KR101386791B1 - A composition of kamisochimtang for anti-depression - Google Patents

Abstract

The present invention relates to an anti-depression composition containing an extract obtained from kamisochimtang which is a mixture of 4.0 parts by weight of Cyperus rotundus, 2.0 parts by weight of lindera roots, 0.5 parts by weight of elecampane, 0.5 parts by weight of Glycyrrhizae radix, 2.0 parts by weight of platycodon, and 2.0 parts by weight of Poncirus trifoliate. The present invention provides an anti-depression composition which has an excellent anti-depression effect, a reduction of side effects, and effectiveness.

Description

  A composition of kamisochimtang for anti-depression

The present invention comprises an antidepressant comprising an extract obtained from Kamiso Oh-Bangtang, which is a mixture of Hyangbuja 4.0 parts by weight, oak 2.0 parts by weight, Mokyang 0.5 parts by weight, licorice 0.5 parts by weight, Gil length 2.0 parts by weight and crust 2.0 parts by weight. It relates to a composition.

 Depression is caused by a decrease in the amount of certain neurotransmitters in the brain. Representative neurotransmitters are serotonin and norepinephrine. Thus, antidepressants have therapeutic effects through increasing the amount of these neurotransmitters in the brain. Many antidepressants used in the past include imipramine, amitriptyline, and trazodone. These drugs have been used in clinical practice because of their good therapeutic effect, but these drugs have the effect of blocking neurotransmitter receptors such as acetylcholine and histamine, causing inconveniences such as constipation, dry mouth and drowsiness. . Recently developed and used antidepressants have minimized these side effects, making patients feel almost uncomfortable when taken. Representative drugs include Prozac, Zoloput, Cerrojat, and Dumyrox, which act to selectively increase the amount of serotonin, especially among the two neurotransmitters. A drug that increases epinephrine at the same time, it is similar in function to antidepressants of the past but has much improved side effects.

     Treatment of depression is emerging as an important social challenge. According to Dr. Lopez's report from the World Health Organization (WHO), 2020 is ranked by disease by a measure that introduces the concept of the Disability-Adjusted Life Year (DALY), `` the number of years a person has lost a healthy life. '' Depression is expected to become the second most common disease group after ischemic heart disease in 2010. In developed countries, the number of patients is increasing at such a rapid rate that depression is predicted to rise to the leading position. Hypotheses about the biological causes of depression vary, but to date, the prevailing view is that disorders of the neurotransmitter system such as adrenaline, dopamine, and serotonin are dominant. This is mainly due to the action of tricyclic antidepressant (TCA) and selective serotonin re-uptake inhibitor (SSRI). SSRI, which is currently used the most, does not show a big difference in comparison with TCA. Another important aspect is the serious problem of drug interactions. In particular, Koreans may have toxicity problems because the poor metabolizer of liver enzymes is four times higher than that of Westerners. Therefore, the need for a new antidepressant for the Korean constitution is increasing.

    Currently, several antidepressants based on herbal medicine are commercially available as an alternative to antidepressants. The most representative of these, "yarsin," is an antidepressant released in August 1999. The main ingredient is LI160. It is derived from a plant called John's Wort (hypericum perforatum), which was used as a folk remedy in the United Kingdom. Efficacy is effective in treating physical symptoms such as depression, anxiety and insomnia, which are accompanied by depression, and the safety is relatively high. The prescription of this drug is six times higher than fluoxetine in Germany. Efficacy comparisons with existing antidepressants range from 6-18% lower response rates than tricyclic antidepressants to reports of mild to moderate outpatient depression with at least the same effect as serline, one of SSRIs. It seems to require further accumulation of research. However, in North America's mainstream psychiatry, where the use of herbal medicines and alternative-complementary medicines is not over 40% of the population, the use of banks in the treatment of dementia and the efficacy of this new antidepressant drug, which is the main ingredient in dementia, It is a movement to admit that there is evidence for this.

