KR102660778B1 - Method for extracting fucoidan for glycemic control from seaweed - Google Patents

Abstract

The present invention relates to a method of extracting fucoidan, which is capable of controlling blood sugar levels, from seaweed. The present invention relates to a method of extracting fucoidan from seaweed, optimizing extraction conditions through an extraction method that can increase the content of fucoidan extracted from seaweed, and reducing the extraction cost and time of fucoidan in large quantities. The aim is to provide a method of extracting fucoidan from seaweed, which can be extracted and at the same time can control blood sugar levels by controlling the activity of the initial starch-degrading enzyme.

Description

Method for extracting fucoidan, which can control blood sugar levels, from seaweed {METHOD FOR EXTRACTING FUCOIDAN FOR GLYCEMIC CONTROL FROM SEAWEED}

The present invention relates to a method of extracting fucoidan, which is capable of controlling blood sugar levels, from seaweed. More specifically, through a fucoidan extraction method that can increase the content of fucoidan extracted from seaweed, extraction conditions are optimized, the extraction cost and time of fucoidan are reduced, enabling mass extraction, while controlling the activity of the initial starch degrading enzyme. The purpose is to provide a method of extracting fucoidan, which can control blood sugar levels, from seaweed.

In Korea, seaweed has been used as food for a long time, but not as widely as marine animals. However, recently, seaweed has been receiving attention as one of the next-generation superfoods in the United States due to the polyphenol-based nutrients, vitamins, vegetable proteins, and minerals it contains. In addition, thanks to the recent wellness trend, research on the nutritional properties of seaweed is being actively conducted.

Approximately 80% of all species on Earth live in the ocean, and a huge amount of resources are reported. In addition, because the marine environment is very different from land, such as high salt, high pressure, and low oxygen, the intracellular components of marine organisms living in the ocean that can be produced as a result of metabolism are expected to be significantly different from those of terrestrial organisms. Therefore, research to discover functional substances from marine organisms and use them as food and food additives is being actively conducted, and marketability is rapidly increasing.

In addition, seaweed has been consumed in Korea and neighboring Asian countries for a long time, but recently, seaweed's potential health promotion effects such as heavy metal removal activity, antihypertensive activity, hyperlipidemia prevention, anticancer activity, anti-inflammatory, antioxidant activity, and antibacterial activity have been discovered. As they become known, Europe is also showing active changes such as not only using seaweed as a raw material for functional foods and medicines, but also consuming it directly as food.

Seaweed consists mostly of water, and has the highest carbohydrate content among the remaining ingredients, making up almost half of the total amount of carbohydrates. Polysaccharides, which are the main components of these seaweeds, can be divided into three types: cell wall polysaccharides, which have a fine fibrous crystal structure in the outer layer, amorphous gel-like viscous polysaccharides that cover them, and storage polysaccharides within the cells.

Seaweed is broadly classified into green algae, brown algae, and red algae, and its composition varies slightly depending on the type. The usefulness of these polysaccharides is as follows: Pharmacological effects include prevention of blood coagulation, reduction of neutral lipids in the blood (seaweed polysaccharide sulfate, fucoidan, carrageenan, saragasic acid), anti-cholesterol effect (seaweed acid polysaccharide), anti-cancer effect (sulfate polysaccharide of sword algae, chives, fucoidan), and nutritional effects. (soluble dietary fiber), energy resources (biomass), and seaweed biotechnology (protoplast production using hezopolysaccharide degrading enzyme).

Among them, brown algae are composed of various minerals in addition to water and polysaccharides, and the polysaccharide components include straight-chain glucans of cell wall polysaccharides, viscous polysaccharides (polysaccharides unique to seaweeds, alginic acid, fucoidan, sargassan), and storage polysaccharides (laminarin). In particular, alginic acid is used as Na-alginate food additive, fiber (Iljinsan rayon), ice cream, cheese stabilizer, medical gauze, and diet food (Sr, Cd absorption inhibition, intestinal function, calorie-free).

Other minerals contained in brown algae include calcium, potassium, magnesium, iodine, iron, and zinc, which are known to be excellent for the growth of humans and animals and the prevention of various diseases, to the extent that they are known as the fifth nutrient. there is.

Seaweed contains various useful substances, but fucoidan and mineral components are especially noteworthy.

In particular, interest in the various functionalities expressed by fucoidan, a high molecular weight polysaccharide contained in brown algae such as seaweed and kelp, which have been consumed for a long time in East Asia such as Korea, Japan, and China, is growing.

Fucoidan is a polysaccharide containing sulfate groups with fucose present in seaweed as its main ingredient, and is generally present in large quantities in brown algae such as kelp, seaweed, and hijiki. Fucoidan is known to have excellent physiological activities such as improving blood circulation and anti-cancer, as well as having whitening, moisturizing, and wrinkle-improving effects, so it is also in the spotlight as a cosmetic raw material.

In addition, fucoidan is a polysaccharide whose main component is L-fucose and contains a large number of sulfate groups, and is contained in large quantities in brown algae such as kelp, seaweed, and hijiki. The chemical structure, physiological activity, and molecular weight of fucoidan vary depending on the type of brown algae, harvest location, and harvest time.

Fucoidan exhibits various physiological activities such as enhancing immune activity, anti-coagulant effect, antioxidant effect, anticancer effect, and antibacterial effect, and based on these effects, research is being actively conducted to apply it to the pharmaceutical, cosmetics, and food industries.

Various extraction methods have been attempted to extract various useful ingredients contained in these seaweeds, but most are extracted through physical manipulation or acid decomposition, which not only causes environmental pollution problems and threatens the health of workers, but also causes high costs in the extraction process. There is a downside to this.

KR 10-2017-0043993 B1 KR 10-2017-0044828 A

The purpose of the present invention is to provide a fucoidan extraction method that can increase the content of fucoidan extracted from seaweed, optimize extraction conditions, reduce fucoidan extraction cost and time, and enable mass extraction of fucoidan.

Another object of the present invention is to provide a method of extracting fucoidan, which is capable of controlling blood sugar levels, from seaweed by controlling the activity of the initial starch-degrading enzyme.

