being conducted to treat or prevent arteriosclerosis using natural substances with relatively few side effects.

On the other hand, hemp is an annual herbaceous plant belonging to the Hemp family, and its scientific name is Cannabis sativa L.. Straight roots extend 30 to 40 cm underground, but side roots do not develop vigorously, so they are easily pulled out. The height is around 3m in temperate zones, but grows up to 6m in the tropics. The stem is longitudinally corrugated and the inner sheath fibers are formed in the parenchyma, which are bast fibers. The leaves are palm-like in shape and consist of 5-9 small leaves, opposite at the bottom of the stem and opposite at the top.

As a cannabis-related technology, a cannabis extract and a method of manufacturing and using the same are known in Korean Patent Application Laid-Open No. 2017-0080608, and a cannabidiol extract extracted from hemp and a colon cancer prevention or use method containing a trail in Korean Patent No. 2041875. Although a therapeutic pharmaceutical composition is known, a composition for preventing, improving or treating arteriosclerosis containing the hemp stem extract of the present invention as an active ingredient has not been disclosed.

The present invention has been derived by the above needs, and the present invention provides a composition for preventing, improving or treating arteriosclerosis containing a hemp stem extract as an active ingredient, and the hemp stem extract is lysophosphatidylcholine (LPC) In (16:0)-induced THP-1 macrophage-derived atherosclerotic cells, cytotoxicity was hardly observed, and the intracellular lipid content was reduced as well as the expression of genes related to the cholesterol biosynthesis pathway. and by confirming that it increases the expression of the cholesterol excreting gene, the present invention was completed.

In order to achieve the above object, the present invention provides a health functional food composition for preventing or improving arteriosclerosis containing hemp stem extract as an active ingredient.

In addition, the present invention provides a pharmaceutical composition for preventing or treating arteriosclerosis comprising a hemp stem extract as an active ingredient.

The present invention relates to a composition for preventing, improving or treating arteriosclerosis containing a hemp stem extract as an active ingredient, wherein the hemp stem extract does not contain CBD and THC-based compounds, and lysophosphatidylcholine (LPC) (16 :0) has little cytotoxicity in THP-1 macrophage-derived arteriosclerotic cells induced, and not only reduces intracellular lipid content, but also reduces cholesterol biosynthetic pathway-related gene expression, cholesterol It has the effect of increasing the expression of the excretion gene.

1 is a result confirming the cytotoxicity of hemp stem extract in arteriosclerosis model cells.
2 is an oil-red staining result confirming the change in lipid content in arteriosclerosis model cells treated with the cannabis stem extract (MIS) of the present invention for 48 hours by concentration, (A) control, (B) LPC 40 μM (C ) LPC 40 μM+oxiLDL 50 μg/ml, (D) LPC 40 μM+oxiLDL 50 μg/ml+MIS 100 μg/ml, (E) LPC 40 μM+oxiLDL 50 μg/ml+MIS 200 μg/ml, (F) LPC 40 μM+oxiLDL 50 μg/ml+MIS 400 μg/ml.
3 is a graph showing the results of oil-red O staining confirming the change in lipid content in arteriosclerosis model cells treated with the cannabis stem extract of the present invention for 48 hours at different concentrations. ** and *** indicate that the lipid content was increased in the arteriosclerosis-inducing group (LPC 40 μM, LPC 40 μM + oxiLDL 50 μg/ml) compared to the normal group, ** is p<0.01, *** is p <0.001, ##, ### indicate that the lipid content in the hemp extract treatment group of the present invention compared to the arteriosclerosis induction group was statistically significantly decreased, ## is p<0.01, ### is p< 0.001.
4 is a result confirming the change in the expression level of the cholesterol synthesis-related gene according to the treatment of the hemp stem extract (MIS) of the present invention. ** and *** indicate that the lipid content was increased in the arteriosclerosis induction group (LPC 40 μM) compared to the normal group, ** is p<0.01, *** is p <0.001, # and ## are The expression level of cholesterol synthesis-related genes in the hemp stem extract-treated group of the present invention compared to the arteriosclerosis-inducing group was statistically significantly decreased, with # being p<0.05 and ## being p<0.01.
5 is a result confirming the change in gene expression level of the SREBP family, a transcription factor of cholesterol synthesis-related genes, by the treatment of the cannabis stem extract of the present invention. * indicates that the expression of the SREBF1 gene was increased in the arteriosclerosis-inducing group compared to the normal group, p<0.05, and #, ## are the 100-400㎍/㎖ cannabis stem extract treatment group of the present invention compared to the arteriosclerosis-inducing group. The expression of the SREBF1 gene was decreased in
6 is a result confirming that the expression of the cholesterol excretion-related gene ABCA1 is increased by the treatment of 400 μg/ml cannabis stem extract of the present invention. *** indicates that the expression of the ABCA1 gene was increased in the arteriosclerosis-inducing group compared to the normal group, p<0.001, and ## indicates the ABCA1 gene expression in the hemp stem extract-treated group of the present invention compared to the arteriosclerosis-inducing group. increased, p<0.01.
7 is an HPLC result of a mixture (B) of a cannabis stem extract (A) of the present invention and a mixture (B) of CBD and THC-based compounds as standard materials.
8 is a GC-MS result for cannabidiol (CBDV) as a standard material and cannabis stem extract (A) of the present invention.

