KR101084939B1 - Composition comprising the extract of sea algae for preventing and treating hypertension - Google Patents

Abstract

The present invention relates to a composition for the prevention and treatment of hypertension containing seaweed extract as an active ingredient, and in detail, the extract of natural seaweed fronds, cultured seaweed fronds, kelp, seaweed ear or seaweed seaweed of the present invention is angiotensin converting enzyme By inhibiting (ACE), not only effectively suppresses the increase in blood pressure caused by the action of angiotensin converting enzyme, but also has high safety for the human body, the composition containing it is a pharmaceutical composition and health functional food useful for the prevention and treatment of hypertension. It can be used to.

Seaweed, Wakame Frond, Kelp, Wakame Ear, Cage Seaweed, Angiotensin Converting Enzyme (ACE), Hypertension

Description

Composition comprising the extract of sea algae for preventing and treating hypertension}

The present invention relates to a composition for the prevention and treatment of hypertension, which contains extracts of wild wakame fronds, cultured wakame fronds, kelp, seaweed ear or seaweed seaweed as active ingredients.

[Document 1] Lin Hyo et al., 2, 水産 加工 利用 學, 螢 雪 出版社, pp.277-278, 1994

[Ref. 2] Redundancy, Hanok Ok, Industrial Use of Seaweeds (1st Report) General Composition and Minerals of Seaweeds, Proceedings of SNU, 6 , p.325, 1972

[Reference 3] Jang Hyun-ki, Nam Gung-seok, Introduction to Food Science, Yurim Culture History, pp.118-119, 1991

[Ref. 4] Byun Jae-Hyung et al., 2, Production of Seaweed and Its Additives, 10 (2) , pp.125-135, 1977

[Reference 5] Yue Woon-Man, Hae-Soon Saeng-Sung Hwa University, Tokyo, Korea, Tokyo, pp.95-100, 1983

[6] Son Hong, et al. 3, Citrus Bioflavonoids Isolation, Purification and Blood Pressure Lowering Effect, Korean Journal of Nutrition and Nutrition, 21 (2) , p.136, 1992

[Ref. 7] Kimda-Shi, Deda-Suk-O, Yu-Sang-Ye-Sang, Kyu-Su-Kin-Sul-Koi-R.-I. Bulletin of the Japanese Society of Scientific Fisheries, 29 (11) , pp. 1020-1023, 1963

[Reference 8] ア 部 部 重 信, 武田 芙 美, 金 田志 水 産物 の ツ ロ ネ ス ミ コ レ ス テ ロ ― ル 代謝 に お よ ほ す 影響 に 關 す る 究 -Ⅶ. Bulletin of the Japanese Society of Scientific Fisheries, 33 (11) , pp.1050-1055, 1967

Oshima, G. et al., Peptide inhibitors of angiotensin-I converting enzyme in digests of gelatin by bacterial cillagenase. Biochim . Biophys . Acta , 566 , p. 128, 1979

Maruyama, S. et al., Angiotensin-I converting enzyme inhibitory activity of the C-terminal hexapeptide of αs1-casein. Agric. Biol. Chem., 51 , pp. 2557-2567, 1987

[11] Miyoshi, S. et al., Structures and activity of angiotensin-converting enzyme inhibitors in an α-zein hydrolysate. Agric. Biol. Chem., 55 , pp. 1313-1318, 1991

[Ref. 12] Yutada Kane, Nara-Yan-Ao, Family-owned Male, High School Self-Goverment, p.101, 1987

[Reference 13] Hypnotherapy 4, Effect of Buckwheat Supplementation on Blood Glucose and Blood Pressure in White Paper. Korean Journal of Nutrition and Nutrition, 20 (4) , p.300, 1991

[Jun 14] Park Joon-Hee et al. 4, A Study on the Blood Pressure Drop Components of Korean Ganoderma Lucidum, Journal of Food Hygiene, 2 (2) , p.57, 1987

[Reference 15] 齊藤 義 幸, 中 村 圭子, 川 戶 章 嗣, 今 安 聰, 酒, お よ ぴ 副 物 中 の ア ン ギ オ テ ソ ツ ソ 變換 酵素 沮 害 物質. Nippon NKaishi, 66 (7) , pp. 1081-1087, 1992

[16] Do Jeong-ryong et al. 3, Angiotensin-I Converting Enzyme Inhibitory Activity of Favorite Beverages, Journal of the Korean Society of Food Science and Technology, 25 (5) , pp.456-460, 1993

Maruyama, S. et al., Angiotensin-I converting enzyme inhibitors derived from Ficus Carica. Agric. Biol. Chem., 53 (10) , p. 2763, 1989

[Research 18] Heung-Soo Yoo, Kang-Ho Lee, Seaweed Protein Extraction 1. Extraction of Water-Soluble Proteins, Journal of the Korean Fisheries Society, 10 , p.151, 1977

[Ref. 19] Yoo Heung-soo et al. 2, A Study on Seaweed Protein Extraction 2. Extraction of Salt-Soluble and Alcohol-Soluble Proteins, Journal of the Korean Fisheries Society, 10 , p.189, 1977

[20] A Study on the Extraction of Seaweed Proteins 3. Yoo Heung-soo et al 2, Extraction of NaOH Soluble Proteins, Journal of the Korean Fisheries Society, 11 , p.85, 1978

[21] Woo-Jung Kim et al. 2, Development of a comprehensive extraction method for effective extraction of kelp, KOREAN J. FOOD SCI. TECHNOL, 26 (1) , pp. 51-56, 1994

[22] Lee, Jung-Keun et al. 3, Effect of Extraction Temperature on the Characteristics of Kelp Extract, Korean Journal of Nutrition and Nutrition, 22 (6) , pp.771-776, 1993

[23] Gu, Jae-Keun et al. 3, Extraction and Purification of Fucoidan from Korean Kelp and Seaweed, J. Korean Fish. Soc. 28 (2) , pp.227-236, 1995

[24] Kim Jun-pyung, Development of Proteins in Non-edible Seaweeds 1. Protein Extraction Conditions and Amino Acid Composition of Isolated Crude Proteins in Non-edible Seaweeds, KOREAN J. FOOD SCI. TECHNOL, 6 (1) , 1974

Cushman, DW and Cheumg, HS, Spectrophotometric assay and properties of the angiotensin converting enzyme of rabbit lung, Biochem. Pharm., 20 , p. 1637, 1971

26 Toshiro Matsui, et al., Colorimetric Measurement of Angiotensin-I Converting Enzyme Inhibitory Activity with Trinitrobenzene Sulfonate Biosci, Biotech. Biochem., 56 (3) , pp. 517-518, 1992

[Reference Document 27] 鈴木 建 夫, 石川宣 子, 目 黑 熙, 食品 中 の ア ソ ゾ オ テ ソ シ ソ I 轉換 沮 害 能 に つ ぃ て. Japan, 57 (11) , pp.1143-1146, 1983

[Reference 28] Hyun-Kyu Cho et al. 5, Food Analysis Method (1), p.246, 1993

[Reference 29] Kosuke Kobe, Food Analysis Hand book, Kenpakusha, Tokyo, pp.53-60, 1969

In Korea, seaweed has been used as food, fertilizer, medicinal food, feed, fertilizer, and raw material for the seaweed industry since ancient times. In recent years, it has been in the spotlight as a so-called health food and a bioactive substance raw material used by extracting a specific ingredient from seaweed. In practice, however, there are not many kinds of seaweeds used for this purpose, and only about 60 to 70 kinds are used (林浩 et al. 2, 水産 加工 利用 學, 螢 雪 出版社, pp.277-278, 1994).

Seaweeds contain high amounts of taste, so they have high value as natural seasonings. However, most of seaweed carbohydrates are non-digestible complex polysaccharides, which are relatively stable to acids and alkalis and are difficult to decompose without special bacterial enzymes. (Dualization, Industrial Use of Seaweeds, Industrial Use of Seaweeds (1st Report) General Composition and Minerals of Seaweeds, Proceedings of SNU, 6 , p.325, 1972).