A method of extracting fucoidan, which is capable of controlling blood sugar levels, from seaweed according to an embodiment of the present invention includes 1) a pretreatment step of drying and desalting seaweed; 2) a mixing step of mixing the seaweed pretreated in step 1) with an aqueous calcium salt solution; 3) a first purification step of centrifuging the stirred mixture of step 2) and filtering it under reduced pressure; 4) precipitating the filtered mixture of step 3) in an alcohol solution; 5) a second purification step of centrifuging the precipitate from step 4) to obtain a supernatant; 6) Precipitating the supernatant of step 5) in an alcohol solution; 7) a third purification step of centrifuging the precipitate from step 6) to remove the supernatant; and 8) removing the supernatant of step 7) and drying the remaining precipitate to obtain Fucoidan.

The calcium salt aqueous solution is selected from the group consisting of calcium chloride (CaCl ₂ ) aqueous solution, calcium carbonate (CaCO ₃ ) aqueous solution, calcium hydroxide (Ca(OH) ₂ ), and mixtures thereof.

The seaweed is selected from the group consisting of seaweed, hijiki, kelp, kelp, seaweed, seaweed, and mixtures thereof.

The fucoidan has a molecular weight of 600 to 900 KDa.

The fucoidan exhibits a blood sugar control effect through its ability to inhibit starch-degrading enzymes, and the starch-degrading enzyme includes alpha-glucosidase.

A food composition for preventing and improving diabetes according to another embodiment of the present invention is manufactured including fucoidan extracted according to the above method.

A pharmaceutical composition for preventing and treating diabetes according to another embodiment of the present invention is prepared including fucoidan extracted according to the above method.

Hereinafter, the present invention will be described in more detail.

A method of extracting fucoidan, which is capable of controlling blood sugar levels, from seaweed according to an embodiment of the present invention includes 1) a pretreatment step of drying and desalting seaweed; 2) a mixing step of mixing the seaweed pretreated in step 1) with an aqueous calcium salt solution; 3) a first purification step of centrifuging the stirred mixture of step 2) and filtering it under reduced pressure; 4) precipitating the filtered mixture of step 3) in an alcohol solution; 5) a second purification step of centrifuging the precipitate from step 4) to obtain a supernatant; 6) Precipitating the supernatant of step 5) in an alcohol solution; 7) a third purification step of centrifuging the precipitate from step 6) to remove the supernatant; and 8) obtaining fucoidan by removing the supernatant of step 7) and drying the remaining precipitate.

Fucoidan is a polysaccharide (average molecular weight 20,000) with a sulfate group attached, and is effective in anticancer, antihypertensive, anti-inflammatory, etc., and is characterized by being present in seaweed.

Step 1) of the present invention is a pretreatment step of drying and desalting seaweed, and is a step of removing moisture and salts from seaweed to extract high purity Fucoidan contained in seaweed.

The moisture of the seaweed is removed through natural drying, which involves drying the seaweed using solar heat and natural wind. In clear weather with little cloud cover, it is preferable to dry the seaweed for about 1 to 2 days.

The salts of the seaweed are removed through desalting after the salts are eluted from naturally dried seaweed in continuously flowing fresh water. The desalting process is performed by using fresh water to sufficiently submerge the dried seaweed and continuously exchanging fresh water. This is done to sufficiently remove moisture after eluting foreign substances on the surface of the seaweed and salts within the seaweed.

The seaweed pretreated in step 1) above may contain not only the fucoidan component, but also polysaccharides such as alginic acid and laminarin, and heavy metal components such as arsenic.

Step 2) of the present invention is a mixing step of mixing the seaweed pretreated in step 1) with an aqueous calcium salt solution, and is a step of mixing with an aqueous calcium salt solution to remove alginic acid contained in the pretreated seaweed. .

The aqueous calcium solution in step 2) can coagulate the alginate in the pretreated seaweed into alginate, and the alginate coagulation action is achieved through stirring.

Step 3) of the present invention is a first purification step in which the stirred mixture of step 2) is centrifuged and filtered under reduced pressure, and the alginate aggregated due to step 2) is separated through centrifugation and purified through reduced pressure filtration. It will happen.

Alginic acid is removed from the mixture filtered through reduced pressure filtration.

The centrifugation is preferably performed at a speed of 6000 rpm to 7000 rpm for 10 to 30 minutes, and the reduced pressure filtration is performed using 20 to 25 ㎛ NO. 4 Filter paper can be used.

In addition, the method for removing alginate is preferably to collect alginate through centrifugation and reduced-pressure filtration, but the method for collecting precipitated alginate is not particularly limited.

Step 4) of the present invention is a step of precipitating the filtered mixture of step 3) in an alcohol solution, and through step 3), alginic acid precipitates laminarin present in the filtered mixture. It's a step.

The laminarin is a carbohydrate contained in the cell wall of brown algae, a storage polysaccharide equivalent to the starch of higher plants, and is mainly found in kelp.

The alcohol solution in step 4) is not a pure ethanol solution, but an ethanol solution diluted in the range of 30% to 50% dilution, and preferably an ethanol solution diluted to 40%.

Step 4) is a step of precipitation for 3 to 6 hours, and is preferably performed at 3 to 6°C.

Step 5) of the present invention is a second purification step in which the precipitate of step 4) is centrifuged to obtain a supernatant. The laminarin precipitated in step 4) is separated by centrifugation, and the supernatant is collected. It is refined through.

The supernatant contains high purity fucoidan from which laminarin has been removed.

The centrifugation is preferably performed at a speed of 6000 rpm to 7000 rpm for 10 to 30 minutes.

The first to third purification steps of the present invention are steps to remove other foreign substances from seaweed, removing polysaccharides such as alginic acid and laminarin, and heavy metal components such as arsenic, excluding fucoidan. Thus, the purity of fucoidan can be further increased.

Step 6) of the present invention is a step of precipitating the supernatant of step 5) in an alcohol solution, and is a step of precipitating high purity fucoidan contained in the supernatant of step 5).

Step 6) is a step of precipitation for 3 to 6 hours, and is preferably performed at 3 to 6°C.