The present invention relates to a health functional food composition for preventing or improving arteriosclerosis containing a hemp stem extract as an active ingredient.

The cannabis stem extract may be prepared by a method comprising the following steps, but is not limited thereto:

(1) extracting by adding an extraction solvent to cannabis sativa stem;

(2) filtering the extract of step (1); and

(3) Concentrating the filtered extract of step (2) under reduced pressure and drying to prepare an extract.

In step (1), the extraction solvent is preferably selected from water, C ₁ to C ₄ lower alcohols or mixtures thereof, and more preferably water, but is not limited thereto.

Cannabis stem extraction in the present invention can use all conventional methods known in the art, such as filtration, hot water extraction, immersion extraction, reflux cooling extraction and ultrasonic extraction. The extraction solvent is preferably extracted by adding 1 to 20 times the weight of the dried hemp stem, and more preferably adding 5 to 15 times. The extraction temperature is preferably 4 ~ 50 ℃, but is not limited thereto. In addition, the extraction time is preferably 0.5 to 10 hours, more preferably 0.5 to 5 hours, but is not limited thereto. In the method, it is preferable to use a vacuum vacuum concentrator or a vacuum rotary evaporator for the concentration under reduced pressure in step (3), but is not limited thereto. In addition, drying under reduced pressure, vacuum drying, boiling drying, spray drying or freeze drying is preferable, but is not limited thereto.

The hemp stem extract is preferably prepared in any one formulation selected from beverages, pills, tablets, capsules, and powders, but is not limited thereto.

The health functional food composition of the present invention may be used as it is, or may be used with other foods or food ingredients, and may be appropriately used according to a conventional method. There is no particular limitation on the type of the health functional food composition. Examples of foods to which the hemp stem extract can be added include meat, sausage, bread, chocolate, candy, snacks, confectionery, pizza, ramen, other noodles, gum, dairy products including ice cream, various soups, beverages, tea, There are drinks, alcoholic beverages, vitamin complexes, etc., and includes all health functional foods in the ordinary sense. A health drink including the composition of the present invention may contain various flavoring agents or natural carbohydrates as additional ingredients like conventional drinks. The above-mentioned natural carbohydrates are monosaccharides such as glucose and fructose, disaccharides such as maltose and sucrose, polysaccharides such as dextrin and cyclodextrin, and sugar alcohols such as xylitol, sorbitol and erythritol. As the sweetener, natural sweeteners such as taumatine and stevia extract, synthetic sweeteners such as saccharin and aspartame, and the like can be used. The proportion of the natural carbohydrate is generally about 0.01 to 0.04 g, preferably about 0.02 to 0.03 g per 100 g of the composition of the present invention.

The health functional food composition of the present invention includes, in addition to the active ingredients, various nutrients, vitamins, electrolytes, flavoring agents, coloring agents, pectic acid and its salts, alginic acid and its salts, organic acids, protective colloidal thickeners, pH regulators, stabilizers, and preservatives. , glycerin, alcohol, a carbonation agent used in carbonated beverages, and the like may be further contained. In addition, it may further contain the pulp for the production of fruit juice or vegetable beverage. These components may be used independently or in combination. The proportion of these additives is not very important, but is generally selected in the range of 0.01 to 2 parts by weight based on 100 parts by weight of the composition of the present invention.

In addition, the present invention relates to a pharmaceutical composition for preventing or treating arteriosclerosis comprising a hemp stem extract as an active ingredient.