The most common seaweeds and kelp among seaweeds are carbohydrates such as mannit and alginic acid, which contain about 12-13% protein, while iodine contains only 0.34% kelp but only 0.03%. About half of the seaweed and seaweed are carbohydrates, 20% of which are fibrous and the remainder are alginic acid and fucoidan (Jang Hyun-gi, Nam Gung-seok, Introduction to Food Science, Yurim Culture History, pp.118-119, 1991).

The differences in the components of the cultured seaweeds and wild seaweeds were studied, and the content of chlorophyll and carotenoids was much lower than that of wild seaweeds, and the content of free amino acids was lower than that of wild seaweeds. . In addition, both aquaculture and natural seaweed are high in threonine, alanine and glutamic acid, and almost no serine and proline, and histidine and cysteine. ), Tyrosine and phenylalanine were found to be extremely low. Glutamic acid and aspartic acid showed significant differences between natural and cultured seaweeds, and essential amino acids and soluble minerals. (About 12%) is excellent. Mannitol and soluble minerals in seaweed were reduced after blanching treatment, and chlorophyll and carotenoids were known to be retained (Strains et al., 2, Seaweeds, and their industrial processes, 10 (2) , pp. 125 -135, 1977).

Recently, studies on physiologically active substances to clarify the regulation of the biological function of food ingredients have been actively conducted. When examining the physiological activity of the seaweed components, porphysin extracted from sesame seeds is shay ) The ulcer was completely inhibited as a dose of 5 mg in a rat (200 g body weight) experiment. Vercosin extracted from chyloviruses showed a 12.5% inhibition rate for shay ulcers and 37.5% for stress ulcers. It is known that the rate of deterrence of the larvae has been shown to be low (坂 上 良 男, 海 恒 の 生 化學 と 利用, 恒星 社 厚生 閣, 東京, pp.95-100, 1983).

As a result of the specificity of blood pressure change in rats of Sprague-Dawley species, which are experimental animals, hesperidin, a major bioflavonoid component of tangerine peel, was significantly increased after administration of drugs. p <0.001) reported that the blood pressure lowering effect was observed (Shon et al. 3, isolates, purification and lowering blood pressure of bioflavonoids of citrus fruits, Journal of the Korean Nutrition Society, 21 (2) , p.136, 1992),

After 10 weeks of drying and powdering green laver (Enteromorpha compessa), tangle (Laminaria japonica), and purple laver (Pophyra tenera), rats were fed with cholesterol at 5% ratio, resulting in green laver and purple laver. Certainly reduced rat plasma cholesterol, tangle was ineffective, and this algae were reported to be ineffective in lowering liver liver cholesterol in rats. Niagara-I. 響 用 藻 海藻 の 效果, Bulletin of the Japanese Society of Scientific Fisheries, 29 (11) , pp. 1020-1023, 1963).

In addition, we compared the amount of cholesterol in the excretion of untreated rats fed with purple laver to determine the metabolism of the significantly lower plasma cholesterol levels in rats fed Porphyra tenera. It has been reported that a lot of cholesterol has been confirmed. (ア 部 重 信, 武田 芙 美, 金 田志 水 産物 の ツ ロ ネ ス ミ コ レ ス テ ロ ー ル 代謝 に お ほ す 影響 に 關 す る 究 -Ⅶ. Bulletin of the Japanese Society of Scientific Fisheries, 33 (11) , pp.1050-1055, 1967).

Hypertension is classified into primary hypertension and secondary hypertension. Primary hypertension, again, can be divided into mild, moderate, and severe, depending on the severity of the symptoms, which can be treated by reducing salt intake and relieving weight control, stress, and lack of exercise. Concurrent blood pressure lowering agents should be given at the same time as therapy. On the other hand, in the case of primary hypertension with a large genetic factor, it was known to limit the protein. However, in order to maintain nitrogen equilibrium, it is necessary to supply high quality protein as long as the function of kidney is able to excrete nitrogen, which is a protein decomposition product. Therefore, the separation of angiotensin converting enzyme (kinase II, Peptidyldipeptide hydrolase, EC 3.4.15.1, ACE hereinafter) inhibitory peptide from food proteins seems to have a strong relationship.

Angiotensin-1 converting enzyme (ACE), one of the factors that increase blood pressure, degrades and inactivates bradykinin, which has a blood pressure lowering effect, and inactivates angiotensin-1 C terminal dipeptides (His-leu). ), Which is activated by angiotensin-II, which exhibits strong blood pressure elevation, causes hypertension. An angiotensin converting enzyme (ACE) inhibitory peptide derived from the food protein was found from hydrolysates of gelatin (Oshima, G. et al. ., Peptide inhibitors of angiotensin-I converting enzyme in digests of gelatin by bacterial cillagenase. Biochim. Biophys. Acta, 566, p.128, 1979). It is known to exist mainly in the hydrolyzate of food protein, and such an active ingredient hydrolyzate is casein (Maruyama, S. et al., Angiotensin-I converting enzyme inhibitory activity of the C-terminal hexapeptide of αs1-). casein.Agric.Biol.Chem., 51 , pp.2557-2567, 1987), zein, a corn protein (Miyoshi, S. et al., Structures and activity of angiotensin-converting enzyme inhibitors in an α-zein hydrolysate.Agric.Biol.Chem., 55 , pp.1313-1318, 1991), etc., peptides derived from shrimp (池田 克己, ナ 良 安 雄, 家 森 幸 男, 高血壓 自然 發症 學會 抄錄, p.101, 1987) , Buckwheat Flour (Effect of Hypnosis 4, Effect of Buckwheat Supplementation on Blood Glucose and Blood Pressure in White Paper. Journal of the Korean Society of Nutrition, 20 (4) , p.300, 1991), Ethanol Available Fraction of Ganoderma lucidum (Jun Hee et al. 4, Korean Ganoderma lucidum) Of Blood Pressure Lowering Components in Korean Journal of Food Hygiene, 2 (2) , p.57, 1987), Cheongju (齊 酒, 中 村 圭子, 川 戶)章 嗣, 今 安 聰, 酒, お よ ぴ 副 物 中 の ア ン ギ オ テ ソ ツ ソ 變換 酵素 沮 害 物質 .Nippon NKaishi, 66 (7) , pp.1081-1087, 1992). ACE-inhibiting effect of the water-soluble fraction of ginger>senticosus>Omija>perilla> Cassia tora>quince> was reported appeared to jujube net (dojeongryong outer 3, Angiotensin symbol beverage ingredients - I-converting enzyme inhibitory activity, Journal of the Korea Food and 25 (5 ), pp.456-460, 1993), others find an ACE inhibitory peptides from various food has been made the study (Maruyama, S. et al, Angiotensin -. I converting enzyme inhibitors derived from Ficus Carica. Agric. Biol. Chem., 53 (10) , p. 2763, 1989).

Looking at the research reports on effective extraction methods for useful substances in seaweeds, Yu et al. Reported the extraction of water-soluble and salt-soluble, alcohol-soluble, and NaOH-soluble proteins in seaweeds (Yung Heung-soo, Lee Kang-ho, Seaweed Protein Extraction 1). Extraction of Water Soluble Proteins, Journal of the Korean Fisheries Society, 10 , p.151, 1977; Yoo Heung-soo et al. 2, A Study on Seaweed Protein Extraction 2. Extraction of Salt Soluble and Alcohol Soluble Proteins, Journal of the Korean Fisheries Society, 10 , p.189, 1977 Yoo Heung-soo et al. 2, Seaweed Protein Extraction 3. Extraction of NaOH Soluble Proteins, Journal of the Korean Fisheries Society, 11 , p.85, 1978), Kim et al. When the kelp is decomposed using the polysaccharide degrading enzymes viscozyme, celluclast, ultrazyme, it is recommended to mix viscozyme and celluclast in a 1: 1 ratio and then to decompose It was reported that the most appropriate in terms of rate. Soluble solids and protein yield increased as the temperature increased during kelp hot water extraction (60 ~ 100 ℃), and gradually increased after 1 hour of extraction (Kim Woo et al. Development of a Comprehensive Extraction Method for Effective Extraction, KOREAN J. FOOD SCI.TECHNOL, 26 (1) , pp.51-56, 1994; Lee, Jung-Keun et al. 3, Effect of Extraction Temperature on the Characteristics of Kelp Extract, Korean Nutrition Journal, 22 (6) , pp.771-776, 1993). In addition, Koo et al reported that the extraction yield of fucoidan from Korean kelp and seaweed was 2.71%, seaweed spore leaf 6.65%, and seaweed frond 0.40%. Extraction and Purification of Fucoidan from Seaweed, J. Korean Fish.Soc. 28 (2) , pp.227-236, 1995).