The alcohol solution in step 6) is not a pure ethanol solution, but an ethanol solution diluted in the 60% to 80% dilution range, and preferably an ethanol solution diluted to 70%.

Step 7) of the present invention is a third purification step in which the precipitate of step 6) is centrifuged to remove the supernatant. Fucoidan precipitated in step 6) is separated by centrifugation and the supernatant is removed. This is to obtain high purity fucoidan.

The centrifugation is preferably performed at a speed of 6000 rpm to 7000 rpm for 10 to 30 minutes.

Step 8) of the present invention is a step of obtaining fucoidan by removing the supernatant of step 7) and drying the remaining precipitate, which removes polysaccharides such as alginic acid and laminarin, and heavy metals such as arsenic. This is the step of obtaining high purity Fucoidan from the sediment remaining after the components are removed.

The calcium salt aqueous solution is selected from the group consisting of calcium chloride (CaCl ₂ ) aqueous solution, calcium carbonate (CaCO ₃ ) aqueous solution, calcium hydroxide (Ca(OH) ₂ ), and mixtures thereof.

If the calcium salt aqueous solution is not added, alginic acid is not sufficiently removed, and there is a problem in that alginic acid remains during Fucoidan extraction.

Preferably, the calcium salt aqueous solution is an aqueous calcium chloride (CaCl ₂ ) solution.

More preferably, the calcium salt aqueous solution may further include a heavy metal adsorption composition.

The heavy metal adsorption composition includes thyme (Thymus vulgaris) extract, carob (Ceratonia siliqua (L.) Taub.) extract, and carob sedge (Carex nubigena var. albata) extract, and can exhibit antibacterial activity, making it effective against seaweed. It can remove harmful bacteria and heavy metals such as arsenic.

Herbal plants such as thyme and carob are herbaceous plants whose flowers, seeds, stems, leaves, and roots are useful to humans, and have played an important role since ancient times as medicinal herbs for pain relief, sedation, etc., as well as for preservatives and insecticides. Today, it is a natural material that is receiving more and more attention due to the toxicity of chemicals to the human body, concerns about environmental pollution, and its excellent efficacy.

The thyme (Thymus vulgaris) is an aromatic plant belonging to the Lamiaceae family and has grayish green small leaves. There are two main varieties, one of which is the southern French variety also called serpolet ( meridionale (wild thyme) and the other (Thymus vulgaris), also called winter thyme (thym dhiver), German thyme (thym allemand), or farigoule, is taller, has wider leaves, and a bitter taste. stronger. Lemon thyme (thym citron, T. citriodorus) is also widely used to give a refreshing flavor to various dishes. Thyme has a very pleasant scent and contains the essential oil thymol, which has antiseptic properties. Thyme, one of the basic cooking herbs, can be used fresh or dried and used alone or used to make a bouquet garni.

The carob {Ceratonia siliqua (L.) Taub.} is an evergreen broad-leaved shrub that grows to a height of about 10 m and is drought tolerant. The leaves contain resin. Small, dark greenish-brown flowers bloom in the fall, and long green pods hang from them. These pods turn chocolate-colored when ripe, and contain small, shiny, hard beans inside. Carob gum, which softens the skin, is popular as a skin pack and is also used to treat diarrhea.

The above-mentioned sedge (Carex nubigena var. albata) is a perennial herb of the monocot family Cyperaceae and grows in mountains and fields. The rhizome is short, and the stem is thin, long, triangular, clustered, and stands upright. The height is about 30cm. The leaves are clustered, line-shaped, and have sharp ends. The leaves are shorter or longer than the stem and are about 3mm wide. Flowers bloom from April to June. The flower spike stands straight at the end of the stem, has a cylindrical shape, and is about 25-40 mm long. There are short, rope-shaped bracts at the base, and the spikelets are bisexual and about 5 mm long. The female flower inflorescence is egg-shaped, has a pointed tip, and is dark brown. The fruit sac is egg-shaped, 4-5 mm long, longer than the inflorescence, and has two styles. It is distributed in Jeollabuk-do, Gyeonggi-do, South Pyongan Province, North Pyongan Province, South Hamgyong Province, and North Hamgyong Province.

The extract is extracted using an extraction solvent selected from the group consisting of water, C ₁ to C ₆ lower alcohols, and mixtures thereof.

Specifically, to prepare the extract, there are steps of washing the natural product; Drying after washing; Grinding the natural product after drying; leaching the pulverized material using an organic solvent; Leaching and drying the sample; Leaching using water; And including the step of leaching, a pine cluster extract can be obtained.

The natural extract extracted using the organic solvent may further include the step of fractionation using the organic solvent.

The method of preparing the extract may be a conventional extraction method in the art, such as ultrasonic extraction, leaching, and reflux extraction. Specifically, it may be an extract obtained by extracting a natural product from which foreign substances have been removed by washing and drying with water, alcohol having 1 to 6 carbon atoms, or a mixed solvent thereof, and may be an extract obtained by sequentially applying the solvents to the sample.

The reflux extraction method is based on 100 mL of water and alcohol having 1 to 6 carbon atoms, 10 to 30 g of pulverized natural product, reflux time of 1 to 3 hours, and alcohol or water having 1 to 6 carbon atoms.

More specifically, based on 100 mL of alcohol with 1 to 6 carbon atoms or 100 mL of water, 10 to 20 g of pulverized natural product, reflux time of 1 to 2 hours, and alcohol or water with 1 to 4 carbon atoms.

The leaching method is carried out at 15 to 30°C for 24 to 72 hours, and water or an alcohol with 1 to 6 carbon atoms is used as an extraction solvent. More specifically, the extraction is performed at 20 to 25°C for 30 to 54 hours, and the extraction solvent is water or an alcohol having 1 to 6 carbon atoms.

The ultrasonic extraction method proceeds at 30 to 50°C for 0.5 to 2.5 hours, and the extraction solvent is water or an alcohol with 1 to 6 carbon atoms. Specifically, extraction is performed at 40 to 50°C for 1 to 2.5 hours, and the extraction solvent is water or an alcohol with 1 to 6 carbon atoms.