The pharmaceutical composition of the present invention may be in various oral or parenteral formulations. In the case of formulation, in addition to the active ingredient, a pharmaceutically acceptable carrier, excipient or diluent may be further included, and generally used fillers, extenders, binders, wetting agents, diluents or excipients such as disintegrants or surfactants are used. can be formulated. Solid preparations for oral administration include tablets, pills, powders, granules, capsules, etc., and these solid preparations include at least one excipient in one or more compounds, for example, starch, calcium carbonate, sucrose or lactose ( lactose), gelatin, etc. In addition to simple excipients, lubricants such as magnesium stearate and talc are also used. Liquid formulations for oral administration include suspensions, solutions, emulsions, syrups, etc. In addition to water and liquid paraffin, which are commonly used simple diluents, various excipients such as wetting agents, sweeteners, fragrances, and preservatives may be included. have. Formulations for parenteral administration include sterile aqueous solutions, non-aqueous solutions, suspensions, emulsions, freeze-dried preparations, and suppositories. Non-aqueous solvents and suspensions may include propylene glycol, polyethylene glycol, vegetable oils such as olive oil, and injectable esters such as ethyl oleate. As the base of the suppository, witepsol, macrogol, tween 61, cacao butter, laurin, glycero gelatin, etc. may be used.

The pharmaceutical composition of the present invention may be administered orally or parenterally, and it is preferable to select an external skin or intraperitoneal, rectal, intravenous, intramuscular, subcutaneous, intrauterine intrauterine or intracerebrovascular injection method for parenteral administration.

The pharmaceutical composition according to the present invention is administered in a pharmaceutically effective amount. In the present invention, "pharmaceutically effective amount" means an amount sufficient to treat a disease at a reasonable benefit/risk ratio applicable to medical treatment, and the effective dose level is the type, severity, and drug activity of the patient. , sensitivity to the drug, administration time, administration route and excretion rate, duration of treatment, factors including concurrent drugs, and other factors well known in the medical field. The pharmaceutical composition of the present invention may be administered as an individual therapeutic agent or in combination with other therapeutic agents, may be administered sequentially or simultaneously with conventional therapeutic agents, and may be administered single or multiple. In consideration of all of the above factors, it is important to administer an amount that can obtain the maximum effect with a minimum amount without side effects, which can be easily determined by those skilled in the art.

The dosage of the composition of the present invention varies depending on the patient's weight, age, sex, health status, diet, administration time, administration method, excretion rate and severity of disease, and the daily dosage is based on the amount of hemp stem extract Based on 0.01 to 2,000 mg/kg, preferably 30 to 500 mg/kg, more preferably 50 to 300 mg/kg, and may be administered 1 to 6 times a day. The pharmaceutical composition of the present invention may be used alone or in combination with methods using surgery, radiation therapy, hormone therapy, chemotherapy, and biological response modifiers.

Hereinafter, the present invention will be described in more detail using examples. These examples are only for illustrating the present invention in more detail, and it will be apparent to those of ordinary skill in the art that the scope of the present invention is not limited thereto.

[Cell Culture]

THP-1 cells, which are monocytes, were purchased from the American Type Culture Collection (ATCC) and contained 10% fetal bovine seum (FBS) and 100 U/ml penicillin/streptomycin (Thermo Fisher Scientific, Pittsburgh, PA, USA). Cultured in RPMI 1640 medium (Thermo Fisher Scientific, Pittsburgh, PA, USA), 200ng/ml PMA (phorbol 12-myristate 13-acetate) was added to the THP-1 cells and cultured for 24 hours to transform into macrophages. differentiation was induced.

Atherosclerosis induction of differentiated macrophages was induced by adding lysophosphatidylcholine 16:0 (LPC:16:0) using the method of Cha et al. (Cha et al. Steroids. 139:28-34.2018). .

Example 1. Hemp Stem Extraction

After 30 liters of water was added to 3 kg of hemp stems grown for about 2 months, hot water extraction was performed at 100-120° C. for 12 hours. This was lyophilized to obtain 108.69 g of extract.