In addition, Kim (19) extracted the protein contained 19% in the perforated seaweed and 17% in the open seaweed of non-edible seaweed, the extraction conditions of crude protein is better to extract with water than Tris buffer, and the recovery and recovery of protein at the boiling point (Junpyeong Kim, Development of Proteins in Non-Edible Seaweeds) 1. Extraction Conditions of Proteins and Amino Acid Composition of Isolated Crude Proteins in Non-Edible Seaweeds, KOREAN J. FOOD SCI.TECHNOL, 6 (1) , 1974 .

On the other hand, research on the biological activity of seaweed is difficult in the seaweed for food many. This is because it is a demanding, cumbersome and uneconomical toxicity test that can only be recognized if it is available for safety testing. Even the seaweeds that are surprisingly frequently eaten are not well studied for the mass separation and purification of antihypertensive components.

Therefore, by trying to isolate and identify the edible seaweeds that have been routinely eaten for a long time, such as seaweed and kelp, where no toxic problems have been observed, the high value of seaweeds, such as the prevention of hypertension, one of the adult diseases, and the extraction of medicinal substances from seaweeds. I want to explore the use of enemies.

Accordingly, the present inventors have continued to study the effects of preventing and treating hypertension by extracts of seaweeds that are safe for toxicity by daily edible and extractable from the east coast, and thus, wild seaweed fronds, cultured seaweed fronds, kelp, seaweed ear or By extracting anchovy wakame inhibits angiotensin converting enzyme (ACE), it was confirmed that effectively inhibit the rise in blood pressure caused by the action of angiotensin converting enzyme to complete the present invention.

In order to carry out the above object, the present invention provides a pharmaceutical composition for the prevention and treatment of hypertension containing seaweed extract as an active ingredient.

The present invention also provides a health functional food for the prevention and improvement of hypertension containing seaweed extract as an active ingredient.

Seaweeds as defined herein are natural Undaria pinnatifida, Cultured Undaria pinnatifida, Laminaria japonica, Sporophyll of Undaria pinnatifida or Agarum cribrosum, preferably seaweed (Laminaria japonica).

The extracts defined herein are water containing purified water, polar solvents such as C ₁ to C ₄ lower alcohols or polar solvent soluble extracts soluble in a mixed solvent thereof, preferably polar solvent soluble extracts such as water or methanol, More preferably, a room temperature water extract extracted with water at room temperature or a hot water extract extracted with water at high temperature, even more preferably, a macromolecular containing water containing a large amount of polymer material having a molecular weight of 20000 Daltons or more, specifically, glucose 0 mg / ml, nitrogen Room temperature water extract fraction containing high molecular weight with 0.01 ~ 0.02 mg%, 0.105 ~ 0.2 mg%, NH ₂ -N 10-20 mg% component analysis range or low molecular weight containing hydrothermal extract containing high molecular weight of less than 20,000 Dalton molecular weight Specifically, glucose 0-0.0005 mg / ml, nitrogen 0.01-0.05 mg%, protein 0.1-0.2 mg%, NH ₂ -N 10-20 mg% A macromolecular water containing high temperature extract fraction having a stone range; Also, the residue remaining after removing the polar solvent soluble extract is extracted with a nonpolar solvent such as ether, acetone, preferably an ether, acetone or a mixed solvent thereof, and more preferably a nonpolar solvent soluble extract obtained by acetone. It includes.

Hereinafter, the method for obtaining the extract of the present invention will be described in detail.

The extract of the present invention is a natural seaweed frond (Natural Undaria pinnatifida), cultured seaweed frond (Cultural Undaria pinnatifida), kelp (Laminaria japonica), seaweed spores (Seaweed; Sporophyll of Undaria pinnatifida) or dried Agarum cribrosum And a first step of grinding; Homogenized by adding an extraction solvent, such as water, about 1 to 1000 times (v / w), preferably about 50 to 200 times (v / w) to each of the ground, and then homogenized at room temperature for 30 minutes to 3 hours, preferably After extraction for 1 hour, the second step of obtaining a room temperature water extract (RWF-1) by vacuum freeze drying the supernatant obtained by centrifugation; Distilled water was added to the residue from which the supernatant liquid of the second step was removed, and extracted at 50 to 150 ° C., preferably at 100 ° C. for 30 minutes to 3 hours, preferably 1 hour, and the supernatant obtained by centrifugation was vacuum-dried and dried. A third step of obtaining an extract (HWF-1); The residue from which the supernatant of the third step was removed was extracted, filtered in the order of acetone, methanol, and concentrated to obtain acetone extract (AF-1) and methanol extract (MF-1). The algae extract of the present invention can be obtained.

Also, three pieces of natural Undaria pinnatifida, cultured Undaria pinnatifida, seaweed (Laminaria japonica), seaweed spores (Seaweed; Sporophyll of Undaria pinnatifida) or dried seaweed (Agarum cribrosum) First step; About 1 to 1000 times (v / w), preferably about 50 to 200 times (v / w) of distilled water was added to each of the ground products, and extracted at room temperature for 30 minutes to 3 hours, preferably 1 hour, and then filtered. Vacuum lyophilization of the filtrate thus obtained to obtain a room temperature water extract (RWF-2); The residue from which the filtrate of the second step was removed was extracted and filtered in the order of ether, acetone and methanol, and concentrated to obtain an ether extract (EF-2), acetone extract (AF-2) and methanol extract (MF-2), respectively. A third step of doing; Distilled water was added to the residue from which the filtrate of the third step was removed, and the supernatant obtained by extraction and centrifugation at 50 to 150 ° C., preferably at 100 ° C. for 30 minutes to 3 hours, preferably 1 hour was subjected to vacuum freeze drying. The seaweed extract of the present invention can be obtained through the extraction method (2) of the fourth step of obtaining the extract (HWF-2).

In addition, the macromolecular weight-containing hot water extract of the present invention or the low molecular weight-containing hot water extract may be extracted and separated and purified by filtration by molecular weight differences, such as ultrafiltration, which is conventional in the art, for the above-mentioned hot water extract or hot water extract.

The present invention provides a pharmaceutical composition for the prevention and treatment of hypertension, containing the extract obtained by the above extraction method as an active ingredient.

The extract of the present invention is a food that is edible to the general public, it is judged that the development potential as a medicine or health food as a safe food without problems such as toxicity and side effects.

In addition, it was confirmed that the extract of the present invention is effective in treating hypertension by effectively inhibiting an increase in blood pressure caused by the action of angiotensin converting enzyme by effectively inhibiting angiotensin converting enzyme (ACE).

The pharmaceutical composition of the present invention comprises 0.1 to 50% by weight of the extract based on the total weight of the composition.

However, the composition is not limited thereto, and may vary depending on the condition of the patient, the type of disease, and the progress of the disease.

The extract of the present invention is a drug that can be used with confidence even when taken for a long time for the purpose of prevention because there is little toxicity and side effects.

The compositions of the present invention may further comprise suitable carriers, excipients and diluents conventionally used in the manufacture of pharmaceutical compositions.