The extraction solvent can be used in an amount of 2 to 50 times, more specifically, 2 to 20 times the weight of the sample. For extraction, the sample may be left for 1 to 72 hours, more specifically, 24 to 48 hours for leaching in the extraction solvent.

After extraction, the extract can be fractionated by sequentially applying a new fractionation solvent. The fractionating solvent used during fractionation is at least one selected from the group consisting of water, hexane, butanol, ethyl acetic acid, ethyl acetate, methylene chloride, and mixtures thereof, and is preferably ethyl acetate or methylene chloride.

After obtaining the extract or fraction, additional methods such as concentration or freeze-drying can be used.

Preferably, the heavy metal adsorbing composition contains 100 parts by weight of carob extract and 40 to 60 parts by weight of cypress sedge extract, based on 100 parts by weight of thyme extract.

In the case within the above range, the heavy metal adsorption effect can be increased through excellent antibacterial action, and various harmful bacteria except fucoidan can be removed, making high purity fucoidan extraction possible.

Preferably, the heavy metal adsorption composition is included in the calcium salt aqueous solution of step 2) of the present invention, and in step 2), 10 to 30 parts by weight of the heavy metal adsorption composition is mixed and stirred with respect to 100 parts by weight of the calcium salt aqueous solution.

When the heavy metal adsorption composition containing the thyme extract, carob extract, and Eucalyptus extract is further mixed and stirred, heavy metals such as residual alginic acid and arsenic that have not been completely removed by the aqueous calcium salt solution can be adsorbed and removed, and within the above range. In this case, the purity of fucoidan can be further increased by providing an excellent alginic acid removal effect and lowering the concentration of other sugars other than alginic acid.

In addition, when added in excess of the above range, more than necessary is added to adsorb and remove heavy metals such as residual alginic acid and arsenic, causing problems in extracting high purity fucoidan.

The seaweed is selected from the group consisting of seaweed, hijiki, kelp, seaweed, seaweed, seaweed, and mixtures thereof.

The seaweed is a type of brown algae and includes, but is not limited to, seaweed, hijiki, seaweed, kelp, seaweed, seaweed, etc., and may include any species that contains a fucoidan component. Preferably, it is better to use hijiki or moxa.

The seaweed ear is a sporophyll (root) that releases the seeds of seaweed. In the past, it was not distributed due to poor texture and lack of a market, but the fucoidan content of seaweed ear is 12.8g per 100g, which is 7 times that of seaweed leaf.

The hijiki (Sargassum fusiforme) is a type of seaweed and is a brown algae plant produced naturally and farmed in the clean waters of Wando and the southwest sea (the southern coast area). Tot belongs to the brown horse group. It is very rich in calcium, iodine, iron, and magnesium.

Sargassum is a general term for algae of the genus Sargassum, and is also used as a general term for a specific species of Sargassum fulvellum or large brown algae. The body has a clear division into roots, stems, and leaves. The roots are ridged and produce one central branch that grows to a height of 1-3 m or more. The stem is triangular or triangular and twisted. The leaves grow from the stem toward the base, are curved, have a spatula shape or an oval shape, and have a midrib extending to the center of the leaf. The upper leaves are lanceolate and have serrated protrusions on the edges, and air bubbles form all over the body starting from the stem. It has a dark tan color and can be found along the entire coast of Korea. It is a representative type of mogaban used for food, and is commonly sold in the market. Caprifolia is a perennial plant that is difficult to digest and is a representative species that forms marine forests along the coast of Korea.

Approximately 20 species of caps are collected, including the earthworm, the hoe-shaped cap, the egg-shaped cap, the twirl-shaped cap, the large-leaf cap, the double-leaf cap, and the double-leaf cap, and about 50 species are growing in Japan and Southeast Asia. Places with lush moss are suitable for various coastal animals to obtain food or lay eggs, and play a very important role in preserving the environment and securing fishing resources. In Korea, many species of the genus Mojapan are edible, and are used as raw materials for the seaweed industry, such as alginic acid, or as fertilizer.

The kelp (Saccharina japonica) is a marine plant belonging to brown algae and is used as a food material in Korea, China, and Japan. Kelp can be cultivated in natural environments as well as in the sea. It contains various pigments including carbohydrates, polysaccharides, carotenes, and chlorophyll, as well as alginic acid, iodine, vitamin B2, and fucoidan.

Gracilaria verrucose is a seaweed of the red algae family Gracilaria verrucose. Its body is divided into feathers and its color is black purple or dark brown. When making agar agar, it is mixed with agar agar. In Jangheung, Wando, Haenam, and Jindo, it is called kosirak, and near Busan, goby fish is also called kosiraegi.

The Pelvetia siliquosa is a seaweed from the brown algae family, growing on the southern and western coasts. Korean moxa is known to be the same species as ‘Pelvetia wrightii’ from Japan, but a Chinese seaweed scientist found that the species from the West Sea, unlike the Japanese one, has a small and upright body, so ‘P. A new name was given, ‘siliquosa’ (1953). Moxa is commonly eaten and is also used as a raw material for alginic acid.

The fucoidan has a molecular weight of 600 to 900 KDa.

The average molecular weight of fucoidan contained in seaweed is MW 20KDa, and it is a basic sugar called fucose combined with a sulfate group. There are two types: F-fucoidan, which comes from sulfated fucoidan, and U-fucoidan, which contains about 20% glucuronic acid.

In particular, fucoidan is a sulfated polysaccharide containing a high proportion of sulfate groups and is a macromolecule with a molecular weight generally exceeding 300 kDa derived from brown algae such as kelp and some marine invertebrates such as sea cucumbers and sea urchins. Fucoidan has been reported to have various biological activities such as anti-inflammatory, anticancer, immunomodulatory, anticoagulant, antithrombotic, and antioxidant. However, since our body cannot break down fucoidan into low molecules on its own, we ingest fucoidan in its high molecular form as is. Even if a large amount is consumed, most of it is excreted without being absorbed into the body.

