CN103446358A - Tibetan medicine Iris uniflora pall antioxidant extract and preparation method and application thereof - Google Patents

Abstract

The present invention relates to an effective extract of Iris chrysalis containing anti-oxidation and a preparation method thereof. The effective extract is selected from petroleum ether extract, chloroform extract, ethyl acetate extract, ortho One of butanol extract and water extract or its combination, studies have shown that petroleum ether extract, chloroform extract and n-butanol extract have obvious ability to scavenge hydroxyl radicals in vitro; ethyl acetate extract and water extract The extract has obvious ability to scavenge DPPH (diphenylpicrylhydrazine) lipid free radicals in vitro, therefore, the effective extract can be used to prepare antioxidant drugs and cosmetics.

Description

Tibetan medicine rock raw iris antioxidant extract and its preparation method and use

technical field

The present invention relates to a kind of anti-oxidation extract of Tibetan medicine Iris chrysalis and its preparation method; the present invention further relates to the pharmaceutical composition of the extract; the present invention also relates to the application of the extract in antioxidant drugs or cosmetics; The field of traditional Chinese medicine.

Background technique

In 1956, the British Harman proposed the "free radical theory", which believed that free radicals attack life macromolecules and cause tissue damage, which is the root cause of body aging and a major cause of tumor malignant diseases. With the in-depth study of free radicals, this theory has been accepted by more and more people. Free radicals refer to "any atom or atomic group containing an unpaired electron", which has a great impact on human health. Under physiological conditions, oxygen is at a low level, and its production and removal are in a dynamic balance, but in some pathological conditions, free radicals are excessive, causing damage to the body. In addition to its own damage to biological macromolecules (proteins, DNA), free radicals can also cause lipid peroxidation (LPO) of polyunsaturated fatty acids, and the end product of LPO reaction is malondialdehyde (MDA), so that Proteins, nucleic acids, cephalins are cross-linked and lose their activity. A large number of studies have shown that diseases and gene mutations related to aging, such as cancer, cardiovascular disease, rheumatoid arthritis, diabetes, neurological diseases, aging, etc., are all related to the damage caused by free radicals, eliminating excess free radicals in the body Aging and disease can be prevented. Therefore, seeking to utilize exogenous antioxidants that are easily absorbed by the human body and can remove excess free radicals in the body has become a new research hotspot. However, as the increasingly prominent potential toxicity and carcinogenic effects of synthetic antioxidants are rejected by people, it has become an inevitable trend in the development of antioxidants to find natural, high-efficiency, and low-toxicity antioxidants from plants. The healthcare industry has pointed the way.

The genus Iirs (Iirs) is a large genus of Iridaceae. There are more than 300 species of plants in the world. The plants of this genus are widely used as traditional folk medicines to treat various diseases, such as cancer, inflammation, bacterial and viral infections. . According to reports, compounds with anti-tumor, anti-oxidation, anti-tuberculosis, and Plasmodium crater activities have been isolated from plants of the genus Iris. Among them, Iris tectorum Maxim, Iris halophila Pall, Iris vetiver ( Irish pallida ), and Shegan ( Belamcanda chinensis (L.) DC) all have antioxidant activity reports, especially the root extract of Iris vetiver Used in some cosmetics. Iris uniflora Pall (Iris uniflora Pall), whose Tibetan name is Heji Zema, is a perennial herb of the family Iridaceae. It is produced in Tibet, Qinghai, Inner Mongolia and other places. Its seeds are often used as medicine, which can kill insects and detoxify. It is used in Tibetan medicine. Deworming; external application on sores can remove putrefaction and regenerate muscle, and treat burns; its flowers can improve eyesight, its roots can cure freckles and ringworm, and the whole plant can be burned to black hair. There is no report on its chemical composition and activity, and there is no report on the research and application of Iris lithophylla and its effective parts on the antioxidant effect. We have conducted preliminary research on the antioxidant activity of its total extract and polar parts of different solvents. The activity experiments show that its total extract and some polar parts have good antioxidant activity.

Contents of the invention

One of the objectives of the present invention is to provide an antioxidant extract of Iris rock and its preparation method. The second object of the present invention is to provide a pharmaceutical composition containing the extract as an active ingredient. The third object of the present invention is to provide the application of the extract in the preparation of anti-oxidation and related medicines and cosmetics.

