KR100458301B1 - Granular and Liquid Type of Tea Containing Extracts of Gastrodia elata Blum - Google Patents

Abstract

The present invention relates to a granular tea and a liquid tea containing the cheonma extract as an active ingredient in more detail to experimentally confirm the various functionalities of cheonma and to granule tea or liquid tea to make it easy for anyone to enjoy the cheonma with such functionality It is about. The granular tea or liquid tea preferably contains 10 to 60% by weight of cheonma extract or may be added in the form of a concentrate. According to the granular tea or liquid tea containing the cheonma extract of the present invention, the results of the quality characteristics test showed excellent functionality and sensory quality, which can be found to be very suitable as a processed food.

Description

Granular and Liquid Type of Tea Containing Extracts of Gastrodia elata Blum}

The present invention relates to a granular tea and a liquid tea containing the cheonma extract as an active ingredient in more detail to experimentally confirm the various functionalities of cheonma and to granule tea or liquid tea to make it easy for anyone to enjoy the cheonma with such functionality It is about.

Cheonma ( 天 astro , Gastrodia elata Blum) is an orchid that grows together with mulberry mushrooms. It is a perennial plant that grows in the forests of humus-rich valleys. It is 60-100m high and has no leaves and potato-like tubers. The tuber of the celestial horse is long oval and the length is usually 10-18 cm. In oriental medicine and private medicine, it is known to be effective for symptoms such as stress, fatigue, as well as serious adult diseases such as hypertension, headache, paralysis, neurological diseases and diabetes.

However, research on the functionality of Chunma has not been done enough, and there are no studies on how to use it as a processed food.

The present inventors attempted to analyze the general components and minerals of cheonma and study physiological activities such as anticancer, antimutation, liver detoxification, hypoglycemia, hypertension control, antioxidant activity, and processed into actual drinkable form based on these results. It is an object of the present invention to provide a teatable granular tea and a liquid tea.

The present invention includes granular tea and liquid tea containing cheonma extract as an active ingredient.

Chunma extract is sufficient to extract using a known extraction method and does not require a special limitation. Therefore, known hot water extraction or extraction using an organic solvent, for example, alcohols and the like may all correspond to this.

Chunma extract is preferably added 10 to 60% by weight, more preferably approximately 10% by weight. If it is added less than 10% by weight is not sufficient to exhibit the functionality of the cheonma, and if it exceeds 60% by weight is not preferable because the taste unique to cheonma. However, since the invention is not impossible even if somewhat out of the above range, the present invention includes equivalent areas within these ranges.

In a preferred embodiment of the present invention, the cheonma extract is provided in the form of a concentrate (about 40 Brix). This concentrate was obtained by injecting water approximately 10 times with respect to the weight of the horse, and repeatedly extracting it several times for 5 to 12 hours at about 70 ° C. to 90 ° C., and compressing the extracted horse horse to concentrate the extract in the state of removing the residue to about 40 Brix. Can be. Although the present invention is carried out with a concentration of about 40 Brix, this is merely a preferred embodiment of the present invention, but does not mean that it is impossible to implement the concentrate at a level slightly beyond this.

When the tea is prepared in the form of a concentrate of about 40 Brix in the preparation of tea as described above, the addition amount is preferably 10 to 60% by weight. More preferably, in the case of granular tea, the addition amount of the concentrate is 10 to 30% by weight, and in the case of liquid tea, 30 to 60% by weight is excellent in terms of sensuality.

As a preferred embodiment of the present invention, the granular tea containing the extract of cheon-ma is used as a subsidiary material in addition to the cheon-ma extract, and various extracts and concentrates such as refined glucose, anhydro-glucose, etc. Celestial concentrates and the like. The addition amount of the submaterial is a matter that can be appropriately selected by those skilled in the art and does not require any particular limitation. For example, 20-40 weight% of refined glucose, 40-60 weight% of anhydrous glucose, and 10-30 weight% of well-known additives, such as citric acid and an inorganic salt, can be added.

In addition, as a preferred embodiment in the second aspect of the present invention, the liquid tea containing the extract of cheon-ma is a subsidiary material in addition to the various saccharides, for example, liquid fructose, honey, etc. And the like. The addition amount of the submaterial is a matter that can be appropriately selected by those skilled in the art and does not require any particular limitation. For example, 30 to 60 weight% of liquid fructose, 5 to 10 weight% of honey, 2 to 10 weight% of jujube extract, and 33 weight% of well-known additives, such as citric acid and an inorganic salt, can be added.