     Since Korea has thousands of years of experience in using herbal medicines, it is highly likely to obtain better antidepressant candidates among herbal medicines than other compounds. To get an antidepressant, you first need the concept of a medical disease that corresponds to depression, and 'depression' is the closest concept. Congestion literally is a symptom of feeling depressed, accompanied by low mood and low motivation, as well as feeling of chest tightness and pain throughout the body. The herbal medicine Kamisoo-Bangtang is a Soo that is proposed to be used for the heart beat due to the problem of qi, called "凡 보 宜… 小 烏 沈 湯" in "Dongyi Bogam". In order to better control the symptoms of chest tightness and chest pain, which are common in depression, it is prescribed by Gil-Sang, which is used to treat depression with physical pain. However, antidepressant effects through objective experimental studies of Kamisosochimtang are not well known, and as a related art related to depression, Korean Patent Publication No. 10-2012-0130973 discloses herbal medicines such as Hyangbuja, Baekbokryeong, and Licorice. And a pharmaceutical composition for preventing and treating depression is disclosed, including extracts of Huangshan, which is the same as the present invention is related to depression, the material is different.

Although depression is one of the common diseases with long duration, high chronicity, relapse rate, psychosocial and physical damage, and high suicide rate, the treatment rate is only 65-70% for any of the many antidepressants currently in use. In addition, current antidepressants can cause many unwanted side effects. Not only are natural products widely used, they have a long history as medicines in Asian societies. Some of them may be safe and effective alternatives to depression treatment, like St John's Wort, a Western-made herbal medicine. Depression occurs when the synaptic concentrations of monoamine neurotransmitters (dopamine, noradrenaline, serotonin, etc.) are out of balance, and various drugs that restore it have antidepressant effects. Meanwhile, depression, which is known to occur in autumn and winter, that is, seasonal affective disorder, is known to be caused by overproduction of melatonin.

The present invention provides an antidepressant composition having an effective antidepressant effect while reducing side effects and having an effective antidepressant effect through a composition comprising flavor extract, ointment, throat, licorice, gilyeong and persimmon extract.

 The present invention provides an anti-depressive composition comprising an extract obtained from a mixture of flavor, medicinal herb, medicinal herb, licorice, gilyeong and crust.

 In another aspect, the present invention provides an anti-depressive composition, characterized in that the weight ratio of the extract, hyangbuja, oakyang, licorice, licorice, clove and crust 4.0: 2.0: 0.5: 0.5: 2.0: 2.0.

Finally, the present invention provides an antidepressant composition, wherein the extract is orally administered at a concentration of 94 mg / kg or 282 mg / kg.

Antidepressant composition according to the present invention significantly reduced immobility time in the forced swimming test of the mouse model, significantly increased serotonin, melatonin in the brain methanol extract, significantly increased serotonin production in the brain turbid solution, Restoration of the vertebral cell layers (CA1, CA3), neurons and dentate gyrus (DG), which disappeared from the lesions of the hippocampus, showed an antistress effect.

1 is a photograph of the forced swimming test of the mouse for the antidepressant composition of the present invention.
Figure 2 is a chart showing the effect on immobility time in the forced swimming test of mice for the antidepressant composition of the present invention.
Figure 3 is an HPLC chromatogram for the antidepressant composition and standard of the present invention.
Figure 4 is a chart showing the effect on serotonin production in the brain turbidity of mice for the antidepressant composition of the present invention.
Figure 5 is an optical micrograph showing brain tissue damage in the hippocampus with chronic restraint stress of the mouse against the antidepressant composition of the present invention.

Hereinafter, the present invention will be described in more detail with reference to examples and application examples. The following examples are merely illustrative of the present invention in detail, and thus, the nature of the technical idea of the present invention is not changed or reduced in scope. It will be apparent to those skilled in the art that various embodiments and applications not shown in the following examples are possible.

First, the drug as a component of the present invention will be described.

Hyangbuja is the root of the perennial herb, Cyperus rotundus L., a member of the Cyperaceae family, which has a flat, spicy, sweet and bitter taste, and helps to break up the qi and bleeding. It is effective.