The invention compares the antioxidant activity of the extracts of different solvents, and determines the extract with antioxidant effect and the preparation method thereof.

The Iris rock extract in the present invention is selected from petroleum ether extract (YS-1), chloroform extract (YS-2), ethyl acetate extract (YS-3), and normal One or a combination of butanol extract (YS-4) and water extract (YS-5); preferably petroleum ether extract (YS-1), chloroform extract (YS -2) or a combination thereof; or preferably an ethyl acetate extract, a water extract, or a combination thereof extracted from the ethanol extract of Iris chrysalis; or preferably a n-butanol extract extracted from the ethanol extract of Iris chrysalis.

The extract of the present invention can be obtained through extraction, concentration and series extraction, and its preparation method comprises the following steps:

(1) Take the whole plant or the underground part of Iris chrysalis, soak it in ethanol, extract it with ultrasonic solution, filter it, and repeat the above extraction with the filter residue, filter it, and combine the extracts;

(2) concentrating the extract under reduced pressure to obtain a concentrated solution, and then mixing the concentrated solution with water to obtain a diluted concentrated solution;

(3) Gradient extraction with solvents of different polarities such as petroleum ether, chloroform, ethyl acetate and n-butanol, each solvent was repeatedly extracted, the same organic phases were combined, and concentrated under reduced pressure to obtain the extracts of each solvent.

The method preferably includes the following steps: (1) Take the whole grass of Iris chrysalis or the coarse powder of the underground part, soak it with 30 times the mass of ethanol for 24 h, extract it with ultrasonic solution for 1 h, filter, repeat the above extraction process twice for the filter residue, and filter , combined three extracts, wherein the ethanol used is preferably 60-90% ethanol; (2) Concentrate the extract under reduced pressure to a relative density of 1.10-1.24 mg/mL (50°C) to obtain a concentrate, and then mix the concentrate with water Mix at a volume ratio of 1:1 to obtain the diluted concentrated solution; (3) gradient extraction with different polar solvents, petroleum ether, chloroform, ethyl acetate, and n-butanol, which is equal to the volume of the diluted concentrated solution, each solvent is respectively Repeat the extraction twice and extract twice, combine the same organic phases, and concentrate under reduced pressure to obtain the extracts of each solvent.

The present invention also provides pharmaceutical compositions of each solvent extract prepared by the method. The pharmaceutical composition takes the extract of the present invention as an active ingredient, is composed of a pharmaceutically acceptable carrier, and exists in the form of a suitable pharmaceutical preparation. These preparations can be tablets (preferably sugar-coated tablets, film-coated tablets, etc.) tablets, enteric-coated tablets and sustained-release tablets), capsules (preferably hard capsules, soft capsules or sustained-release capsules), oral liquids, mixtures, buccal preparations, granules, granules, pills, powders , ointment, pill, suspension, solution, injection, powder injection, freeze-dried powder injection, suppository, ointment, plaster, cream, spray, dripping pill, dripping pill, patch, etc.

The present invention further provides the application of various extracts in the preparation of medicines and cosmetics for treating and preventing oxidation. The anti-oxidation-related experiments carried out for this purpose used scavenging DPPH free radicals and hydroxyl free radicals as activity indicators. The experimental results show that the ethyl acetate extract and water extract have obvious ability to scavenge DPPH lipid free radicals in vitro; the petroleum ether extract and chloroform extract have better scavenging hydroxyl free radicals, have anti-oxidation effect, and can be used to prepare antioxidant Drugs and Cosmetics.

Specific implementation plan:

Example 1:

The preparation method of each extract:

1) Total extract extraction

Take 20 g of the whole Iris rock powder, soak it in 600 ml of 90% ethanol for 24 h, extract it with ultrasonic solution for 1 h, filter, repeat the above extraction process twice for the filter residue, filter, and combine the three extracts.

2) concentrated

Recover ethanol from the extract under reduced pressure to obtain a concentrated solution (relative density 1.12g/mL, 50°C), and then mix the concentrated solution with water at a volume ratio of 1:1 to obtain a diluted concentrated solution (100 ml).

