KR0168721B1 - Process for preparing the extracts of cassiae semen, and pharmaceutical compositions containing them useful for antioxidant, free radical scavenger, antimutagent and opthalimic disease preventing agent - Google Patents

Abstract

Cassiae semen (Cassiae semen) is a seed of the superdestructor (Cassia obtusifolia L.), or Cassia tora L. is used as a sanitary, formal, diuretic, and laxative in oriental medicine, and in the sense of consent, it is called as a blue-eyed name (淸 肝 明目). When boiled with boiled water, the liver is known to brighten the eyes.

The present invention finds that the active ingredient of no-lubrofusarin is not contained at all when the boiled extract is extracted by only boiling with water, and to obtain an extract with a high content of noro-brobrofusarin, which is the main ingredient of the defector. It is to provide a method for producing a deflector extract containing a high concentration of nor- lubrofusarin by extracting the deficiency powder with an aqueous 30% to 100% ethanol solution.

In addition, the present invention is to prepare extracts having excellent activity against the deficiency known to be good for the eye since ancient times, and to check the antioxidant activity, free radical scavenging effect and antimutagenic effect of these extracts and the active ingredient contained in the eye It is to provide a use for the prevention of aging and eye diseases.

Description

Method for preparing the extract of C. oleracea and its antioxidant, free radical scavenging and antimutagenic agents

The present invention relates to a method for the preparation of extracts containing a large amount of active ingredients from the cultivars and to their use, in particular, to the antioxidant activity, free radical scavenging activity and antimutagenicity of these extracts and active ingredients contained in the extract will be.

Cassiae semen (Cassiae semen) is the seed of the superdestructor (Cassia obtusifolia L.) or Cassia tora L. is used as a sanitary, formal, diuretic, and laxative in oriental medicine, and in the sense of consent, it is called the bluish name. It is known that if you drink it, it will be good for your liver and brighten your eyes. Therefore, until recently, many people have decoctioned the boiled water with boiling water and drink it as tea.

In the meantime, anthraquinones such as chrysophanol, emodin, and physcion, obtusifolin, obtusin, chryso-obtusin, nord Anthraquinone glycosides, such as nor-rubrofusarin and rubrofusarin, have been isolated and identified. However, to date, it is not clear what component of the defect is active.

In addition, the present inventors found that when the extract was boiled with water only and extracted and drinked, no active ingredient nor-robrofusarin was contained.

Accordingly, it is an object of the present invention to provide a method for producing an extract having a high content of nolu-brobrofusarin, which is a main component of the deficiency.

Another object of the present invention is to identify the use of the extract and the active ingredient contained in the extract by the above-described method to provide a food or pharmaceutical use.

As a result of several experiments with a variety of extraction solvents, the present inventors have found that the method of extracting 30% to 100% of an ethanol aqueous solution contains the most active ingredient, nor lubrofusarin. In this case, it was possible to obtain the extract of the deficiency containing no more than 0.005% of nor- lubrofusarin.

In addition, through in vitro experiments, it was found that these extracts were excellent in antioxidant activity, free radical scavenging activity and antimutagenic activity. In particular, free radicals and free radicals produced in the body weaken the various regulating muscles of sensory organs such as the eye and harden the arteries to the eye to reduce blood flow to the retina, while also reducing the transparency of the lens. In addition, free radicals and free radicals injure β cells in the body, leading to diabetes, and secondly to ophthalmic diseases such as cataracts. In addition, ultraviolet rays and various mutagenic substances can also cause cataracts by mutating in the body. Therefore, the antioxidant extract, free radical scavenging activity and antimutagenic activity of the extract of the present invention provides the use of the eye as an anti-aging and eye disease prevention agent.

Hereinafter, the present invention will be described in detail with reference to Examples, but the present invention is not limited to these Examples.

Example 1

· Isolation and Assay of Nor-Lupurofusarin

After clarification of the deflector, the mixture was refluxed with a 70% methanol and reflux-cooled for 3 days while heating. The filtrate was dried under reduced pressure after filtration. 50 g of the dried extract was dissolved in a small amount of methanol, and then repeatedly purified by silica gel chromatography using ethyl acetate and methanol as a solvent to obtain about 50 mg of pure nor- lubrofusarin. The structure of this material was identified by comparison with published data and the structure is shown in Figure 1.

In the quantitative determination of nor-brobrofusarin, the extractor was powdered, extracted with 70% ethanol solution for 3 days, filtered and concentrated under reduced pressure. Dissolve 100 mg of the sample in 50 ml of methanol, and filter it to obtain a sample solution for HPLC. At this time, the column was ODS (manufactured by Shimazu), the mobile phase was Methanol: Water: Acetic acid = 40: 60: 0.5, the detector was UV 254 nm, and the flow rate was 0.8 / ml. In this case, nor-brobrofusarin showed a holding time of about 30 minutes.