Linderae Radix is a root of the camphor tree, which is harvested in the winter and spring and sun-dried. It is warm and tastes hot. The upper part enters the rain and lungs to swell the chest abdomen, and the lower part enters the neural bladder to remove the bladder's cold air and pain.

Mokhyang (木香) is a perennial herb that belongs to Asteraceae. It has a strong fragrance and contains abundant volatiles, which makes it good for use in the digestive organs. It treats severe stomach pain and weakness of the stomach, and swells and pains in the chest and abdomen, vomiting, diarrhea, and pulling from the lower abdomen to the thigh. Indigestion, stomach expansion, stomach swelling and vomiting when you do not vomit when you write with a new song.

Licorice (甘草) is the root and mycelia of the perennial herb, Glycyrrhiza ssp. It is warm, sweet, without poison, releasing a hundred poisons and vomiting tablets, and alleviating 72 stones and 1200 herbaceous ingredients and making it effective with all medicines.

Gilkyung (桔梗) is a dried perennial bellflower root, flat nature, taste is bitter and spicy. It has been used since ancient times for anti-inflammatory effects, especially for sore throat.

The crust is cold, bitter, sour, very, and nonpoisonous. It is better to cough with scrapping, to break through the wall in the chest, to pass through the large intestine, to cut the window, to flock to the peninsula, and to open the blockage. It cleans the walls, drains the water, heals the walls and the frauds, and heals the itchy and paralyzed winds, long winds, and hemorrhoids.

The researcher concluded that Kamisoo-chimtang discovered the possibility of antidepressant through long research, and completed the present invention by obtaining antidepressant effect from animal model, biochemical and molecular biological profile.

Hereinafter, examples and experimental examples of the present invention will be described in detail. It should be noted, however, that the following examples and experimental examples are illustrative of the present invention and that the present invention is not limited to the following examples and experimental examples.

Preparation and Administration of Complex Herbal Extracts

Selected by purchasing at Dunsan Oriental Hospital of Daejeon University, Kamiso Ochi-tang is a prescription consisting of 4g of Hyangbuja, 2g of oak, 0.5g of Mokyang, 0.5g of licorice, 2g of Gilkyung, and 2g of crust.

2,000 ml of distilled water was added to the 2-pack of Kamisoo-Jangtang (22 g), and the liquid obtained by extraction in a hot water extractor for 3 hours was filtered by suction, and then concentrated by a rotary vaccum evaporator. This was again obtained by using a freeze dryer (freeze dryer) 4.7g of completely dried Kamiso sediment extract was stored at -84 ℃ frozen and diluted to an appropriate concentration according to the purpose and then used as a sample of the following test example.

Kamisoochitang was administered orally at the concentration of 94 mg / kg and 282 mg / kg 2 hours before the immobilization stress for 21 days daily.

<Reference example>

1. Experimental animals

Supplied with 6-week-old male 20-22g mice C57bl / 6 (Daehan Biolink, Korea), supplies solid feed (without antibiotics, Samyang feed) and water until the day of the experiment. Temperature 22 ± 2 ℃, humidity 55 ± 15 % Was used in the experiment after one week of adaptation in a light-dark cycle environment of 12 hours.

2. Statistical Processing

All measurements were once averaged ± standard deviation to facilitate comparison. In order to verify the statistical significance of the measured values, nonparametric disease was used in consideration of the nature of the experiment. The Kruskal-Wallis test was used for the comparison between the whole groups, and the Mann-Whitney test was used for the comparison between the two groups. SPSS for Windows (version 10.0) was used as a program for statistical processing.