The compositions of the present invention can be used in the form of powders, granules, tablets, capsules, suspensions, emulsions, syrups, aerosols and the like, oral formulations, external preparations, suppositories, and sterile injectable solutions, respectively, according to conventional methods. Carriers, excipients and diluents that may be included in the composition comprising the extract include lactose, dextrose, sucrose, sorbitol, mannitol, xylitol, erythritol, maltitol, starch, acacia rubber, alginate, gelatin, calcium phosphate, calcium silicate , Cellulose, methyl cellulose, microcrystalline cellulose, polyvinyl pyrrolidone, water, methylhydroxybenzoate, propylhydroxybenzoate, talc, magnesium stearate and mineral oil. When formulated, diluents or excipients such as fillers, extenders, binders, wetting agents, disintegrating agents, and surfactants are usually used. Solid preparations for oral administration include tablets, pills, powders, granules, capsules, and the like, and the solid preparations may include at least one excipient such as starch, calcium carbonate, sucrose in the extract. ) Or lactose, gelatin and the like are mixed. In addition to simple excipients, lubricants such as magnesium styrate talc are also used. Examples of the liquid preparation for oral use include suspensions, solutions, emulsions, and syrups. In addition to water and liquid paraffin, simple diluents commonly used, various excipients such as wetting agents, sweeteners, fragrances, preservatives and the like may be included . Formulations for parenteral administration include sterilized aqueous solutions, non-aqueous solutions, suspensions, emulsions, freeze-dried preparations, and suppositories. As the non-aqueous solvent and suspending agent, propylene glycol, polyethylene glycol, vegetable oil such as olive oil, injectable ester such as ethyl oleate, and the like can be used. As the base of the suppository, witepsol, macrogol, tween 61, cacao butter, laurin butter, glycerogelatin and the like can be used.

The preferred dosage of the composition of the present invention varies depending on the condition and the weight of the patient, the degree of disease, the type of drug, the route of administration and the period of time, but can be appropriately selected by those skilled in the art. However, for the desired effect, the extract of the present invention is preferably administered at 0.5 g / kg to 5 g / kg, preferably 1 g / kg to 3 g / kg per day. The administration may be carried out once a day or divided into several doses. Accordingly, the dosage is not limited in any way to the scope of the present invention.

The composition of the present invention may be administered to mammals such as rats, mice, livestock, humans, and the like in various routes. All modes of administration may be expected, for example, by oral, rectal or intravenous, intramuscular, subcutaneous, intra-uterine or intracerebroventricular injections.

The present invention also provides a health functional food for the prevention and improvement of hypertension containing the extract obtained by the above extraction method as an active ingredient.

The composition containing the extract of the present invention can be used in various ways, such as drugs, foods and drinks for the prevention and improvement of hypertension. Examples of the foods to which the extract of the present invention can be added include various foods, beverages, gums, tea, vitamin complexes, health supplements and the like, and they can be used as powders, granules, tablets, capsules or beverages have.

Extract of the present invention may be added to food or beverage for the purpose of preventing and improving hypertension. At this time, the amount of the extract in the food or beverage is generally the health food composition of the present invention can be added to 1 to 5% by weight of the total food weight, the health beverage composition is 0.02 to 10 g, preferably based on 100 ml Can be added in a ratio of 0.3 to 1 g.

In addition to containing the extract as an essential ingredient in the indicated ratio, the health beverage composition of the present invention is not particularly limited in the liquid component and may contain various flavors or natural carbohydrates as additional ingredients, as in general beverages. Examples of the above-mentioned natural carbohydrates include monosaccharides such as disaccharides such as glucose and fructose such as maltose, sucrose and the like and polysaccharides such as dextrin, cyclodextrin and the like Sugar, and sugar alcohols such as xylitol, sorbitol, and erythritol. As flavoring agents other than those described above, natural flavoring agents (tauumatin, stevia extract (e.g., Rebaudioside A, glycyrrhizin, etc.) and synthetic flavoring agents (saccharin, aspartame, etc.) can be advantageously used. The proportion of natural carbohydrates is generally about 1 to 20 g, preferably about 5 to 12 g per 100 mL of the composition of the present invention.

In addition to the above-mentioned composition, the composition of the present invention can be used as a flavoring agent such as various nutrients, vitamins, minerals (electrolytes), synthetic flavors and natural flavors, coloring agents and intermediates (cheese, chocolate etc.), pectic acid and its salts, Salts, organic acids, protective colloid thickening agents, pH adjusting agents, stabilizers, preservatives, glycerin, alcohols, carbonating agents used in carbonated beverages and the like. In addition, the compositions of the present invention may contain flesh for the production of natural fruit juices and vegetable beverages. These components can be used independently or in combination. The proportion of such additives is not so critical, but is generally selected in the range of 0 to about 20 parts by weight per 100 parts by weight of the composition of the present invention.

Extracts of natural seaweed fronds, cultured seaweed fronds, kelp, seaweed or seaweed seaweed of the present invention inhibits angiotensin converting enzyme (ACE), effectively inhibiting blood pressure rise caused by the action of angiotensin converting enzyme, Because of its high safety, the composition containing the same can be used for pharmaceutical compositions and health functional foods useful for the prevention and treatment of hypertension.

Hereinafter, the present invention will be described in detail by reference examples, examples and experimental examples.

However, the following reference examples, examples and experimental examples are merely to illustrate the invention, the content of the present invention is not limited to the following reference examples, examples and experimental examples.

Example  1. Preparation of Seaweed Extract

Natural seaweed fronds (including stems, Natural Undaria pinnatifida, hereinafter referred to as “NUP”) were collected and dried in June in Jeong-dong, East Sea, and cultured seaweed fronds (CUP ”) were used in Uljin. Collected and dried in March, laminaria japonica (LJ) and seaweed spores (Seaweed; Sporophyll of Undaria pinnatifida, SUP) were harvested and dried in June in Jumunjin. Iron seaweed (Agarum cribrosum, hereinafter referred to as “AC”) was collected and dried in June at Goseong, sliced into 1cm squares, and then pulverized three times with a mixer and stored in a low temperature room.

1-1. Preparation of Extract by Extraction Method (1)

In the extraction method (1) shown in FIG. 1, 10 g of natural seaweed fronds (NUP), cultured seaweed fronds (CUP), kelp (LJ), seaweed ear (SUP), and seaweed seaweed (AC) prepared above were distilled water. Each 1000ml was added and homogenized for 1 minute, then extracted by stirring at room temperature for 1 hour, followed by centrifugation (HITACHI, 6000rpm, 15 ° C, 10min) to obtain a supernatant and a residue. The supernatant was vacuum-dried to obtain a room temperature extract (hereinafter referred to as “RWF-1”). 1000 ml of distilled water was added to the residue, followed by extraction in a water bath (KMC-1205 SW1, 100 ° C.) for 1 hour, followed by centrifugation (HITACHI). , 6000rpm, 15 ° C, 10min) to freeze-dried the supernatant to obtain a hot water extract (hereinafter referred to as "HWF-1"), the residue was extracted and filtered in the order of 1000ml acetone, 1000ml methanol. 4) and concentrated with a rotary vacuum evaporator (Japan, A-3S) to obtain acetone extract (hereinafter referred to as "AF-1") and methanol extract (hereinafter referred to as "MF-1"), respectively.

1-2. Preparation of Extract by Extraction Method (2)

In the extraction method (2) shown in Fig. 2, 10 g of the natural seaweed frond (NUP), cultured seaweed frond (CUP), kelp (LJ), seaweed ear (SUP), or seaweed seaweed (AC) prepared as described above are distilled water. 1000 ml of each was added and stirred for 1 hour at room temperature, followed by filtration (Toyo fiter No. 4) to obtain a filtrate and a residue. The filtrate was lyophilized to obtain room temperature extract (hereinafter referred to as “RWF-2”), and the residue was extracted and filtered in the order of 1000 ml of ether, 1000 ml of acetone, and 1000 ml of methanol, and a rotary vacuum evaporator (Japan, A-3S). ) To give an ether extract (hereinafter referred to as "EF-2"), acetone extract (hereinafter referred to as "AF-2") and methanol extract (hereinafter referred to as "MF-2"), respectively. 1000 ml of distilled water was added to the residue, and extracted for 1 hour in a water bath (KMC-1205 SW1, 100 ℃) and centrifuged (HITACHI, 6000rpm, 15 ℃, 10min). The supernatant was vacuum-dried to dry and the hot water extract (hereinafter, “HWF -2 ").