Hereinafter, the content of the present invention will be described in more detail with reference to Examples. However, the following examples are merely examples for understanding the contents of the present invention, but the scope of the present invention is not limited thereto.

Example 1 Preparation of Chunma Extract and Concentrate

100 g of the dry horse was chopped and ground to a size of 0.5 cm or less. Water was injected 10 times with respect to the pulverized cheonma weight, and extracted three times for 9 hours at 80 ℃. The extracted cheonma was compressed to obtain an extract, and the residue was removed. The extract was concentrated to 40Brix, and the results of investigating the properties of the concentrate were as shown in Table 1 (extract) and Table 2 (concentrate).

In addition, the results of observing the change in yield with temperature and time during the extraction of cheonma are shown in Table 3 below.

Table 1 General Component Analysis and Mineral Content (Unit:%)

division moisture View minutes Crude protein Crude fiber Crude fat mineral Ca K Mg P Thousand horses 8.11 2.61 5.41 3.34 3.56 0.09 1.15 0.02 0.30

General protein content of cheonma extract was high as crude protein (41.41%), and mineral content of potassium was high.

<Table 2> Analysis of general components and quality characteristics of concentrate (unit:%)

division General ingredient Quality characteristic moisture View minutes Crude protein Brix pH Thousand horses 61.53 2.53 2,1 40 4.57

<Table 3> Yield according to extraction temperature and time (unit:%)

division 60 ℃ 80 ℃ 100 ℃ 3 hours 6 hours 9 hours 3 hours 6 hours 9 hours 3 hours 6 hours 9 hours Yield 14.60 (2.10) 15.30 (3.27) 14.22 (2.50) 17.90 (2.53) 16.01 (2.67) 17.09 (2.60) 17.26 (3.03) 19.42 (3.80) 17.30 (3.40)

The extraction yield according to the extraction temperature and time of Chunma was highest at 100 ℃ for 6 hours and the yield for solid content was extracted at 100 ℃ for 6 hours and 9 hours.

Example 2 Test of Functionality Using Chunma Extract (Antitumor Activity)

Experiment process

(1) Cell line and medium composition

The cell lines used in this experiment were Hep3B (hepatocellular carcinoma, human), human lung cancer cells, A549 (Lung carcinoma, human), human lung cancer cells, and MCF7 (brest adenocarcinoma, pleural effusion, human) cells. As a normal cell to determine the cytotoxicity of the sample itself, human normal liver cells WRL68 (Human embryo liver) was used. Among the cells used in the experiment, Hep3B, MCF7, and WRL68 were cultured by adapting DMEM medium to 10% heating-inactivated FBS in RPMI 1640 medium for A549.

(2) SRB Essay

The SRB (sulforhodamine B) assay is a method of measuring cell proliferation or toxicity by staining cellular proteins.The cells to be tested are WRL68, Hep3B, MCF7 (10% FBS, DMEM medium) and A549 (10% FBS, RPMI 1640 medium). 100 μl of each well of a 96 well plate was added at a concentration of 4-5 × 10 ⁴ cells / ml, followed by incubation for 24 hours (37 ° C., 5% CO ₂ ). 100 μl each was added at 0.6, 0.8, 1.0 mg / ml and incubated for 48 hours. After the incubation was completed, the supernatant was removed and 100 µl of cold 10% (w / v) TCA (trichloroacetic acid) was added thereto, and left at 4 ° C for 1 hour. After drying, 100 μl of 0.4% (w / v) SRB solution dissolved in 1% (v / v) acetic acid was added to each well and stained at room temperature for 30 minutes. The unbound SRB dye solution was washed 4-5 times with 1% acetic acid and dried, and then 100 μl of 10 mM Tris buffer was dissolved to dissolve the dye solution. The absorbance was measured at 540 nm using a microplate reader. Tables 4, 5, 6, and 7 are shown below.