Extraction method of Clarifier extract

In order to know the effect of the factors on the extraction of the deficiency extracter powder (60g) as a sample was extracted with varying concentration of ethanol to 600ml ethanol aqueous solution. At this time, when ethanol 0% (aqueous solution 100%) was boiled for 1 day, 30%, 70%, 100% when extracted for 3 days at 20 ℃. The extract was filtered and concentrated under reduced pressure to obtain the extract.

The final yields of these extracts and the content of nor- lubrofusarin contained in these extracts are shown in Table 1 below.

As can be seen in Table 1, the higher the ethanol concentration, the lower the final yield of the extract of C. oleracea, but no-rubrofusarin in the C. extract was hardly detected when extracted with 100% aqueous solution, but in 70% ethanol and 100% ethanol. The content was high.

Example 2

Antioxidant activity.

Determination of antioxidant activity of 70% ethanol extract and nor- lubrofusarin extracted from the defector was measured by TBA assay using a lipid peroxidation system by the Fenton reaction. In other words, 5 ml of 25 mM Trizma HCl / 0.75 mM KCl buffer containing 10ul of ethyl linoleate, 0.2% SDS, ferrous chloride, and 2umol of hydrogen peroxide were added to the ethanol extract of the samples, followed by incubation for 16 hours at 55 ° C. The reaction is stopped after adding 50ul of% BHT. 0.3 ml of this solution was added to a test tube, and 0.05 ml of HCl 3.1 ml and 0.67% TBA solution were added and boiled for 30 minutes. Next, after cooling the reaction solution, 4.0 ml of 85% butanol was added, and after centrifugation, the absorbance of the butanol layer was measured at 535 nm to obtain an antioxidant effect (%). The measurement results of this antioxidant effect are shown in Table 2 below.

As can be seen in Table 2, the extract of C. vulgaris showed higher activity at 1mg / ml and 0.01mg / ml as the ethanol content was higher in the extraction solvent. M-10 The dose-dependent antioxidant activity was shown between M.

Free radical scavenging action

Determination of free radical scavenging activity of 70% ethanol extract and nolu-brobrofusarin at 60 μM 1,1-diphenylyl-2-picryl hydrazyl (1,1-diphenyl-2-picrylhydrazyl; DPPH) ethanol solution 2 ml 2 ml of DMSO solution was added to the sample, reacted at room temperature for 30 minutes, and the absorbance was measured at 520 nm to determine the free radical scavenging effect (%). The measurement results are shown in Table 3 below.

As can be seen in Table 3, the extract of C. vulgaris showed more activity at 0.01 mg / ml and 1 mg / ml than 70% ethanol extract, and no-rubrofusarin 10 M -10 A dose-dependent free radical scavenging action was shown between M.

Antimutagenic activity

Antimutagenic activity of 70% ethanol extract and no-robrofusarin was determined as follows.

That is, the mouse performs the micronucleus suppression effect using peripheral blood. At this time, the positive control material used was intraperitoneal injection of 1 mg / kg of mitomycin C (MMC), and peripheral blood was collected from the tail after 48 hours. The effect of suppressing micronucleation (%) according to the oral dose of fusarin was observed. The measurement results are shown in Table 4 below.

As can be seen in Table 4, the extract of L. oleracea showed activity at 1 mg / kg and 10 mg / kg, and no-rubrofusarin at 0.01 mg / kg and 0.1 mg / kg for micronucleation for mitomycin seed (MMC). Inhibition showed antimutagenic activity.

As shown in the above results, the composition showed antioxidant and free radical scavenging effects at 0.01-1 mg / ml in vitro and effective at a single oral dose of 1-10 mg / kg in mice (ICR, 6-8 weeks of age). Antimutagenic effects were shown.

Example 3

[Production of Preparation]

Tablets are prepared by mixing 20-200 mg of this composition with 150 mg of lactose, 25 mg of starch and 10 mg of magnesium stearate per tablet. The solution is prepared as a solution by adding 20-200 mg of the composition, 2 g of isomerized sugar, 2 g of white sugar, and 10 g of pear juice per 10 ml and dissolving it in purified water.

Example 4

[Acute Toxicity]

For reference, the acute toxicity test of this composition was applied to mice (ICR, 6-8 weeks of age). As a result, no males died at 5 g / kg (oral administration) in male mice. Did not appear abnormal. The same results were obtained in the male mice as in the male, and thus the composition was considered safe.

As shown in the above examples, the extraction of the active ingredient from the cultivars according to the method of the present invention can extract nor- lubrofusarin in high yield, in particular, these extracts have antioxidant, free radical scavenging and antimutagenic activity. It is useful as an anti-aging and eye disease preventive agent of the eye supported by these activities.

Claims (2)

Method for producing a defect extract containing nolu-brobrofusarin characterized in that the powder to the defect is extracted with an aqueous 30 to 100% ethanol. It is characterized in that the extract is extracted from the 30% to 100% ethanol aqueous solution, as a main component, to which antioxidant, free radical scavenging and anti-oxidation, to which isosaccharides, excipients and other pharmaceutically acceptable additives can be added. Mutant.