Cold water extracts (RWF-1 and RWF-2) and hot water extracts (HWF-1 and HWF-2) were dissolved in distilled water and used as sample liquid. Acetone extracts (AF-1 and AF-2) and ether extracts (EF- 2) was dissolved in acetone (HPLC Grade, UV Cutoff 330nm) and used as sample liquid. Methanol extracts (MF-1 and MF-2) were dissolved in DMSO (Dimethyl Solfoxide, C ₂ H ₆ SO) and stored at refrigeration temperature. Was used.

Reference Example  1. Preparation of reagents

Lung acetone powder (from Rabbit) and Hippuryl-His-Leu (Acetate Salt), which are antihypertensive reagents, were purchased from Sigma * (United States), and Trinitrobenzene Sulfonate (TNBS) was purchased from Tokyo. Other than that, the first-class reagent for analysis was purchased and used.

Experimental Example  1. Extraction yield measurement

Yield of the extract obtained in the above Example was calculated as the following formula (1) as a percentage of the total Soluble Solid content of the extract to the dried product of the sample used for extraction are shown in Table 1 and Table 2.

수율(%) =

Yield (%) =

As a result of measuring the yield of each extraction section obtained by sequentially extracting five samples for each solvent, as shown in Table 1, looking at the yield by sample solvent extraction section in the extraction method (1), room temperature water extract (RWF-1) In the case, the minimum cultured seaweed fronds (CUP) was 12.5%, and the maximum wild seaweed fronds (NUP) was up to 42.1%, and the average yield was 34%, but the hydrothermal extract (HWF-1) and acetone extract (AF -1) and methanol extracts (MF-1) showed average yields of 10.5%, 3.1%, and 2.8%, respectively, yielding very low yields compared to room temperature water extracts (RWF-1).

In addition, as shown in Table 2, the yield of each sample solvent extraction section in the extraction method (2), in the normal temperature water extract (RWF-2), the minimum cultured seaweed fronds (CUP) is 13.5%, the maximum seaweed return (SUP) 42.9%, the average yield was 28.6%, which was relatively high like the extraction method (1), ether extract (EF-1), acetone extract (AF-2), methanol extract (MF-2) and hot water extract (HWF). In -2), the average yield was 7.0%, 2.0%, 1.5%, and 9.2%, respectively, indicating that the yield was lower than that of RF-2.

Yield (%) RWF-1 HWF-1 AF-1 MF-1 Natural Wakame Fronds (NUP) 42.1 9.0 2.66 0.3 Cultured Seaweed Fronds (CUP) 12.2 11.7 3.6 0.5 Kelp (LJ) 37.7 12.3 1.0 10.4 Seaweed ear (SUP) 40.1 8.7 5.4 1.3 Iron Seaweed (AC) 38.2 10.8 2.8 1.6

Yield (%) RWF-2 EF-2 AF-2 MF-2 HWF-2 Natural Wakame Fronds (NUP) 33.4 7.8 0.6 0.9 10.5 Cultured Seaweed Fronds (CUP) 13.5 6.7 0.3 0.1 10.2 Kelp (LJ) 30.3 4.3 1.0 1.2 5.6 Seaweed ear (SUP) 42.9 11.5 7.7 3.1 13.4 Iron Seaweed (AC) 23.0 4.5 0.6 2.3 6.1

Experimental Example 2. Antihypertensive Test

The antihypertensive activity of the sample obtained in the above Example was confirmed by applying the method disclosed in the literature to inhibit the activity of Angiotensin-I Converting Enzyme.

2-1. Angiotensin-I convertiong enzyme (ACE) preparation

ACE coenzyme preparation was prepared according to the method of Cushman and Cheung (Cushman, DW and Cheumg, HS, Spectrophotometric assay and properties of the angiotensin converting enzyme of rabbit lung, Biochem. Pharm., 20 , p.1637, 1971). It prepared by the method shown.

That is, 10 ml of sodium borate buffer (PH 8.5) was added to 1 g of Lung acetone powder (rabbit, sigma Co.), and stirred at 5 ° C. for 24 hours, followed by centrifugation (10000 rpm, 30 min). It was stored and used.

2-2. Determination of Angiotensin-Ⅰ converting enzyme (ACE) inhibitory activity

To measure the ACE inhibitory activity, Toshiro Matsui, et al., Colorimetric Measurement of Angiotensin -I Converting Enzyme Inhibitory Activity with Trinitrobenzene Sulfonate Biosci, Biotech.Biochem., 56 (3) , pp .517-518, 1992).

That is, 50 μl of Hip-His-Leu (2.5 mM in borate buffer containing 200 mM NaCl, pH 8.3) was added to 25 μl of the sample solution, and then 50 μl of the ACE coenzyme solution prepared in Experimental Example 2-1 was added 50 times 37 It was incubated for 1 hour at ℃. 250 μl of 0.5 N HCl was added as a reaction stopper, and the pH was adjusted to 250 μl of Kolthoff buffer (0.1 M Na ₂ HPO ₄ -1.0 N NaOH (1: 2)), followed by incubation for 20 minutes with 25 μl of TNBS Solution. After the addition of Sulfite (4mM Na ₂ SO ₃ in 0.2M NaH ₂ PO ₄ ) to measure the absorbance at 416nm with an ultraviolet-visible spectrophotometer (Ultraviolet-visible Spectrophotometer; Beckman England) to determine the ACE inhibition rate using the following equation 2 The amount of ACE inhibitor required to inhibit the enzyme activity by 50% was calculated as IC ₅₀ , and this activity measurement pattern was shown in FIG. 4.

AS: Absorbance of Sample Addition

AB: Absorbance of Sample-Free Sphere

AC: absorbance of the control

Vol (μl) Blank Control Sample Inhibitor - - 25 H ₂ O 25 25 - ACE solution 50 50 50 0.5N HCl 400 - - 12.5mM Hip-His-Leu 150 150 150 Incubated at 37 ℃ for 90min 0.5N HCl - 400 400 Kolthoff buffer ^{* 1} 250 250 250 0.1M TNBS solution 25 25 25 Incubated at 37 ℃ for 20min Sulfite ^{* 2} 4500 4500 4500 Measured at 416 nm ^{* 1} 0.1 M Na ₂ HPO ₄ . 1.0N NaOH (1: 2)
^{* 2} 4 mM Na ₂ SO ₃ in 0.2 M NaH ₂ PO ₄

As shown in Table 4, in the extraction method (1), IC ₅₀ (μg / ml) was 12.4 (μg / ml) in the hot water extract (HWF-1) of kelp (LJ), and ACE at the lowest concentration. Inhibitory effects were observed in 15.9 (μg / ml) and 14.9 (μg) extracts from RWF-1 of wild wakame fronds (NUP), cultured wakame fronds (CUP), kelp (LJ), and boiled seaweed (AC). / ml), 15.5 (μg / ml), 13.3 (μg / ml), and 14.4 (μg / ml) of hot water extracts (HWF-1) of wild wakame fronds (NUP) and boiled seaweed (AC), respectively. ml), 13.3 (μg / ml). Acetone extract (AF-1) showed a high ACE inhibitory effect of 20.3 (μg / ml) on cultured wakame fronds (CUP). Methanol extract (MF-1) showed 27ACE (μg / ml) ACE inhibitory effect only in kelp (LJ).