Example 2. Confirmation of cytotoxicity

Using the CCK-8 kit, the cytotoxicity of the hemp stem extract was confirmed. 1×10 ⁵ cells were dispensed in a 96-well plate and treated with cannabis stem extract at a concentration of 50 to 600 μg/ml for 24 hours and 48 hours. After each hour, 10 μl of CCK-8 solution was administered, and incubated at 37° C. for 2 hours. Then, the absorbance was measured at a wavelength of 450 nm using an ELISA reader.

As a result, the hemp stem extract showed little toxicity in the arteriosclerosis model cells (FIG. 1).

Example 3. Confirmation of changes in intracellular lipid content

The intracellular lipid content was measured using the Oil-red O staining method. First, cells induced with PMA (phorbol 12-myristate 13-acetate) were treated with 40 μM of lysophosphatidylcholine (LPC) and 50 μg/ml of oxi-LDL. At the same time, the stem extract was treated up to 100-400 μg/ml. Thereafter, the supernatant was removed, fixed with 10% formalin, and stained with Oil-red-O for 10 minutes. After washing three times with distilled water, the intracellular oil-red-o was eluted using 100% isopropanol, and the absorbance was measured at 490/630 nm to determine the intracellular lipid content.

As a result, as shown in FIGS. 2 and 3, when LPC and oxi-LDL were treated compared to the control group, it was confirmed that the lipid accumulation increased by about 45% or more, and in contrast to this, when the hemp stem extract was treated, the concentration dependently decreased intracellular lipid content.

Example 4. Confirmation of Cholesterol-Related Gene Expression

RNA isolation was performed according to the Trizol method, and was isolated according to the manufacturer's method using an Easy spid-RNA extraction kit (Intron, Seoul, Korea). cDNA from each RNA was synthesized using the iScript cDNA synthesis kit (BioRad, Hercules, CA, USA), and the expression of each gene was performed using a CFX96 Real Time PCR instrument (BioRad, Hercules, CA, USA) and SsoAdvenced ^TM universal Sybrgreen supermix The expression of each gene was compared by q-PCR method using (BioRad, Hercules, CA, USA). The primer information used is shown in Table 1.

gene primer information gene directional sequence (5'->3') SEQ ID NO: LSS forward ACTTCGCCAGCATTGACTGG SEQ ID NO: 1 reverse GTCCACATACCAGCGCACAA SEQ ID NO: 2 MVD forward CGAGTCACACTGGCCTGAAC SEQ ID NO: 3 reverse GGATGATGCGCCAGGAGATG SEQ ID NO: 4 HMGCR forward TGCTTTGGCTGCATGTCAGT SEQ ID NO: 5 reverse GCTATCCAGCGACTGTGAGC SEQ ID NO: 6 MVK forward GTGCTGGCCTTTCTTTACTT SEQ ID NO: 7 reverse GGAAGGCCCACTTGTTTATT SEQ ID NO: 8 ABCA1 forward CTGCTAAGGAGGGAGCCTTT SEQ ID NO: 9 reverse AAAAGGGCCACAAACTGTTG SEQ ID NO: 10 SREBF1 forward AGTGGAGGACACACTGACC SEQ ID NO: 11 reverse CAGGACAGGCAGAGGAAGAC SEQ ID NO: 12 SREBF2 forward CAAGCTTCTAAAAGGGCATCG SEQ ID NO: 13 reverse GAGAGGCACAGGAAGGTGTAG SEQ ID NO: 14 GAPDH forward GAGTCAACGGATTTGGTCGT SEQ ID NO: 15 reverse GTTGCATGGATGACCTTGG SEQ ID NO: 16

(1) Inhibition of expression of cholesterol synthesis-related genes

Expression of HMGCR (HMG-CoA reductase), MVK (Mevalonate kinase), MVD (Mevalonate Diphosphate Decarboxylase) and LSS (Lanosterol Synthase) genes involved in cholesterol biosynthesis in order to confirm the inhibitory effect of the hemp stem extract on intracellular cholesterol metabolism Change was confirmed.

The expression of MVK, MVD, and LSS genes increased by LPC was decreased in a concentration-dependent manner of the hemp stem extract. In the case of HMGCR, the expression change by LPC was increased to insignificant level, but when the hemp stem extract of the present invention was treated at a concentration of 400 μg/ml, it was confirmed that HMGCR expression was statistically significantly suppressed (Fig. 4). .

In addition, the genes are known to promote transcription by the transcription factor SREBP, the SREBP family consists of SREBF1 and SREBF2.