3) Gradient extraction

Sequentially use petroleum ether (100 ml/time x 3 times), chloroform (100 ml/time x 3 times), ethyl acetate (100 ml/time x 3 times), n-butanol (100 ml/time x 3 times) Extract the diluted concentrated solution, combine the same organic phases, and concentrate under reduced pressure to obtain petroleum ether extract (YS-1, 0.35 g, extraction rate: 1.75%), chloroform extract (YS-2, 0.62 g, extraction rate: 3.1%), ethyl acetate extract (YS-3, 0.23 g, extraction rate: 1.15%), n-butanol extract (YS-4, 0.14 g, extraction rate: 0.7%) and water extract (YS- 5, 0.67 g, extraction rate: 3.35%).

Embodiment 2 :

The preparation method of each extract:

1) Total extract extraction

Take 20 g of the whole Iris rock powder, soak it in 600 ml of 80% ethanol for 24 h, extract it with ultrasonic solution for 1 h, filter, repeat the above extraction process twice for the filter residue, filter, and combine the three extracts.

2) concentrated

Recover ethanol from the extract under reduced pressure to obtain a concentrated solution (relative density 1.16g/ml, 50°C), and then mix the concentrated solution with water at a volume ratio of 1:1 to obtain a diluted concentrated solution (80 ml).

3) Gradient extraction

Sequentially use petroleum ether (80 ml/time x 3 times), chloroform (80 ml/time x 3 times), ethyl acetate (80 ml/time x 3 times), n-butanol (80 ml/time x 3 times) Extract the diluted concentrated solution, combine the same organic phases, and concentrate under reduced pressure to obtain petroleum ether extract (YS-1, 0.21 g, extraction rate: 1.05%), chloroform extract (YS-2, 0.95 g, extraction rate: 4.75%), ethyl acetate extract (YS-3, 0.33 g, extraction rate: 1.65%), n-butanol extract (YS-4, 0.14 g, extraction rate: 2.8%) and water extract (YS- 5, 0.29 g, extraction rate: 1.45%).

Embodiment 3 :

Scavenging DPPH free radical activity experiment:

Activity experiments were carried out using the polar sites prepared in Example 1.

The antioxidant activity is expressed by the ability to scavenge DPPH free radicals. According to the experimental conditions determined in the pre-experiment, the activity of scavenging lipid free radical DPPH of different samples of Iris petus was determined. Take 2.0 mL of sample solutions with different concentrations, add 2.0 mL of 1.52×10 ^-4 mmol/L DPPH absolute ethanol solution and mix well, and after standing in the dark at room temperature for 30 min, use the solvent as a blank control, and measure the absorbance A at 517 nm ₁ ; Use the same volume of solvent instead of sample solution as a negative control to measure absorbance A ₂ ; use the same volume of absolute ethanol instead of DPPH solution as a blank group to measure absorbance A ₃ . With VC as the control, each group was repeated three times, and the average value was calculated. Calculate the clearance rate according to the following formula:

Clearance rate of sample to DPPH·(%)=[1-(A ₁ -A ₃ )/A ₂ ]×100%

Embodiment 4 :

Hydroxyl free radical scavenging activity test:

Activity experiments were carried out using the polar sites prepared in Example 1.

Antioxidant activity is expressed by the ability to scavenge hydroxyl radicals. According to the experimental conditions determined in the pre-experiment, the activity of scavenging hydroxyl radicals of different samples of Iris lithosum was determined. Mix 1.5 mL of 1.8 mol/L salicylic acid solution and 2.0 mL of 1.8 mol/L FeSO ₄ solution in the test tube, add 1 mL of sample solutions of different concentrations, then add 0.1 mL of 0.03% H ₂ O ₂ , shake well, Incubate at 37 ℃ for 30 min, measure the absorbance A _i of the sample group at 510 nm; at the same time replace the H ₂ O ₂ in the sample group with 0.1 mL distilled water, measure the absorbance A _j of the sample reference group; replace the sample in the sample group with 1.0 mL solvent, measure the absorbance A ₀ of the blank group. With VC as the control, each group was repeated three times, and the average value was calculated. Calculate the clearance rate according to the following formula:

OH scavenging rate (%) = [A ₀ -(A _i -A _j )] /A ₀ ×100%

Table 1 The in vitro scavenging activity of DPPH free radicals in various parts of Iris lithophylla