As mentioned above, despite the numerous advantages of fucoidan, which exhibits various biological activities, the molecular weight of fucoidan extracted from brown algae is not only 20 KDa, but also contains polysaccharides including alginic acid, making it a high molecular weight substance of over 1,000 KDa, so its absorption rate is very low when ingested. has a disadvantage, so it is desirable to consume low-molecular-weight products to absorb fucoidan, so the development of a low-molecular-weight manufacturing method for fucoidan is required.

Fucoidan extracted according to the method of the present invention has a molecular weight of 600 to 900 KDa, which can increase the absorption rate in the body when ingested.

The fucoidan exhibits a blood sugar control effect through its ability to inhibit starch-degrading enzymes, and the starch-degrading enzyme includes alpha-glucosidase.

The alpha-glucosidase is a decomposition enzyme that hydrolyzes carbohydrates to produce alpha-glucose, and provides sugar for essential energy supply to organisms.

In humans, alpha-glucosidase acts in the final step of carbohydrate breakdown, so alpha-glucosidase-mediated production of blood sugar supplies the body with energy to maintain a healthy state. Low concentrations of alpha-glucosidase result in serious pathological changes such as Pompe's disease and impaired glycogen storage caused by alpha-glucosidase deficiency in lysosomes, in contrast, when the activity of the enzyme is reduced. In high cases, blood sugar levels may increase.

Therefore, Fucoidan extracted according to the present invention can exhibit a blood sugar control effect through the starch-degrading enzyme inhibitory ability of α-glucosidase.

A food composition for preventing and improving diabetes according to another embodiment of the present invention is manufactured including fucoidan extracted according to the above method.

The term “food” refers to a natural product or processed product containing one or more nutrients, preferably in a state that can be eaten directly after a certain degree of processing. In the usual sense, it means food. , which includes all food additives, functional foods, and beverages.

Foods to which the food composition can be added include, for example, various foods, beverages, gum, tea, vitamin complexes, functional foods, etc. Additionally, special nutritional foods (e.g., milk formula, infant and baby food, etc.), processed meat products, fish products, tofu, jelly, noodles (e.g., ramen, noodles, etc.), breads, health supplements, seasoned foods (e.g., soy sauce) , soybean paste, red pepper paste, mixed paste, etc.), sauces, confectionery (e.g., snacks), candy, chocolate, gum, ice cream, dairy products (e.g., fermented milk, cheese, etc.), other processed foods, kimchi, pickled foods (various types of kimchi) , pickles, etc.), beverages (e.g., fruit drinks, vegetable drinks, soy milk, fermented drinks, etc.), and natural seasonings (e.g., ramen soup, etc.), but are not limited thereto. The food, beverage or food additive can be manufactured by conventional manufacturing methods.

In addition, the above-mentioned “functional food” or “health functional food” refers to a food group or food composition that has added value to the food by using physical, biochemical, bioengineering methods, etc. to function and express the function of the food for a specific purpose. It refers to food designed and processed to fully express the body's regulatory functions related to defense rhythm control, disease prevention and recovery, etc., and specifically, it may be a health functional food. The functional food may include food auxiliary additives that are foodologically acceptable, and may further include appropriate carriers, excipients, and diluents commonly used in the production of functional foods.

The type of health supplement is not limited thereto, but may be in the form of powder, granule, tablet, capsule, or beverage.

The advantages and features of the present invention and methods for achieving them will become clear with reference to the embodiments described in detail below. Hereinafter, the present invention will be described in detail through examples. However, these examples are for illustrating the present invention in detail, and the scope of the present invention is not limited to these examples.

A pharmaceutical composition for preventing and treating diabetes according to another embodiment of the present invention is prepared including fucoidan extracted according to the above method.

Fucoidan extracted according to the present invention is formulated in the form of oral dosage forms such as powders, granules, tablets, capsules, suspensions, emulsions, syrups, aerosols, external preparations, suppositories, and sterile injection solutions according to conventional methods. It can be used interchangeably.

In addition, carriers, excipients, and diluents that may be included in the pharmaceutical composition for preventing and treating diabetes include lactose, dextrose, sucrose, sorbitol, mannitol, xylitol, erythritol, maltitol, starch, acacia gum, alginate, gelatin, Various compounds or mixtures including calcium phosphate, calcium silicate, cellulose, methyl cellulose, microcrystalline cellulose, polyvinyl pyrrolidone, water, methylhydroxybenzoate, propylhydroxybenzoate, talc, magnesium stearate and mineral oil. I can hear it. When formulated, it is prepared using diluents or excipients such as commonly used fillers, extenders, binders, wetting agents, disintegrants, and surfactants.

Solid preparations for oral administration include tablets, pills, powders, granules, capsules, etc. These solid preparations contain at least one excipient such as starch, calcium carbonate, or sucrose. ) or prepared by mixing lactose, gelatin, etc. In addition to simple excipients, lubricants such as magnesium stearate and talc are also used. Liquid preparations for oral use include suspensions, oral solutions, emulsions, syrups, etc. In addition to the commonly used simple diluents such as water and liquid paraffin, various excipients such as wetting agents, sweeteners, fragrances, and preservatives may be included. . Preparations for parenteral administration include sterile aqueous solutions, non-aqueous solutions, suspensions, emulsions, lyophilized preparations, and suppositories. Non-aqueous solvents and suspensions may include propylene glycol, polyethylene glycol, vegetable oil such as olive oil, and injectable ester such as ethyl oleate. As a base for suppositories, witepsol, macrogol, tween 61, cacao, laurin, glycerogeratin, etc. can be used.

The preferred dosage of the composition of the present invention varies depending on the patient's condition and weight, degree of disease, drug form, administration route and period, but can be appropriately selected by a person skilled in the art. However, for desirable effects, the composition of the present invention is preferably administered at 0.0001 to 100 mg/kg per day, preferably at 0.001 to 100 mg/kg. Administration may be administered once a day, or may be administered several times. The above dosage does not limit the scope of the present invention in any way.

According to the method of extracting fucoidan capable of controlling blood sugar levels from seaweed of the present invention, the extraction conditions are optimized through an extraction method that can increase the content of fucoidan extracted from seaweed, and the extraction cost and time of fucoidan are reduced, enabling mass extraction. At the same time, it is possible to provide a method of extracting fucoidan, which can control blood sugar levels, from seaweed by controlling the activity of the initial starch-degrading enzyme.