<Experimental Example>

1. Depression Mouse Model by Body Restraint Stress

A male C57bl / 6 mouse was used to make a 50ml conical tube (FALCON, BD) with a number of holes in the top, side, and stopper. The depression model of mice exposed to long-term restraint stress was subjected to a forced swimming test. The forced swimming test was performed by using a clean tap water in a cylindrical acrylic cylinder (20 cm in diameter and 40 cm in height) with the top open. Pour water temperature 25 ± 2 ° C. up to 15 cm from the bottom of the cylinder. The depressed mouse model is forced to swim for 15 minutes, pulled out of the water, wiped with a dry towel and returned to the breeding box (pretest session). After 24 hours the mice were placed on the same equipment for 5 minutes (test session). The entire test run was recorded using a recording device such as a camcorder, and the time taken for the next animal's behavior to be measured during the test time (5 minutes) was recorded from the recorded images. Alternatively, the time and frequency of three animal behaviors were measured by sampling every 5 seconds. Forced swimming tests used immobility behavior as an indicator of despair, with no movement other than the movement required to bring the head out of the water in an unscathed, floating motion. The horizontal movement around the cylinder was classified into a more aggressive swimming behavior than the immobility behavior, and a rather intense climbing behavior in which the forefoot was kicked out of the water toward the wall. Each behavior may be measured during a test session or compared to the last 5 minutes of the pretest 15 minutes and the 5 minutes of the test session, as shown in FIG. 1.

2. Forced swimming test

As a result of observing the effect of long-term administration of Kamisoochi-tang on immobility time in the forced swimming test in the model of chronic restraint stress mice, as shown in FIG. 2, the control group (CIS-NC) compared to the normal group (C57bl / 6). In this forced swimming test, the immobility time increased, and the experimental group treated with long-term Kamisoo-tang (SOCTH) showed statistically significant decrease in immobility time compared to the control group. However, the positive control group fluoxetine did not show a significant difference from the control group (CIS-NC).

3. Analysis of Standard Components and Neurotransmitters Using Liquid Chromatography (HPLC)

 HPLC-DAD detector for brain methanol extract analysis was used and Chemstation software was used. As a condition of liquid chromatography, the temperature of the column oven was 25 ° C., the analytical wavelength was 220 nm, the flow rate was 1.0 ml / min, and the column was a Luna C18 column (250 mm ⅹ 4.6 mm; particle size 5 μm, Phenomenex, Torrance, CA). , USA). The mobile phase was used by adding 100% tertiary distilled water (containing 0.1% TFA) as solvent A and 0.1% formic acid as solvent B to 100% acetonitrile, respectively. The solvent system is shown in Table 1 as a solvent gradient. The standard component samples were serotonin and melatonin from Sigma USA, and the components were analyzed in brain methanol extract.

Solvent Slope Conditions End time (min) Flow rate (ml / min) A ^a B ^b 0 0.3 85 15 10 0.3 80 20 30 0.3 65 35 35 0.3 65 35 40 0.3 85 15

* ^a 0.1% formic acid solution

^b Acetonitrile with 0.1% formic acid

Standards were identified by identifying the patterns of serotonin and melatonin through matching of retention times through HPLC-DAD in the identification of the standards contained in the components of depressed mice (FIG. 3A). As a result of depression animal experiments, serotonin and melatonin in normal C57bl / 6 mice (Fig. 3B) were significantly decreased in the depression mice control group (Chronic immobilization stress (CIS-NC)) as shown in [Figure 3]. (FIG. 3C), the positive control fluoxetine-treated group showed an increase in melatonin compared to the control group (CIS-NC) (FIG. 3D), and the Kamisoo-Bangtang 94 mg / kg-treated group in the control group (CIS-NC). In comparison, serotonin was increased (FIG. 3E), and serotonin and melatonin were significantly increased in the Kamisoochitang 282 mg / kg administration group (FIG. 3F).