In addition, as shown in Table 5, looking at the ACE inhibitory effect in the extraction method (2), the same as the extraction method (1) in the cold water extract (RWF-2) and hot water extract (HWF-2) of kelp (LJ) 10.4 (μg / ml), respectively, and high temperature water extract (RWF-2) showed 21.4 (μg / ml) of wild seaweed fronds (NUP), cultured seaweed fronds (CUP) and poached seaweeds (AC). , 14.4 (μg / ml), 24 (μg / ml) and relatively high.In ether extract (EF-2), natural seaweed fronds (NUP) and cultured seaweed fronds (CUP) were 30.2 (μg / ml), 68.7 (μg / ml) showed a relatively high effect, while sea tangle (LJ) and clasp seaweed (AC) were 103.9 (μg / ml) and 150.1 (μg / ml), showing a lower effect than seaweed. In acetone extract (AF-2), the cultured seaweed frond (CUP) was the highest at 31.4 (μg / ml), while the wild seaweed frond (NUP), kelp (LJ) and poached seaweed (AC) were 288.5 (μg / ml), respectively. ml), 184.0 (μg / ml), 336.8 (μg / ml), lower than cultured seaweed fronds (CUP). In methanol extract (MF-2), only kelp (LJ) showed 169.4 (μg / ml). In hot water extract (HWF-2), wild seaweed fronds (NUP) and cultured seaweed fronds (CUP) were relatively high at 16.3 (μg / ml) and 40.8 (μg / ml), respectively. Seaweed ear (SUP) was found to have a weak ACE inhibitory effect in all extracts as in the extraction method (1).

On the other hand, 鈴木, etc., the ACE inhibitory factor present in foods is stable to heat treatment, and is expected to be useful in that it is a low-molecular substance that is easily absorbed by the body, and its inhibitory activity is lower than blood pressure lowering agents, but it is present in regular foods. (보고 木 建 鈴, 石川宣 子, 目 黑 熙, 食品 中 の ア ソ ゾ オ テ ソ シ ソ I) 沮 害 能 に つ ぃ て. 日本 化學 會 志, 57 (11) , pp.1143-1146, 1983) In the experiments, the solvent extract extract having the maximum antihypertensibility by sample showed a high effect in the room water extracts (RWF-1 and RWF-2) and hot water extracts (HWF-1 and HWF-2), which are strong polar water extracts. In general, it was confirmed that the antihypertensive substance in seaweed was extracted from a polar solvent rather than a nonpolar solvent.

IC ₅₀ ^{* 1} (μg / ml) RWF-1 HWF-1 AF-1 MF-1 Natural Wakame Fronds (NUP) 15.9 14.4 177.0 ND ^{* 2} Cultured Seaweed Fronds (CUP) 14.9 9.1 20.3 N.D. Kelp (LJ) 15.5 12.4 110.1 272.7 Seaweed ear (SUP) N.D. N.D. N.D. N.D. Iron Seaweed (AC) 13.3 13.3 317.7 N.D. ^{* 1} IC ₅₀ means the sample concentration to inhibit 50% of ACE inhibitory ratio
^{* 2} ND: not detected

IC ₅₀ ^{* 1} (μg / ml) RWF-2 EF-2 AF-2 MF-2 HWF-2 Natural Wakame Fronds (NUP) 21.4 30.2 288.5 ND ^{* 2} 16.3 Cultured Seaweed Fronds (CUP) 14.4 68.7 31.4 N.D. 40.8 Kelp (LJ) 10.4 103.9 184.0 169.4 10.4 Seaweed ear (SUP) N.D. N.D. N.D. N.D. N.D. Iron Seaweed (AC) 24 150.1 336.8 N.D. N.D. ^{* 1} IC ₅₀ means the sample concentration to inhibit 50% of ACE inhibitory ratio
^{* 2} ND: not detected

Experimental Example  3. Ultra  Separation and Recovery of Bioactive Substances by Filtration

In Experimental Example 2, ultrafiltration was performed for mass separation and purification of the active substance against the extract having high antihypertensive activity. The ultrafiltration unit is a system with a plate and frame type module (DDS, Japan; KSA / ISO 9001 SAM YEON), with a membrane area of 0.0336 m ² , and the membrane used is FS 61 PP with a fractional molecular weight of 20,000. . The feed was continuously stirred during the experiment, and the temperature was kept constant before entering the module by connecting a constant temperature circulator to the heat exchanger. After the experiment was completed, the filter membrane was sufficiently washed with water until the initial water flux was restored.

The recovery yield of each component recovered in the permeate was calculated by the following equation.

C _p : solid content of permeate

C _F : solid content of treatment liquid

As a result of comparing ACE inhibitory effect with IC ₅₀ (㎍ / ml), sequential solvent extraction of extraction method (1) and extraction method (2) showed that the extracts of kelp (LJ) and hot water extracts of kelp (LJ) showed strong antihypertensive activity. An ultrafiltration device having a filter membrane cutoff of 20000 was used for the extraction of room temperature water (RWF-1) and hot water extract (HWF-1) of wild kelp (LJ) obtained from the extraction method (1) near the daily diet. After separation by separation, the natural water extract (RWF-1) and hot water extract (HWF-1) of natural kelp (LJ) were extracted into stock solution, high molecular weight (more than 20000 Daltons) and low molecular weight (more than 20000 Daltons) By dividing the recovery and ACE inhibitory effect is shown in Table 6 below.

As a result of the experiment, as shown in Table 6, the recovery was 9% for the molecular weight of more than 20000 Daltons, 78.8% for the molecular weight less than 20000 Daltons of the kelp room temperature water extract (LJ-RWF-1), the kelp hot water extract (LJ -HWF-1) was relatively high, with a molecular weight of at least 20000 Daltons being 64% and a molecular weight of at most 20000 Daltons being 28.7%. In addition, ACE inhibitory effect, IC ₅₀ (㎍ / ml) of the kelp hot water extract (LJ-HWF-1) has a molecular weight of 20000 Dalton or less 24.9 (㎍ / ml), the highest activity, the kelp water extract ( LJ-RWF-1) had a high activity of 25.1 (µg / ml) at a molecular weight of 20000 Dalton or more. On the other hand, the kelp room temperature extract (LJ-RWF-1) has a molecular weight of less than 20,000 Daltons of 297.6 (㎍ / ml), the activity was slightly lower than other extracts, but the absolute value, it was confirmed that the strong activity.

The antihypertensive activity of alginate and cellulose in seaweed was measured. Sodium alginate 300, sodium alginate 900 and IC ₅₀ (µg / ml) of cellulose were measured. Were identified as 134.6 (µg / ml), 111.1 (µg / ml), and 53.1 (µg / ml), respectively.

weight (% recovery) IC ₅₀ (μg / ml) Kelp Water Extract (LJ-RWF-1) (Stock) 0.2 g 316.5 Polymer (more than 20000 Da) 0.018g (0.9%) 25.1 Low molecular weight (below 20000 Da) 0.1575 g (78.8%) 297.6 Kelp Hot Water Extract (LJ-HWF-1) (Stock) 0.06 g 93.9 Polymer (more than 20000 Da) 0.0384g (64.0%) 42.4 Low molecular weight (below 20000 Da) 0.0172g (28.7%) 24.9 Sodium alginate 300 134.6 Sodium alginate 900 111.1 Cellulose 53.1

Experimental Example  4. Qualitative Detection of Seaweed Extracts

A 20% sodium carbonate solution is sprayed onto the sampled spot and a further ciluted Folin-ciocalteau reagent-H ₂ O (50:50, v / v) solution is used to spray blue spots. The phenol component was detected by.

In addition, 0.25 g of ninhydrin (triketohydridene) reagent is mixed with 100 ml of ethanol, sprayed on a spot where the sample liquid is spotted, and then heated to 120 to 150 ° C. for 10 to 15 minutes to develop an amino acid and an amino sugar in blue color. Detected amino acids and aminosaccharides, reddish violet colored lipids, and violet colored vitamins detected.

1 ml of a saturated solution of silver nitrate (AgNO ₃ -H ₂ O ≒ 22: 10, w / w) was dissolved in 253 ml of acetone and 2% NaOH solution. washed in _{5% Na 2 S 2 O 3} (sodium thiosulfate) was washed with water retention dried and then detecting the color of dark brown sugar by spot. Sugars appear as dark brown spots according to the reducing power, and the detection sensitivity is high enough to develop even 1 μg of glucose.