Figure 1 shows a step-by-step method of extracting fucoidan, which is capable of controlling blood sugar levels, from seaweed according to an embodiment of the present invention.
Figure 2 shows the results of analyzing the inhibition rate of alpha-glucosidase enzyme activity according to fucoidan treatment according to an embodiment of the present invention.
Figure 3 shows the yield of the extract and the cost of consumable items required to produce 100 g of the extract according to an embodiment of the present invention.
Figure 4 shows the molecular weight distribution of an extract according to an embodiment of the present invention.
Figure 5 shows the alpha-glucosidase inhibitory ability of an extract according to an embodiment of the present invention.

Hereinafter, embodiments of the present invention will be described in detail so that those skilled in the art can easily implement it. However, the present invention may be implemented in many different forms and is not limited to the embodiments described herein.

[Preparation Example 1: Preparation of heavy metal adsorption composition]

1. Preparation of herbal extract

The thyme was thoroughly washed under running water and then completely air-dried. The dried thyme was ground in a blender and then made into powder. The powder sample was completely immersed in 50% diluted ethanol as an extraction solvent at a ratio of 1:10 (w;v), and then extracted three times for 3 hours while refluxing at 80°C. The extract was Whatman No. 2 Filtered with filter paper. The filtrate was concentrated under reduced pressure at 60°C to prepare thyme extract (TH).

Carob extract (KH) was prepared using the same method as the method for producing thyme extract (TH).

2. Preparation of Doraengi sedge extract

Dorangi sedge was washed, dried, and then ground. The pulverized product was mixed with ethanol diluted to 60%, extracted for 2 hours, cooled, and filtered with Whatman filter paper, and the filtrate was obtained to prepare a sedge extract (CE).

3. Preparation of heavy metal adsorption composition

The thyme extract (TH), carob extract (TK), and cypress sedge extract (CE) were mixed as shown in Table 1 below to prepare the heavy metal adsorption composition of the present invention.

BSC1 BSC2 BSC3 BSC4 BSC5 BSC6 T.E. 100 100 100 100 100 100 KE 100 100 100 100 100 100 C.E. - 30 40 50 60 70

(Unit: parts by weight)

[Preparation Example 2: Extraction of fucoidan]

In order to confirm the effect of the method of extracting fucoidan, which can control blood sugar levels, from seaweed according to the present invention, each step was configured as shown in Table 2 below to extract fucoidan from seaweed.

The following FUCO4 to FUCO9 are fucoidan from seaweed by adding 20 parts by weight of the heavy metal adsorption composition BSC1 to BSC6 prepared in Preparation Example 1 to 100 parts by weight of the calcium salt aqueous solution (CaCl ₂ ) in step S2 of the present invention. is extracted.

Fuco1 Fuco2 Fuco3 Fuco4 Fuco5 Fuco6 Fuco7 Fuco8 Fuco9 S1 O O O O O O O O O S2 O O O O(+BSC1) O(+BSC2) O(+BSC3) O(+BSC4) O(+BSC5) O(+BSC6) S3 O O O O O O O O O S4 - O O O O O O O O S5 - O O O O O O O O S6 - - O O O O O O O S7 - - O O O O O O O S8 O O O O O O O O O

S1: Pretreatment step of drying and desalting hijiki

S2: Mixing step of mixing the pretreated seaweed with CaCl ₂ at 85°C for 4 hours

S3: A first purification step in which the stirred mixture is centrifuged at 6,500 rpm for 20 minutes and filtered under reduced pressure on NO 4 filter paper.

S4: Precipitating the filtered mixture in 40% diluted ethanol solution for 4 hours

S5: a second purification step of centrifuging the precipitate at 6,500 rpm for 20 minutes to obtain a supernatant;

S6: Precipitating the supernatant in an ethanol solution diluted to 70% for 4 hours

S7: Third purification step of removing the supernatant by centrifuging the precipitate at 6,500 rpm for 20 minutes

S8: Obtaining fucoidan by drying the final product with hot air for 24 hours

[Experimental Example 1: Analysis of the content of fucoidan extracted according to the present invention]

The following experiment was conducted to analyze the content of fucoidan (FUCO1 to FUCO9) obtained in Preparation Example 2.

50 μL of fucoidan FUCO1 to FUCO9 obtained in Preparation Example 2 diluted to 0.3 mg/mL and 200 μL of sulfuric acid (18 M) were added to each 96 well plate, stirred well, and heated at 80°C for 30 minutes. After cooling, 8 μL of 3% (w/v) L-cysteine hydrochloride was added to the plate and stored at 4°C for 1 hour.

The content of fucoidan was determined using HPAEC (High Performance Anion Electric Chtomatography), and the detector was analyzed using PAD (Pulsed Amperometry Detector).

The results are shown in Table 3 below.

Fuco1 Fuco2 Fuco3 Fuco4 Fuco5 Fuco6 Fuco7 Fuco8 Fuco9 Fucoidan content 52% 62% 80% 83% 85% 93% 95% 96% 88%

(unit: %)

As shown in Table 3, compared to fucoidan FUCO1 extracted including only the first purification step of the present invention, fucoidan FUCO2 extracted further including a second purification step and further comprising a second purification step and a third purification step Fucoidan FUCO3 showed excellent fucoidan content.

In particular, when extracting fucoidan including the heavy metal adsorption composition (BSC1 to BSC6) of the present invention in step S2, polysaccharides such as alginic acid and laminarin, and heavy metal components such as arsenic are removed, further It showed excellent fucoidan content.

[Experimental Example 2: Blood sugar control effect of fucoidan extracted according to the present invention]

In order to confirm the blood sugar control effect of the fucoidan (FUCO1 to FUCO9) obtained in Preparation Example 2, the inhibitory activity of alpha-glucosidase according to the fucoidan FUCO1 to FUCO9 treatment was confirmed.

The inhibitory activity of the fucoidans FUCO1 to FUCO9 against alpha-glucosidase was measured using ρ-nitrophenyl-alpha-D-glucopyranoside as a substrate.