Experiment result

Table 4 Liver Cancer Cells (% / mg / ml)

sample Solvent Hep3B (liver cancer) 0.2 0.6 1.0 Thousand horses Distilled water 36 57 76 Effect criteria 50% less than Black method SRB assay

Table 5 Breast Cancer Cells

sample Solvent MCF-7 (Human Breast Cancer Cell) 0.2 0.6 1.0 Thousand horses Distilled water 35 51 79 Effect criteria 50% less than Black method SRB assay

Table 6 Lung Cancer Cells

sample Solvent A549 (Human Lung Cancer Cell) 0.2 0.6 1.0 Thousand horses Distilled water 25 43 72 Effect criteria 50% less than Test method SRB assay

Table 7 Gastric Cancer Cells

sample Solvent AGS (Human Gastric Cancer Cell) 0.2 0.6 1.0 Thousand horses Distilled water 35 57 83 Effect criteria 50% less than Test method SRB assay

As a result of measuring the cancer cell growth inhibition ability of cheonma, higher than 76, 79, 72% at a concentration of 1.0% by weight in the inhibitory activity against human liver cancer cells (Hep3B), breast cancer cells (MCF-7) and lung cancer cells (A549) Activity was shown. In particular, the inhibitory activity of gastric cancer cells (AGS) was 83%, which was higher than that of other cancer cells.

Example 3 Test of Functionality Using Chunma Extract (Antimutagenicity)

Experiment process

(1) Rec-essays

a) strain and pretreatment

The strains used in this experiment were Bacillus sertilis PB 1652 rec + and PB 1791 rec-, and the lyophilized strains were inoculated in Petri dishes solidified with nutrient medium using a scalpel in a cleanbench. It was. When the incubation was completed, platinum was added to the sterilized TSB liquid medium and incubated at 37 ° C. for 24 hours to use in the experiment.

b) spore agar plate preparation and experimental method

After incubation, 10 ml of soft broth agar containing 0.1 ml spore solution was added to the Petri dish and solidified. Then, the paper disk (p.disc) was placed, and 20 µl of each extract and 10 µl of control MNNG were inoculated. After incubation at 12 to 24 hours, the inhibition zone was measured.

c) mutagenicity measurement

Under a sterile cleanbench, MNNG (2 μg / μl), a strong mutagen among several mutagenic substances, was added to the sterilized paper disc in the amount of 20 μg / paper disc as mutagenic control and each extract was added to Mutagenicity was measured by adding 100 μg / paper disc and measuring the difference between the inhibition region (cm) formed by the rec (+) and rec (−) bacteria.

d) measurement of antimutagenicity

After confirming mutagenicity, MNNG (10µg / p.disc) and extracts (50, 100µg / p.disc) were mixed at a ratio of 1: 1 (10µl: 10µl) and cultured in the same manner as in mutagenicity measurement. Comparison of the inhibition zones (cm) and difference ratio (cm) between rec (+) and rec (-) bacteria to determine how much each extract inhibits the mutagenicity of MNNG, a mutagen, ratio) (sample + MNNG / MNNG). The control was treated only with MNNG. The measurement results are shown in Table 8 below.

Experiment result

Table 8 Antimutagenicity

sample Solvent amounttest (µg / disc) Inhibition halodiameter (cm) differnce zone (cm) differnce ratio (sample / MNNG) rec ⁺ rec ^- Thousand horses Distilled water 50 1.2 2.4 1.2 0.71 100 1.0 2.2 1.2 0.71 Stability and Effectiveness Criteria 50% or more Test method Rec assay

As a result of performing mutagenesis experiment of the cheonma extract, it was determined that the mutant extract itself did not have mutagenicity by confirming that no clear zone was generated. In addition, the mutant inhibitory effect according to the extract concentration was tested by mixing 1: 1 extract of cheonma extract (50, 100㎍ / p.disc) and MNNG (10㎍ / p.disc).

Compared with the control group MNNG (20㎍ / p.disc), the mixture of MNNG and extract 1: 1 showed a relatively high antimutagenicity. The antimutagenicity of extracts was determined based on MNNG, a potent mutagenic substance used as a control, and as a result, Chunma extract showed 29% mutation inhibition rate at 50, 100㎍ / p.disc concentration against MNNG. It can be inferred to have antimutagenicity by inhibiting the binding of the strain to DNA or RNA to some extent.