Note: IC ₅₀ ±s, n=3

Table 2 Hydroxyl free radical scavenging activity test in vitro of various parts of Iris rockus

The results show that the petroleum ether extract (YS-1), chloroform extract (YS-2), ethyl acetate extract (YS-3), and n-butanol extract (YS-4) prepared by the process of the present invention and water extract (YS-5) have good antioxidant activity, among which petroleum ether extract and chloroform extract have good ability to scavenge hydroxyl radicals, and their antioxidant capacity is about 5/6 and 3/4 of VC respectively; ethyl acetate Ester extract and water extract have good ability to scavenge DPPH free radicals, and the antioxidant power is 1/7 and 1/11 of VC respectively; while the ability of n-butanol to scavenge hydroxyl free radicals is in the range of 60-1250 μg/mL. It is maintained at about 50%, and it is stronger than VC when the concentration is lower than 250 μg/mL.

Claims (9)

1. A kind of active extract of Iris rock with anti-oxidative effect, it is characterized in that described extract is selected from petroleum ether extract (YS-1), chloroform extract (YS-2) that extracts from the alcohol extract of Iris rock ), ethyl acetate extract (YS-3), n-butanol extract (YS-4), water extract (YS-5) or a combination thereof. 2. An active extract of Iris rock according to claim 1, characterized in that said extract is selected from petroleum ether extract (YS-1), chloroform extract (YS-1) extracted from Iris rock alcohol extract. -2) or a combination thereof. 3. A kind of active extract of Iris rock according to claim 1, it is characterized in that described extract is selected from the ethyl acetate extract that extracts from the alcoholic extract of Iris rock, water extract or its combination. 4. an active extract of Iris rock according to claim 1, characterized in that said extract is selected from the n-butanol extract extracted from Iris rock extract. 5. A method for preparing the active extract as claimed in any one of claims 1-4, characterized in that the method comprises the following steps: (1) Take the whole grass of Iris petrifuse or the coarse powder of the underground part, soak it with ethanol, Ultrasonic assisted extraction, filtration, and the filter residue repeated the above extraction, filtration, and combined extracts; (2) Concentrate the extract under reduced pressure to obtain a concentrated solution, and then mix the concentrated solution with water to obtain a diluted concentrated solution; (3) Use Gradient extraction with solvents of different polarities, petroleum ether, chloroform, ethyl acetate and n-butanol, repeated extraction, merging the same organic phases, and concentrating under reduced pressure to obtain the extracts of each solvent; The method preferably comprises the following steps: (1) Take the whole grass of Iris chrysalis or the coarse powder of the underground part, soak it with 30 times the mass of ethanol for 24 h, extract it with ultrasonic solution for 1 h, filter, repeat the above extraction process twice for the filter residue, and filter , combined three extracts, wherein the ethanol used is preferably 60-90% ethanol; (2) Concentrate the extract under reduced pressure to a relative density of 1.10-1.24 mg/mL (50°C) to obtain a concentrate, and then mix the concentrate with water Mix at a volume ratio of 1:1 to obtain the diluted concentrated solution; (3) Gradient extraction with different polar solvents petroleum ether, chloroform, ethyl acetate and n-butanol equal to the volume of the diluted concentrated solution, and repeat for each solvent Extracted twice, combined the same organic phases, concentrated under reduced pressure to obtain the extracts of each solvent. 6. A pharmaceutical composition comprising the active extract of any one of claims 1-4 and a pharmaceutically acceptable carrier. 7. according to the described pharmaceutical composition of claim 6, the preparation form of described pharmaceutical composition can be tablet, capsule, oral liquid, mixture, buccal preparation, granule, electuary, pill, powder, ointment, pill , suspensions, solutions, injections, powder injections, freeze-dried powder injections, suppositories, ointments, plasters, creams, sprays, dripping pills, dripping pills, patches, etc. 8. according to the described pharmaceutical composition of claim 7, it is characterized in that: described tablet is selected from sugar-coated tablet, film-coated tablet, enteric-coated tablet or sustained-release tablet; Described capsule is selected from hard capsule capsules, soft capsules or sustained-release capsules. 9. The application of the active extract described in any one of claims 1-4 in the preparation of medicines and cosmetics for treating and preventing oxidation.