Experimental results, as shown in Table 7 below, the qualitative reaction of the polymer and low molecular weight to the kelp room temperature extract (LJ-RWF-1) and the hot water extract (LJ-HWF-1), Folin's detection indicator of the Phenol component In the reagents, all of the four sample solutions showed no reaction.The peptide, ACE inhibitory factor, was detected by the ninhydrin reagent, which is an indicator of detection of amino acid and peptide components, and as a result, low-temperature kelp extract (LJ-RWF-1) From the strong detection of the material, it was confirmed that the ACE inhibitory peptide, which is a low-molecular substance of kelp, was recovered by the extraction of room temperature water.

In addition, antihypertensive properties may be expressed as a sugar component, and sugar component detection was strongly detected in low molecular weight kelp extract (LJ-RWF-1) as a result of detecting a saturated aqueous solution of silver nitrate (AgNO ₃ ) as an indicator. It was confirmed that the keratin water extract (LJ-HWF-1) was also weakly detected in the low molecular weight material.

Kelp Water Extract (LJ-RWF-1) Kelp Hot Water Extract (LJ-HWF-1) Polymer
(More than 20000 Da) Low molecular weight
(20000 Da or less) Polymer
(More than 20000 Da) Low molecular weight
(20000 Da or less) Folin's Indicators - - - - Ninhydrin Reagent - +++ - - AgNO ₃ aqueous solution - +++ - + +++: dark color, +: light color

Experimental Example  5. Quantitative Component Analysis of Seaweed Extracts

5-1. Reducing Sugar Quantities

Equivalence was quantified using the Somogyi method (CHO Hyun et al. 5, Food Analysis Method (1), p.246, 1993). That is, 5 ml of sample solution A (Na ₂ CO ₃ : C ₄ H ₄ O _{6 ·} K · Na · 5H ₂ O = 25 g: 25 g) was added to 500 ml of water, and then 10 ml of a 10% CuSO ₄ solution, 20 g of NaHCO ₃ , Add 5 ml of KI 5g, 200 ml of N / 10 KIO ₃ solution to 11), add 5 ml, heat in a water bath for 15 minutes, cool for 3 minutes, add 1 ml of 5H H ₂ SO _4, and cool for 2 minutes. 0.1ml was added, titrated in B solution (20 times diluted N / 10 Na ₂ S ₂ O ₃ ), and the equivalent was quantified using Equation 4 below.

B: blank titration value (ml)

A: Sample liquid titration value (ml)

f: 0.005N-Factor of Na ₂ S ₂ O ₃

D: dilution drainage

S: sample sample (g)

5-2. Protein Quantitation

Proteins were quantified using Kjeldahl nitrogen assay (Cho Hyun-kyu et al. 5, Food Assay (1), p.246, 1993). That is, 5 ml of the sample solution was water-substituted with a decomposition accelerator and 25 ml H ₂ SO ₄ and titrated with 0.1 N NaOH to quantify the protein using the following Equations 5 to 7.

V ₀ : ml of 0.1N NaOH consumption of blank test

V ₁ : ml of 0.1N NaOH consumption required for this experiment

f: 0.1N-NaOH standard solution factor

S: Sample Collection

N: Nitrogen conversion factor

5-3. Aminotenitrogen ( NH ₂ -N) quantification

Copper salt method (小 原 哲 二 郞, Food Analysis Hand book, Kenpakusha, Tokyo, pp.53-60, 1969) is used to form a blue complex by the reaction of amino acids and peptides with Cu. Substituted with alanine Cu-complex salt to remove the difference in absorbance between each amino acid and absorbance at 620 nm was measured.

That is, 5 ml of the sample solution was added to 70 ml of water, heated in a water bath for 20 minutes, cooled for 10 minutes, centrifuged, 5 ml of the supernatant was added, 5 ml of copper phosphate suspension was added, and centrifuged again. The supernatant was 620 nm. The absorbance was measured to quantify aminotenitrogen using Equation 8 below.

1260/13: Slope on calibration curve

D: dilution factor

As a result of the experiment, as shown in Table 8, sugar, protein, and aminotetrogen of high-molecular and low-molecular substances separated by ultrafiltration from kelp room temperature water extract (LJ-RWF-1) and hot water extract (LJ-HWF-1) It was confirmed that the content ratio of (NH ₂ -N) was maintained at a similar level between the two molecular weight groups.

In other words, it can not be said that the separation and purification was still achieved only by ultrafiltration, but as shown in Table 6 above, as a result of measuring the antihypertensive activity of the extract, in the case of kelp extract (LJ-RWF-1), ultrafiltration Since only highly active fractions could be obtained by themselves, kelp was extracted from the room temperature water extract (LJ-RWF-1) and hot water extract (LJ-HWF-1) because it is highly viscous to separate and purify by general column method. Separation of active substance by sewage can be said to have great significance.

Glucose
(mg / ml) nitrogen
(mg%) protein
(%) NH ₂ -N
(mg%) Kelp Water Extract (LJ-RWF-1) Polymer (more than 20000 Da) 0 0.0168 0.105 13.956 Low molecular weight (below 20000 Da) 0.001 0.0182 0.1138 13.375 Kelp Hot Water Extract (LJ-HWF-1) Polymer (more than 20000 Da) 0.0005 0.0238 0.1488 8.14 Low molecular weight (below 20000 Da) 0.0002 0.0238 0.1488 14.538

Hereinafter, an example of the preparation of the composition comprising the seaweed extract of the present invention, but the present invention is not intended to limit it, it is intended to explain in detail only.

Formulation Example 1 Preparation of Powder

Kelp Room Temperature Extract (LJ-RWF-1) 20 mg

Lactose 100 mg

Talc 10 mg

The above ingredients are mixed and filled in an airtight cloth to prepare a powder.

Formulation Example 2 Preparation of Tablet

Kelp Hot Water Extract (LJ-HWF-1) 10 mg

Corn starch 100 mg

Lactose 100 mg

2 mg magnesium stearate

After mixing the above components, tablets are prepared by tableting according to a conventional method for preparing tablets.

Formulation Example 3 Preparation of Capsule

Kelp Water Extract (LJ-RWF-1) 10 mg

3 mg of crystalline cellulose

Lactose 14.8 mg

Magnesium Stearate 0.2 mg

According to a conventional capsule preparation method, the above ingredients are mixed and filled into gelatin capsules to prepare capsules.

Formulation Example 4 Preparation of Injection

Kelp Hot Water Extract (LJ-HWF-1) 10 mg

Mannitol 180 mg

Sterile distilled water for injection 2974 mg

Na ₂ HPO ₄ , 12H ₂ O 26 mg

According to the conventional method for preparing an injection, the amount of the above ingredient is prepared per ampoule (2 ml).

Formulation Example 5 Preparation of Liquid

Kelp Room Temperature Extract (LJ-RWF-1) 20 mg

10 g of isomerized sugar

5 g of mannitol

Purified water

After dissolving each component in purified water according to the usual method of preparing a liquid solution, adding lemon flavor appropriately, mixing the above components, adding purified water, adjusting the whole to 100 ml by adding purified water, and then filling into a brown bottle. The solution is prepared by sterilization.

Formulation Example 6 Preparation of Healthy Food

Kelp Hot Water Extract (LJ-HWF-1) 1000 mg

Vitamin mixture quantity

70 [mu] g of vitamin A acetate

Vitamin E 1.0 mg

0.13 mg vitamin B1

0.15 mg of vitamin B2

0.5 mg vitamin B6

0.2 [mu] g vitamin B12

10 mg vitamin C

Biotin 10 μg

Nicotinic acid amide 1.7 mg

50 ㎍ of folic acid

Calcium pantothenate 0.5 mg

Mineral mixture quantity

1.75 mg of ferrous sulfate

0.82 mg of zinc oxide

Magnesium carbonate 25.3 mg

15 mg of potassium phosphate monobasic

Secondary calcium phosphate 55 mg

Potassium citrate 90 mg

100 mg of calcium carbonate

Magnesium chloride 24.8 mg

Although the composition ratio of the above-mentioned vitamin and mineral mixture is comparatively mixed with a composition suitable for health food as a preferred embodiment, the compounding ratio may be arbitrarily modified, and the above ingredients are mixed according to a conventional method for producing healthy foods , Granules can be prepared and used in the manufacture of health food compositions according to conventional methods.