Example 4 Test of Functionality Using Gastrodiae cava Extracts

Experiment process

Experiments were performed using α-glucosidase, which plays a critical role in raising blood glucose levels in vivo. First, the enzyme was prepared by dissolving the enzyme in 10 mM PIPES buffer, and then mixed 20 mM maltose and each extract with 10 µl, 40 µl and 10 µl, respectively, and the final volume was mixed with 60 µl. Incubate for minutes. 1 ml DNS reagent was added to 60 µl of the reaction solution, and the reaction was stopped by boiling water treatment at 100 ° C. (10 minutes), and then absorbance was measured at 540 nm to quantify the reducing sugar produced by the enzymatic reaction. Comparing with the enzyme activity inhibition rate was calculated and the results are shown in Table 9 below.

Experiment result

TABLE 9 Blood sugar drop (% / mg / ml)

sample Solvent Concentration 0.2 0.6 1.0 Thousand horses Distilled water 42 58 64 Effect criteria 50% or more Test method α-glucosidase

α-glucosidase not only plays a crucial role in raising blood glucose in vivo, but is also involved in various metabolic disorders, diabetes, viral adhesion, bacterial infection, and carcinogenesis. In this experiment, the enzyme activity was inhibited by adding the cheonma extract to the substrate (maltose). As a result, the high enzyme activity inhibition rate was 58% at the concentration of 0.6% by weight and 64% at the concentration of 1.0% by weight. Indicated.

Example 5 Test of Functionality Using Chunma Extract

Experiment process

Since ACE is an enzyme that induces hypertension, the ACE reagent was used to measure the inhibitory activity of this enzyme. Before the experiment, all the reactants were maintained at 37 ° C., ACE reagent 1 viral was dissolved in 10 ml of 37 ° C. distilled water, and 1 ml each was taken into an Eppendorf tube. 100 μl of extract and ACE calibrator for each concentration were added to each Eppendorf tube, and then reacted at 37 ° C. for 5 minutes to measure absorbance at 340 nm. The absorbance measured after 5 minutes was defined as final A. It was. As a control, 100 μl of distilled water was added instead of the extract. The ACE (U / L) value of the control was measured by the change in absorbance when nothing was added, and the activity of ACE was calculated according to the following formula.

ACE (U / L) = (Initial A-Final A) _Test / (Initial A-Final A) _Control × active of ACE reagent (50U / L)

Experiment result

<Table 10> Blood Pressure Drop (% / mg / ml)

sample Solvent Concentration 0.2 0.6 1.0 Thousand horses Distilled water 63 82 89 Effect criteria 50% or more Test method ACE system

The main mechanism of blood pressure rise in vivo is the renin angiotensin system, which plays an important role in ACE. Angiotensin II inhibits the excretion of water and sodium by promoting the secretion of aldosterin in the body, and inactivates bradykinin, which has a vasorelaxant effect, thereby raising blood pressure.

Therefore, direct inhibition of ACE could suggest the possibility of treatment of hypertension.

In this experiment, ACE uses the principle of decomposing FAPGG (furylacryloylphenylalanyl glycylglycine) into FAP (furylacryloylphenylalanine) and GG (glycilyglycine). It showed the highest inhibitory activity of 89%, especially at a concentration of 1.0% by weight.

Example 6 Test of Functionality Using Chunma Extracts (Liver Function Improvement Effect)

Experiment process

In order to measure the degree of detoxification of the sperm, the activity of GST (glutathione-S-transferase), one of the liver's important detoxification mechanisms, was measured. The prepared reaction reagent was used as a control. The reaction solution without the extract was used as a control. Each extract was added at different concentrations and reacted at 37 ° C. for 5 minutes, and then 1-chloro-2,4 dinitrobenzene was added as a substrate. The reaction was carried out at 37 ° C. for 2 minutes.

After the reaction, 20% TCA was added to terminate the reaction. After centrifugation, the supernatant was measured for absorbance at 340 nm, and then the inactivation and activity ratio of GST were calculated as follows.

Total activity (units) = (A ₃₄₀ /9.6) × dilution factor × (3 ml / 0.1) × crude extract (ml)

Inactive (units / ml protein) = Total active / total protein

% Active = inactive _test / inactive _control × 100

Experiment result

<Table 11> Improvement of liver function (% / mg / ml)

sample Solvent Concentration 0.2 0.6 1.0 Thousand horses Distilled water 175 202 221 Stability and Effectiveness Criteria 100% or more Test method GST

GST is a major detoxification mechanism that catalyzes the reaction of binding various electrophiles to reduced glutathione, and many studies have also been conducted.