Therefore, it was confirmed that the gene expression change of the transcription factor according to the treatment of the hemp stem extract was confirmed. As a result, while there was no significant expression change in SREBF2, SREBF1 was increased by about 2 times or more by LPC, and normal by the hemp stem extract. It was confirmed that it decreased to the level (FIG. 5).

(2) Confirmation of increase in the expression of cholesterol excretion-related genes

In order to suppress the accumulation of cholesterol and lipids in the cell, the cholesterol inside the cell must be released to the outside. The gene expression level of ABCA1 was confirmed.

As a result, when the hemp stem extract of the present invention was treated, it was confirmed that the expression of the ABCA1 gene was significantly increased compared to the LPC-induced group (FIG. 6).

Example 5. CBD in Hemp Stem Extract ( cannabidiol ) and content analysis of THC (delta-9 tetrahydrocannabinol) compounds

About 150 kinds of CBD compounds are known from cannabis, and among them, delta-9 tetrahydrocannabinol (THC) is known as a narcotic substance that causes hallucinations.

In order to confirm the presence of CBD and THC-based compounds in the hemp stem extract used in the present invention, using HPLC, four basic CBD and THC-based compounds, CBDV (Cannabidivarin), CBDA (cannabidiolic acid), and THCV (Tetrahydrocannabivarin) ) and THCA (tetrahydrocannabinolic acid) were checked.

100% methanol was added to each of the four CBDVs, CBDA, THCV and THCA to make the final assay concentration 5 μg/ml, and the freeze-dried hemp stem hot water extract of the present invention was accurately weighed and 1 ml of water was added. After adding and dissolving at a concentration of 1 mg/ml, it was filtered using a 0.2 μm membrane filter, and 20 μl of the obtained filtrate was injected and analyzed.

Analysis was performed using HPLC (Prominence, Shimadzu) including a pump (LC-20AD), auto sampler (SIL-20A), column oven (CTO-20A) and UV/PDA detector (SPD-20A). For the analysis of the extract, a Kinetex C18 column (5 μm, 150×4.6 mm) was used, and the column temperature was maintained at 30°C. As the mobile phase, water (A) and acetonitrile (B) containing 50 mM ammonium formate were used, and the mobile phase was developed at a flow rate of 1.0 mL per minute. The optimized mobile phase is a solvent mixed in a ratio of water (A): acetonitrile (B) = 35:75, and UV absorbance at 210 nm was detected.

As a result, the peaks for the four types of CBDV, CBDA, THCV and THCA were not observed, so it was determined that there were no CBD and THC-based compounds (FIG. 7).

Example 6. Gas chromatography-mass spectrometry ( GC -MS) to analyze the content of CBD and THC-based compounds

In addition to CBDV, CBDA, THCV, and THCA confirmed in Example 5, various CBD and THC-based compounds may exist, and thus, the presence or absence of CBD and THC was confirmed through additional GC-MS analysis.

After accurately weighing the freeze-dried hemp stem hot water extract, 1 ml of methanol was added and dissolved to a concentration of 1 mg/ml, filtered using a 0.2 μm membrane filter, and 2 μl of the obtained filtrate was injected and analyzed.

For the GC-MS system used for the analysis, a mass spectrometer (5977A, Agilent) connected to gas chromatography (7890B, Agilent) was used, and an HP-5MS (30m×0.25mm, 0.25㎛ ID) column was used for component separation. did. The sample was ionized with 70 eV energy using the EI (Electronic Ionization) mode, and to improve the analytical sensitivity, the split ratio of the sample was analyzed at 1/10 to determine the retention time and mass spectrum of each sample. Confirmed. It was developed at a flow rate of 1.0 ml per minute using helium as the carrier gas. Other analysis conditions are shown in Table 2.

analysis conditions equipment GC (Agilent 7890B), MSD (Agilent 5977A) column HP-5MS (30m×0.25mm, 0.25μm ID) Oven 80℃(initial) → 80℃(1min hold) → 20℃/min (240℃) → 5℃/min (260℃) → 20℃/min (300℃, 10 min hold) Carrier gas He (99.99%) injection volume 2 μl injection temperature 280℃ Ionization mode EI (70 eV) MS Quad 300℃ TIC scan range 40 to 450 (m/z)

As a result of GC-MS analysis, no substance was detected at the CBD (Cannabidiol) site, and no THC was identified. Therefore, it was determined that there is no CBD and THC in the hemp stem extract, or a trace amount that cannot be detected even if there is (FIG. 8).