10 μL of the fucoidan (FUCO1 to FUCO9) was added to 40 μL of 0.1 M phosphate buffer solution (pH 6.8), and then 25 μL of the diluted enzyme solution was added. After adding 25 μL of ρ-nitrophenyl-α-Dglucopyranoside, react at 37°C for 30 minutes and add 100 μL of 0.2 M sodium carbonate solution. Upon termination, the amount of free ρ-nitrophenol produced was measured at 405 nm.

The enzyme activity inhibition rate was measured from the difference between the value obtained by adding the same amount of phosphate buffer solution instead of the fucoidan (FUCO1 to FUCO9) of the present invention to the reaction solution, and the enzyme activity inhibition rate of alpha-glucosidase according to the fucoidan FUCO1 to FUCO9 treatment. is shown in Figure 2.

As shown in Figure 2 below, it was found that fucoidan FUCO1 to FUCO9 of the present invention inhibit the enzymatic activity of alpha-glucosidase enzyme in a concentration-dependent manner, and fucoidan FUCO1 extracted including only the first purification step of the present invention In comparison, fucoidan FUCO2 extracted by further comprising a second purification step and fucoidan FUCO3 further comprising a second purification step and a third purification step showed excellent alpha-glucosidase enzyme inhibition ability.

In addition, when extracting fucoidan including the heavy metal adsorption composition (BSC1 to BSC6) of the present invention in step S2 of the present invention, polysaccharides such as alginic acid and laminarin, and heavy metal components such as arsenic are removed. It was confirmed that as the fucoidan content became more excellent, the alpha-glucosidase enzyme inhibitory ability also increased.

[Experimental Example 3: Comparison of fucoidan extraction processes from seaweed]

1. Economic comparison

The yield of the extract according to the extraction method, hot water extraction method, acid treatment extraction method, and cold water extraction method of the present invention and the price of reagents, which are consumable items required to produce 100 g of extract (calculated as $1 = 1,000 won) were evaluated.

The seaweed was tested on wakame seaweed, hijiki, kelp, seaweed, and moxa, and extraction was performed three times for each extraction process. The values were expressed as mean ± standard deviation, and the results are shown in Figure 3 below. It was.

As shown in Figure 3 below, the yield of the extract showed significant differences depending on the process, and the yield was confirmed to be excellent in the following order: the extraction method of the present invention, the hot water extraction method, the acid treatment extraction method, and the cold water extraction method (Figure 3a).

In addition, as a result of calculating the price of consumable items (reagents, etc.) required for each process to obtain 100g of seaweed extract, the development method required $272, which was about 70% of the production cost of a commercial product. In particular, when compared to the cold water extraction method, it took 1/7 the cost, confirming economic feasibility (Figure 3b).

2. Comparison of starch degrading enzyme inhibition ability

In order to confirm the blood sugar control functionality of the seaweed extract extracted according to each of the above extraction methods, the inhibitory ability of alpha-amylase, which is involved in starch decomposition, and the α-glucosidase, which is involved in glucose production, were tested. The activity was evaluated and shown in Figure 5 below.

As shown in Figure 5a below, alpha-amylase (α-amylase) inhibitory ability was not observed, but as shown in Figure 5b, the alpha-glucosidase (α-glucosidase) inhibitory ability of the extracts depending on the extraction method was It was confirmed that there was a significant difference.

In addition, the IC- ₅₀ value of α-glucosidase of the extracts by manufacturing process was 0.031 mg/mL for the extraction method of the present invention, 0.080 mg/mL for the cold water extraction method, 0.125 mg/mL for the hot water extraction method, and 0.162 mg/mL for the commercially available product. Acarbose, the main ingredient of anti-diabetic drugs, was measured at 0.146 mg/mL, and it was confirmed that the seaweed extract extracted using the extraction method of the present invention had the best α-glucosidase inhibitory ability.

3. Comparison of physicochemical properties

In order to compare and evaluate the physicochemical properties of each of the above extraction methods, carbohydrates, sugars, uronic acid, and sulfate (protein) according to the extraction method, hot water extraction method, acid treatment extraction method, and cold water extraction method of the present invention. sulfate) content and molecular weight were confirmed.

The results are shown in Table 4 below.

Chemical composition (%) Mw (kDa) Carbohydrate Protein Uronic acid Sulfate thermal water extraction 38.9±0.4 5.3±0.0 2.9±0.2 48.0±0.5 2736.7±78.3 acid treatment extraction 17.3±0.4 6.7±0.3 2.3±0.2 53.2±0.6 77.9±11.1 cold water extraction 36.0±0.8 2.3±0.1 8.7±0.8 34.4±0.5 352.1±38.8 Extraction according to the invention 24.8±0.7 2.3±0.0 28.3±0.8 18.8±0.5 887.3±6.6 For commercial use 35.2±0.2 3.1±0.0 12.7±0.3 37.3±0.2 2292.7±23.8

As shown in Table 4, the seaweed extract extracted according to the present invention had a molecular weight of 887 KDa and a sulfur content of 18.8%, and in the case of other extraction processes, the sulfur content was more than 34%, which was higher than the extraction method of the present invention. However, as confirmed above, it was confirmed that the alpha-glucosidase inhibitory ability was not excellent.

In addition, in the case of the acid extraction method, although the molecular weight was the lowest at 77.9 KDa and the sulfur content was the highest at 53.2%, the alpha-glucosidase inhibition ability was not excellent, so the molecular weight of the extracted fucoidan was It can be seen that differences appear depending on the extraction method, and it was confirmed that low molecular weight and high sulfur content are not the main factors for starch decomposition enzyme inhibition ability.

4. Comparison of molecular weight distribution

In order to compare the molecular weight distribution of each of the above extraction methods, the molecular weight distribution of the seaweed extract according to the extraction method of the present invention, hot water extraction method, acid treatment extraction method, and cold water extraction method was confirmed.

The molecular weight distribution was examined using RI profile, and the results are shown in Figure 4 below.