Therefore, GST activity measurement can be used to identify medicinal or natural substances that can detoxify or inactivate chemical carcinogens. The results of this experiment showed that Chunma extract increased the activity of more than 175% at all concentrations, and showed a high activity of 221%, especially at the concentration of 1.0% by weight.

Example 7 Test of Functionality Using Chunma Extract (Immune Activity)

Experiment process

Immunity refers to the defense mechanism against stimulation and damage from inside and outside of our body. The measurement of immune activity is carried out through the measurement of immune substances, the growth of immune cells, etc. The increase and decrease of immune cells may be an indicator of the abnormality of our body's immune system.

The experimental immune cell line Raji (RPM1-1640 medium) was dispensed to reach the normal cell growth period (takes about 7 days), and then 96-well plates (100 μl / well of medium adjusted to 5 × 10 ⁴ Cells / ml) were 37 Incubate for 24 hours at 5 ° C. with 5% CO ₂ , administer 100 μl of sample at each concentration, incubate for 48 hours at 37 ° C. with 5% CO ₂ , remove supernatant and cool 10% (w / v) TCA. 100 μl was added and left at 4 ° C. for 1 hour, washed 4-5 times with distilled water to remove TCA, dried at room temperature and 0.4% (w / v) dissolved in 1% (v / v) acetic acid in each well. ) SRB solution was added 100 μl and stained at room temperature for 30 minutes.

Unbound SRB stain was washed 4-5 times with 1% acetic acid and dried, and then 100 µl of 10 mM Tris buffer was added to dissolve the stain, and the absorbance was measured at 540 nm using a microplate reader.

Cancer cell growth (%) = test / control × 100

As it measures the growth of immune cells Raji, it suggests that the food can increase immunity as cell growth increases. Immune function enhancement effect was verified using B cells (Raji), a human immune cell. Cell growth was incubated at 5% CO ₂ , 37 ℃ in RPMI 1640 medium containing 10% FBS, immune function enhancement effect was measured using SRB assay.

Experiment result

TABLE 12 Immune Activity (% / mg / ml)

sample Solvent Concentration 0.2 0.6 1.0 Thousand horses Distilled water 105 125 149 Effect criteria More than 100% Test method SRB assay

Experiments using SRB assays with human B cells (Raji), which play an important role in the production of antibodies in the human immune system, showed that the activity of Raji increased in a concentration-dependent manner. In addition, the cheonma extract showed an increase of 1.49 times or more at a concentration of 1.0% by weight.

<Example 8> Functional assay using cheonma extract (antioxidant activity)

Experiment process

Oxidation causes damage in cell membranes, promotes aging, and is closely related to carcinogenesis and adult diseases. Antioxidant active material is required in terms of aging and prevention of geriatric diseases. In the search for an antioxidant, 0.5 ml of the sample was placed in 5 ml of 4.5 × 10 ^-3 linoleic acid aqueous solution, and incubated at 50 ° C with 1.1 ml each. After every 4 days, 1 ml of TCA, 2 ml of TBA, 0.1 ml of BHT, and 1 ml of SDS were mixed, blown with N ₂ gas, sealed, incubated in a boiling water bath for 15 minutes, allowed to cool, and then cooled with 1 ml of acetic acid and 2 After mixing and shaking the ml chloroform, the supernatant was measured for absorbance at 532 nm by centrifugation at 2,500 rpm for 10 minutes.

Linoleic acid is an essential fatty acid and easily oxidized to polyunsaturated fatty acids. These foods, which exhibit antioxidant properties against linoleic acid, are effective in preventing aging and preventing adult diseases. The search for antioxidants was measured by TBA method.