Compound names of each peak (1 to 7) from the GC-MS spectrum of the cannabis stem extract of the present invention disclosed in FIG. 8(A) Peak No. compound name Retention time (min) Content ratio of each peak in hemp stem extract (%) reliability
(%) One Octadecane 3.645 4.21 50 2 2,3-dihydrobenzofuran 4.544 4.25 64 3 2,4-di-tert-bytylphenol 6.476 33.44 97 4 methyl palmitate 8.759 2.64 94 5 methyl stearate 9.763 9.18 99 6 monopalmitin 12.216 11.04 93 7 2,3-dihydroxypropyl stearate 13.823 35.23 90

In Table 3, the reliability of each peak identified in the GC-MS analysis was selected by selecting a compound that is likely to appear at each peak position through mapping with the GC-MS database, and the reliability of the selected compound in % The closer to 100, the higher the reliability.

<110> Korea Institute of Oriental Medicine <120> Composition for preventing, ameliorating or treating arteriosclerosis containing Cannabis sativa stem extract as effective component <130> PN19442 <160> 16 <170> KopatentIn 2.0 <210> 1 <211> 20 <212> DNA <213> Artificial Sequence <220> <223> LSS-F <400> 1 acttcgccag cattgactgg 20 <210> 2 <211> 20 <212> DNA <213> Artificial Sequence <220> <223> LSS-R <400> 2 gtccacatac cagcgcacaa 20 <210> 3 <211> 20 <212> DNA <213> Artificial Sequence <220> <223> MVD-F <400> 3 cgagtcacac tggcctgaac 20 <210> 4 <211> 20 <212> DNA <213> Artificial Sequence <220> <223> MVD-R <400> 4 ggatgatgcg ccaggagatg 20 <210> 5 <211> 20 <212> DNA <213> Artificial Sequence <220> <223> HMGCR-F <400> 5 tgctttggct gcatgtcagt 20 <210> 6 <211> 20 <212> DNA <213> Artificial Sequence <220> <223> HMGCR-R <400> 6 gctatccagc gactgtgagc 20 <210> 7 <211> 20 <212> DNA <213> Artificial Sequence <220> <223> MVK-F <400> 7 gtgctggcct ttctttactt 20 <210> 8 <211> 19 <212> DNA <213> Artificial Sequence <220> <223> MVK-R <400> 8 ggaaggccca cttgttatt 19 <210> 9 <211> 20 <212> DNA <213> Artificial Sequence <220> <223> ABCA1-F <400> 9 ctgctaagga gggagccttt 20 <210> 10 <211> 20 <212> DNA <213> Artificial Sequence <220> <223> ABCA1-R <400> 10 aaaagggcca caaactgttg 20 <210> 11 <211> 20 <212> DNA <213> Artificial Sequence <220> <223> SREBF1-F <400> 11 aggtggagga cacactgacc 20 <210> 12 <211> 20 <212> DNA <213> Artificial Sequence <220> <223> SREBF1-R <400> 12 caggacaggc agaggaagac 20 <210> 13 <211> 20 <212> DNA <213> Artificial Sequence <220> <223> SREBF2-F <400> 13 caagcttcta aagggcatcg 20 <210> 14 <211> 21 <212> DNA <213> Artificial Sequence <220> <223> SREBF2-R <400> 14 gagaggcaca ggaaggtgta g 21 <210> 15 <211> 20 <212> DNA <213> Artificial Sequence <220> <223> GAPDH-F <400> 15 gagtcaacgg atttggtcgt 20 <210> 16 <211> 20 <212> DNA <213> Artificial Sequence <220> <223> GAPDH-R <400> 16 gttgtcatgg atgaccttgg 20

Claims (5)

A health functional food composition for preventing or improving arteriosclerosis, comprising a hemp stem hot water extract as an active ingredient. delete The health function for preventing or improving arteriosclerosis according to claim 1, wherein the cannabis stem hot water extract is prepared in any one formulation selected from beverages, pills, tablets, capsules, and powders. food composition. A pharmaceutical composition for preventing or treating arteriosclerosis comprising a hemp stem hot water extract as an active ingredient. The pharmaceutical composition for preventing or treating arteriosclerosis according to claim 4, further comprising a pharmaceutically acceptable carrier, excipient or diluent in addition to the active ingredient.

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