Formulation Example 7 Preparation of Healthy Drink

Kelp Water Extract (LJ-RWF-1) 1000 mg

Citric acid 1000 mg

100 g of oligosaccharide

Plum concentrate 2 g

Taurine 1 g

Purified water was added to a total of 900 ml

The above components were mixed according to a conventional health drink manufacturing method, and the mixture was heated at 85 DEG C for about 1 hour with stirring, and the solution thus prepared was filtered to obtain a sterilized 2-liter container, which was sealed and sterilized, &Lt; / RTI &gt;

Although the composition ratio is a mixture of the components suitable for the preferred beverage as a preferred embodiment, the blending ratio may be arbitrarily varied according to the regional and national preferences such as the demand level, the demanding country, and the intended use.

[National R & D project supporting this invention]

[Task unique number]

RRC00200-0000-00 / B0010517-2008-01 / F20815508H220000110

[Name of Buddha]

Ministry of Knowledge Economy / Ministry of Knowledge Economy / Korea Maritime Affairs and Fisheries Technology Development Institute

[Name of research project]

Regional Cooperation Center Project / Regional Industry Promotion Project / Fisheries Specific R & D Industry

[Name of Research Project]

Donghae Coastal Marine Biological Resources Research Center Performance Utilization Project / Sokcho Well-being Salted Fish Specialty Product Development Project / High-Quality Processed Marine Products of Essence Oil Applications as Additives and Commercialization of Functional Materials

[Host]

Gangneung-Wonju National University / Gangneung-Wonju National University

[Research period]

January 01, 2009-December 30, 2009 / July 01, 2008-June 30, 2011 / December 01, 2008-November 30, 2011

1 is a view showing a manufacturing process of the extract by the extraction method (1),

2 is a view showing a manufacturing process of the extract by the extraction method (2),

Figure 3 is a diagram showing the process of preparing angiotensin-I convertiong enzyme (ACE) coenzyme solution,

4 is a diagram showing the absorbance (against borate buffer at 416 nm) of TNMS (-, blank) and TNP-HIs-Leu (-, control),

5 is a diagram showing the Angiotensin-Iconverting enzyme (ACE) inhibitory activity of the extract of room temperature (RWF-1) by the extraction method (1),

6 is a diagram showing the Angiotensin-Ⅰconverting enzyme (ACE) inhibitory activity of the hot water extract (HWF-1) by the extraction method (1),

7 is a diagram showing angiotensin-Ⅰconverting enzyme (ACE) inhibitory activity of acetone extract (AF-1) by the extraction method (1),

8 is a diagram showing the Angiotensin-Ⅰconverting enzyme (ACE) inhibitory activity of the room temperature water extract (RWF-2) by the extraction method (2),

9 is a diagram showing the Angiotensin-Iconverting enzyme (ACE) inhibitory activity of the ether extract (EF-2) by the extraction method (2),

10 is a diagram showing the angiotensin-Ⅰconverting enzyme (ACE) inhibitory activity of acetone extract (AF-2) by the extraction method (2),

11 is a diagram showing the Angiotensin-Ⅰconverting enzyme (ACE) inhibitory activity of the hot water extract (HWF-2) by the extraction method (2),

12 is a result of detection of amino acids and peptides of high-molecular and low-molecular substances separated from kelp room temperature water extract (LJ-RWF-1) and hot water extract (LJ-HWF-1) by ultrafiltration (R-1: kelp room temperature water extract ( Polymer of LJ-RWF-1) / R-2: Low molecular material of kelp room temperature extract (LJ-RWF-1) / H-1: Polymer of kelp hot water extract (LJ-HWF-1) / H-2: It is a diagram showing the low molecular weight material of kelp hot water extract (LJ-HWF-1),

FIG. 13 is a diagram illustrating a detection result of sugar components of high-molecular substances and low-molecular substances in which kelp water extract (LJ-RWF-1) and hot water extract (LJ-HWF-1) are separated by ultrafiltration.

Claims (7)

Cutting and grinding the dried seaweed (Laminaria japonica) or Agarum cribrosum dried seaweed from East Sea; 50 to 200 times (v / w) of water was added to the pulverized product, homogenized, extracted for 30 minutes to 3 hours at room temperature, and the supernatant obtained by centrifugation was vacuum-dried and dried to obtain room temperature extract (RWF-1). Obtaining a second step; Hot water extract (HWF-1) obtained by adding distilled water to the residue from which the supernatant was removed in the second step and extracting at 50 to 150 ° C. for 30 minutes to 3 hours, and then vacuum-drying the supernatant obtained by centrifugation; Or a first step of cutting and pulverizing the seaweed dry matter; 50 to 200 times (v / w) of distilled water was added to each of the pulverized products, and extracted at room temperature for 30 minutes to 3 hours, and then the filtrate obtained by filtration was lyophilized under vacuum to obtain room temperature water extract (RWF-2). step; The residue from which the filtrate of the second step was removed was extracted and filtered in the order of ether, acetone and methanol, and concentrated to obtain an ether extract (EF-2), acetone extract (AF-2) and methanol extract (MF-2), respectively. A third step of doing; Distilled water was added to the residue from which the filtrate of the third step was removed, extracted for 30 minutes to 3 hours at 50 to 150 ° C., and the supernatant obtained by centrifugation was vacuum-dried to obtain a hot water extract (HWF-2). A high-temperature water-containing high temperature extract (RWF-1) containing a high molecular weight of at least 20,000 Daltons of molecular weight extracted from kelp (Laminaria japonica) or Agarum cribrosum of East Sea, obtained through the extraction method of (2) , For prevention and treatment of hypertension, which contains a high-temperature water extract (RWF-2) or a low molecular weight-containing hydrothermal extract (HWF-1) or a hydrothermal extract (HWF-2) containing a large amount of low-molecular weight substances with a molecular weight of 20000 Daltons or less as an active ingredient Pharmaceutic composition. delete delete delete delete Cutting and grinding the dried seaweed (Laminaria japonica) or Agarum cribrosum dried seaweed from East Sea; 50 to 200 times (v / w) of water was added to the pulverized product, homogenized, extracted for 30 minutes to 3 hours at room temperature, and the supernatant obtained by centrifugation was vacuum-dried and dried to obtain room temperature extract (RWF-1). Obtaining a second step; Hot water extract (HWF-1) obtained by adding distilled water to the residue from which the supernatant was removed in the second step and extracting at 50 to 150 ° C. for 30 minutes to 3 hours, and then vacuum-drying the supernatant obtained by centrifugation; Or a first step of cutting and pulverizing the seaweed dry matter; 50-200 times (v / w) of distilled water was added to each of the ground products, followed by extraction for 30 minutes to 3 hours at room temperature, and the filtrate obtained by filtration was lyophilized under vacuum to obtain room temperature water extract (RWF-2). step; The residue from which the filtrate of the second step was removed was extracted and filtered in the order of ether, acetone and methanol, and concentrated to obtain an ether extract (EF-2), acetone extract (AF-2) and methanol extract (MF-2), respectively. A third step of doing; Distilled water was added to the residue from which the filtrate of the third step was removed, extracted for 30 minutes to 3 hours at 50 to 150 ° C., and the supernatant obtained by centrifugation was vacuum-freeze-dried to obtain a hydrothermal extract (HWF-2). A high-temperature water-containing high temperature extract (RWF-1) containing a high molecular weight of at least 20,000 Daltons of molecular weight extracted from kelp (Laminaria japonica) or Agarum cribrosum of East Sea, obtained through the extraction method of (2) , For prevention and improvement of hypertension containing high temperature water extract (RWF-2) or low molecular weight containing hydrothermal extract (HWF-1) or hot water extract (HWF-2) containing a large amount of low molecular weight substances below molecular weight 20000 Daltons Health functional food. delete

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