Experiment result

TABLE 13 Antioxidant Activity (% / mg / ml)

sample Solvent Concentration 0.2 0.6 1.0 Thousand horses Distilled water 23 35 46 Effect criteria 50% or more Test method TBA assay

Linoleic acid, an essential fatty acid, is very easy to oxidize to polyunsaturated fatty acids. Inhibiting the oxidation of such linoleic acid will be effective in preventing aging and preventing adult diseases. At 1.0% by weight of the extract, 46% of antioxidants exhibited antioxidant activity. Antioxidant activity is an antioxidant that inhibits the oxidation of essential fatty acids in foods that are needed in the body when food is produced from plants as an ingredient. Effective in the prevention of

Example 9 Preparation of Granular Tea Containing Chunma Extract

The concentrate obtained in Example 1 was changed to 11, 16, 21% by weight, and refined glucose, anhydroglucose and the like were mixed in the composition as shown in Table 14 to prepare granular tea by a known method, and then the quality characteristics and The results of measuring the functionality are shown in Tables 15 and 16 below.

TABLE 14 Composition of Granular Tea (% / mg / ml)

concentrate Tablet sugar Glucose-Free Cheongung Concentrate Pair Purification Jujube Jujube flavor Ginseng flavor Caramel 11 (40BX) 34.3 50.2 2.6 0.1 0.7 0.05 0.05 0.5 16 31.8 47.7 2.6 0.1 0.7 0.05 0.05 0.5 21 29.3 45.2 2.6 0.1 0.7 0.05 0.05 0.5

<Table 15> Quality Characteristics (% / mg / ml)

content(%) Brix pH Turbidity Chromaticity Solubility (sec) Sensory tests (1-5) ♪ L a b color incense flavor Synthesis 11 10.2 6.01 1.25 71.59 1.48 47.14 30 3.5 3.3 3.2 3.3 16 10.1 5.99 1.55 67.73 3.72 52.61 30 3.6 3.4 3.6 3.5 21 10.0 5.22 1.19 75.42 1.03 49.37 30 3.5 3.4 3.3 3.4

TABLE 16 Functionality (% / mg / ml)

Concentrate addition Anticancer (Stomach Cancer) Hypertension Liver function improvement 16% 56 65 158

Brix showed 10.0-10.2 as the quality characteristics of the granulated tea, and the sensory test showed that the 16% addition of cheonma concentrate was the best.

In addition, the results of all functional tests of granular tea were excellent, but the hypertension control effect was 65% / mg / ml.

Example 10 Preparation of Liquid Tea Containing Chunma Extract

The concentrate obtained in Example 1 was changed to 36.9, 41.9, 46.9% by weight, mixed with liquid fructose, honey, jujube extract and the like as shown in Table 17 to prepare a liquid tea by a known method The results of measuring quality characteristics and functionality are shown in Tables 18 and 19.

Table 17. Composition of Liquid Tea (% / mg / ml)

concentrate Liquid fructose honey Jujube Farmland Cinnamon concentrate Pair Purification Sodium citrate 36.9 (40BX) 55 5 2 0.25 0.5 0.25 0.1 41.9 50 5 2 0.25 0.5 0.25 0.1 46.9 45 5 2 0.25 0.5 0.25 0.1

<Table 19> Quality Characteristics (% / mg / ml)

content(%) Brix pH Turbidity Chromaticity Sensuality (1-5) L a b color flavor Synthesis 36.9 16.0 4.69 3.21 26.53 5.92 30.56 3.5 3.2 3.4 41.9 13.5 4.70 3.07 28.60 5.50 30.81 3.6 3.5 3.6 46.9 13.5 4.63 3.47 24.40 6.21 30.27 3.4 3.3 3.4

TABLE 20 Functionality (% / mg / ml)

Concentrate addition Anticancer (Stomach Cancer) Hypertension Liver function improvement 41.9% 78 82 202

As a quality characteristic of the prepared liquid tea, 16.0, 4.9, and 46.9% of Brix were added 13.5, 4.9, and 46.9%, respectively. As a result of the assay, the high concentration of the concentrate showed high functional effects such as anticancer and hypertension control.

According to the present invention, anyone can easily use Cheonma in the form of granular tea or liquid tea, which has excellent efficacy in anticancer activity, hypoglycemia, hypotension, detoxification, and prevention of various adult diseases. It can also contribute to income increase for farmers.

Claims (6)

delete delete delete In liquid tea, 30 to 60% by weight of cheonma extract as an active ingredient, 30 to 60% by weight of liquid fructose, 5 to 10% by weight of honey, 2 to 10% by weight of jujube extract. delete The liquid tea according to claim 4, wherein the cheonma extract is a concentrate of about 40 Brix.