As shown in Figure 4 below, it was confirmed that the extraction method of the present invention, the hot water extraction method, and the commercially available extract had a constant molecular weight distribution and were composed of similar molecular weights.

5. Analysis of monosaccharides in the extract

The results of analyzing the monosaccharides of the seaweed extract extracted by each of the above extraction methods are shown in Tables 5 and 6 below.

Monosaccharide content (%) Extraction method hot water extraction Acid treatment extraction method cold water extraction method Extraction method of the present invention For commercial use rhamnose
(Ramnose) - 3.6±0.1 - 2.0±0.1 - Fucose 35.2±0.4 16.0±0.2 42.0±0.3 23.3±0.3 42.7±0.6 Xylose 0.8±0.1 1.1±0.1 - 1.0±0 - Mannose 4.7±0.2 4.7±0.2 7.0±0.2 20.2±0.4 5.0±0.2 Glucose 2.8±0.1 16.4±0.2 4.2±0.1 2.4±0.3 3.2±0.1 Galactose 56.5±0.7 58.2±0.7 46.7±0.6 51.1±0.8 49.1±0.5

Monosaccharide content (mg/g) Extraction method hot water extraction Acid treatment extraction method cold water extraction method Extraction method of the present invention For commercial use rhamnose
(Ramnose) - 8.7±0.5 - 5.3±0.3 - Fucose 116.6±3.0 39.3±1.9 181.3±8.9 59.3±2.9 187.3±4.9 Xylose 3.3±0.1 3.6±0 - 3.1±0 - Mannose 15.8±0.9 12.0±0.4 30.6±1.0 51.7±1.8 22.0±1.3 Glucose 7.0±0.2 30.1±1.7 13.5±0.8 4.5±0.3 10.4±0.4 Galactose 171.9±1.3 131.0±4.1 184.8±5.8 119.3±3.8 197.8±1.6 total 314.5±3.5 224.8±6.8 410.2±14.5 243.2±4.8 417.5±5.6

As shown in Tables 5 and 6, the seaweed extract according to the extraction method of the present invention was confirmed to have a relatively high mannose ratio (20.2%) compared to extracts from other methods.

The mannose is a type of sugar, but only a very small amount is used in metabolism, and since most of it is excreted outside the body, it does not affect blood sugar levels, making it a safe ingredient for diabetic patients.

[Experimental Example 4: Palatability evaluation]

A palatability evaluation was conducted on fucoidan FUCO1 to FUCO9 extracted according to the present invention. After manufacturing tea containing fucoidan FUCO1 to FUCO9, it was provided to 20 adult men and women, and evaluation of taste and aroma was requested.

Evaluation scores were requested from 1 to 10 points for each item, and the evaluation results were converted into average scores. The higher the number, the higher the preference for the index.

FUCO1 FUCO2 FUCO3 FUCO4 FUCO5 FUCO6 FUCO7 FUCO8 FUCO9 taste One 2 3 5 5 7 7 7 7 incense 2 2 4 5 6 6 7 8 5 Comprehensive preference (average) 1.5 2.0 3.5 5.0 5.5 6.5 7.0 7.5 6.0

(Unit: index)

Referring to Table 7, compared to fucoidan FUCO1 extracted including only the first purification step of the present invention, fucoidan FUCO2 extracted further including a second purification step and further comprising a second purification step and a third purification step It was confirmed that fucoidan FUCO3 showed excellent preference.

In addition, it was confirmed that when extracting fucoidan including the heavy metal adsorption composition (BSC1 to BSC6) of the present invention in step S2 of the present invention, the preference level lowered due to the natural extract was increased.

Although the preferred embodiments of the present invention have been described in detail above, the scope of the present invention is not limited thereto, and various modifications and improvements made by those skilled in the art using the basic concept of the present invention defined in the following claims are also possible. falls within the scope of rights.

Claims (7)

1) Pretreatment step of drying and desalting seaweed;
2) a mixing step of mixing the seaweed pretreated in step 1) with an aqueous calcium salt solution;
3) a first purification step of centrifuging the stirred mixture of step 2) and filtering it under reduced pressure;
4) mixing the filtered mixture of step 3) with an ethanol solution diluted to 40% to precipitate laminarin present in the mixture;
5) a second purification step of centrifuging the precipitate from step 4) to obtain a supernatant;
6) Precipitating the supernatant of step 5) in an alcohol solution;
7) a third purification step of centrifuging the precipitate from step 6) to remove the supernatant; and
8) A method of extracting fucoidan capable of controlling blood sugar from seaweed, comprising the step of obtaining fucoidan by removing the supernatant of step 7) and drying the remaining sediment,
The calcium salt aqueous solution further includes a heavy metal adsorption composition,
The heavy metal adsorption composition includes thyme (Thymus vulgaris) extract, carob (Ceratonia siliqua (L.) Taub.) extract, and sedge sedge (Carex nubigena var. albata) extract.
A method of extracting fucoidan, which can control blood sugar levels, from seaweed. According to clause 1,
The calcium salt aqueous solution is selected from the group consisting of calcium chloride (CaCl ₂ ) aqueous solution, calcium carbonate (CaCO ₃ ) aqueous solution, calcium hydroxide (Ca(OH) _-2 ), and mixtures thereof.
A method of extracting fucoidan, which can control blood sugar levels, from seaweed. According to clause 1,
The seaweed is selected from the group consisting of seaweed, hijiki, moba, kelp, seaweed, seaweed, and mixtures thereof.
A method of extracting fucoidan, which can control blood sugar levels, from seaweed. According to clause 1,
The fucoidan has a molecular weight of 600 to 900 KDa.
A method of extracting fucoidan, which can control blood sugar levels, from seaweed. According to clause 4,
The fucoidan exhibits a blood sugar control effect through starch-degrading enzyme inhibition,
The starch-decomposing enzyme includes alpha-glucosidase (α-glucosidase).
A method of extracting fucoidan, which can control blood sugar levels, from seaweed. Containing fucoidan extracted according to any one of claims 1 to 5
Food composition for preventing and improving diabetes. Containing fucoidan extracted according to any one of claims 1 to 5
Pharmaceutical composition for preventing and treating diabetes.