4. Determination of Serotonin Level in Mouse Brain Turbidity

After the experiment, the brains of the mice were extracted and frozen at -20 ° C, and then the cortex, the hippocampus, and the striatum were separated. 200-500 ml of extraction buffer (0.3 M sucrose, 0.15 mM spermine, 0.5 mM spermidine, 10 mM HEPES (pH 7.9), 1.5 mM MgCl2, 10 mM KCl, 0.5 mM DTT, 0.2 mM PMSF, 0.1% protease inhibitor, 0.1% phosphatase inhibitor, 0.5% NP 40) was added to homogenizer and homogenized, followed by addition of extraction buffer to a final volume of 0.2ml, followed by lysis at 4 ° C for 2 hours. After the reaction was completed, the sample was placed in a 1.5 ml tube, vortexed three times for five minutes, centrifuged at 4 ° C. and 15,000 rpm for 15 minutes, and 100 μl of the supernatant was taken. Serotonin EIA kit (ADI-900-175, Enzo life Sciences) was used to wash each well with wash buffer according to the manufacturer's instructions, and 100 μl of Assay diluent was added to block wells for 1 hour and then incubated at room temperature. After diluting the standard and diluting the supernatant 20 times, the microplate was washed and 100 μl of each standard and the supernatant were added. The wells were blocked for 2 hours and then incubated at room temperature. After washing the microplate and making a waring detector, put 100µl into each well, block the well for 1 hour, and incubate at room temperature. Then, wash the microplate, make a substrate solution, put 200µl into each well, and put it in the dark for 30 minutes at room temperature. Incubated with. 50 ㎕ of each stop solution was added to each well, and the absorbance was measured at 405 nm using a microplate spectrophotometer.

As a result of observing the effect of long-term administration of Kamisoochimtang on serotonin production in brain turbidity in the model of chronic constrained stress, serotonin production in the control group (CIS-NC) compared to the normal group as shown in FIG. The experimental group which received fluoxetine (p <0.01) and long-term Kamisoo-Bangtang (p <0.001) as a positive control group showed a statistically significant increase in serotonin production compared to the control group. It could be confirmed that having.

5. Brain H & E staining

After completion of the experiment, the brains of the mice were fixed in 10% formaldehyde solution, and then rinsed with running water for 8 hours. This was sliced with a microtome, hematoxylin & Eosin staining and observed on an optical microscope.

 Brain disease is usually known to begin in the hippocampus area of the brain, and the hippocampus is the part of the brain that is first damaged when a brain disease occurs, and is known to be responsible for memory in the brain. So the hippocampus has two thin layers of neurons that overlap each other, one called the dentate gyrus and the other the Ammon's horn. Two of the four sites of Ammon's angle, two of which are important, are CA3 and CA1, and CA3 {Hippocampal area CA3 (cornu Ammonis = Ammon's horn)} and CA1 {Hippocampal area CA1 (cornu Ammonis = Ammon's horn)} are cornu Ammonis The abbreviation for Ammon is Latin. Thus, the CA1 and CA3 regions of the hippocampus are related to recent memory, and learning and memory can occur at synapses, which are useful for studying synaptic plasticity in the hippocampus.

In this study, stratum orion, stratum radiatum, oligodendrocytes-like cells, astrocytes-like cells, pyramidal cell layers (CA1, CA3), neurons, and dentate gyrus, DG) and the like can be seen in Figure 5 disappearance of the control group (CIS-NC) compared to the normal group (C57bl / 6-WT). The experimental group (94 mg / kg, 282 mg / kg) that received fluoxetine (p <0.01) and Kamisoo-Bangtang (p <0.001) as the positive control group had the cortical cell layer (CA1, CA3) disappeared at the lesion site compared to the control group. , Neurons and dentate gyrus (DG) were found to have recovered, indicating that they have antidepressant activity.

Claims (4)

Antidepressant composition, characterized in that it comprises an extract obtained from a mixture of Hyangbuja, Yakyang, Mokko, Licorice, Giltyeong and Crust.
The antidepressant composition according to claim 1, wherein the extract has a weight ratio of hyangbuja, oak, neck, licorice, gilyeong and crust 4.0: 2.0: 0.5: 0.5: 2.0: 2.0.
The antidepressant composition according to claim 2, wherein the extract is orally administered at a concentration of 94 mg / kg.
The antidepressant composition according to claim 2, wherein the extract is orally administered at a concentration of 282 mg / kg.

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