CN1164267C - Medical use of chlorogenic acid and its preparation - Google Patents

Abstract

The present invention discloses chlorogenic acid and isochlorogenic acid prepared from natural plants. Pharmacological experiments prove that the chlorogenic acid and isochlorogenic acid have obvious activity resisting SARS viruses. The chlorogenic acid and the isochlorogenic acid with the activity can be prepared into compositions of different preparations, wherein the chlorogenic acid, the isochlorogenic acid and vitamin c are used as main active components, and excipient or auxiliary agents or carriers are added to the main active components according to different requirements. The compositions can be applied to the preparation of medicine for treating and preventing diseases caused by SARS viruses.

Description

Medical use of chlorogenic acid and its preparation

technical field

The present invention relates to compounds with biological activity, more specifically chlorogenic acid, and its anti-SARS virus activity used as a drug for preventing and treating atypical pneumonia.

Background technique

Chlorogenic acid, isochlorogenic acid, flavonoids, etc. exist in a variety of plants, such as honeysuckle, coffee, Eucommia, Artemisia annua and other plants.

Honeysuckle, also known as double flower, silver flower, and honeysuckle, is the flower bud of the perennial winding woody vine of Lonicera family Lonicera. It is distributed throughout the north and south of my country. Pick when the buds are in buds in early summer, dry in the sun or in the shade, and use raw or stir-fried. Cold in nature, sweet in taste, enters the lung, stomach, and large intestine meridians. Function: heat-clearing and toxic substances removing, evacuating wind-heat, cooling blood and stopping dysentery. Chinese medicine believes that honeysuckle is sweet and cold, light in quality, and its air is fragrant. It can not only clear away heat and detoxify, but also dispel wind-heat. . Chinese folks often make tea with honeysuckle and raw licorice in moderation, which can be used to prevent heatstroke and temperature in summer, clear heat and quench thirst. There are also honeysuckle, Ophiopogon japonicus, Platycodon grandiflorum, ebony, licorice and other bubble drinks to treat acute and chronic pharyngitis.

Chlorogenic acid (Chlorogenic acid), synonym: coffee tannic acid, is one of the most important natural products in phenylacryloylquinic acid ester compounds, mainly exists in some traditional Chinese medicines with heat-clearing and detoxifying properties, such as honeysuckle, It is one of the functional ingredients of these traditional Chinese medicines. The structural formula of chlorogenic acid is as follows:

Chlorogenic acid has a wide range of antibacterial effects. Oral or intraperitoneal injection can increase the central excitability of rats. It can increase the intestinal motility of rats and mice and the tension of rat uterus. Conducive to the role of gallbladder, can enhance the secretion of bile in rats. It is sensitizing to humans. After inhalation of plant dust containing this product, asthma and dermatitis may occur. However, after ingestion, it can become non-sensitizing substances through the action of small intestinal secretions. It can stop bleeding, increase white blood cells, and shorten blood coagulation and bleeding time. The toxicity of chlorogenic acid is very small, and the oral LD ₅₀ of young rats is greater than 1g/Kg and the intraperitoneal injection is greater than 0.25g/Kg.

Contents of the invention

The object of the present invention is to provide a new medical application of chlorogenic acid, especially having anti-SARS virus activity, which can be used as a medicine for preventing and treating SARS virus diseases.

Another object of the present invention is to prepare a suitable dosage form with chlorogenic acid having anti-SARS virus activity as a main component.

The chlorogenic acid of the present invention is decocted twice with water from the plant honeysuckle, the decoction is concentrated to a certain volume under reduced pressure, the pH of the dilute lime milk suspension is adjusted to alkaline, centrifuged, and the precipitate is ground and stirred with 2 times the amount of ethanol Add dilute sulfuric acid to make the pH to 3, centrifuge, adjust the pH of the supernatant with 2N NaOH, and recrystallize the precipitate from water to obtain pure chlorogenic acid.

The structure proves to be:

(Chlorogenic Acid Structure)

Chlorogenic acid has been proved by pharmacological tests that it has obvious anti-SARS virus activity. Vero-E6 cells are used as virus host cells (susceptible cells) for activity determination, and the detection index of the protective effect of chlorogenic acid on virus-infected cells is cell degeneration Response, and the rate of protection of infected cells.

The virus strain is BJ-01, and 7 dilutions of samples are used for screening, and the results prove that it has obvious anti-SARS virus activity.

According to the obvious anti-SARS virus activity of chlorogenic acid, different dosage forms can be made by adding adjuvants as needed.

The present invention contains an effective amount of chlorogenic acid and pharmaceutically acceptable excipients, carriers or diluents formulated into different pharmaceutical dosage forms, which can prepare chlorogenic acid and vitamin C with typical pharmaceutical carriers and excipients in oral preparations. Oral preparations, pharmaceutical carriers and excipients include lactose; starch and its derivatives such as corn starch, sodium carboxymethyl starch, etc.; cellulose derivatives, such as methyl and ethyl cellulose; gelatin; inorganic compounds such as light magnesium oxide, talc, etc.; oils such as vegetable oil, sesame oil, etc.; inclusion agents such as β-cyclodextrin; and glycols such as polyethylene glycol. Oral preparations can be made into tablets, capsules, emulsions, solutions, etc., and can also be applied to solid dispersion tablets, liposome preparations, targeted preparations, and controlled release preparations.

The present invention contains an effective amount of chlorogenic acid and pharmaceutically acceptable excipients or diluents to prepare injections including isotonic saline, 5% glucose aqueous solution, water for injection, tea oil, soybean oil, sesame oil, ethanol, glycerin, propylene glycol, poly Ethylene glycol, benzyl benzoate; Tween 80, lecithin, sodium deoxycholate, egg yolk phospholipids, polyvinylpyrrolidone; acetic acid, sodium acetate, citric acid and its sodium salt, lactic acid, tartaric acid and its sodium salt, dihydrogen phosphate Sodium, sodium monobasic phosphate, sodium bicarbonate, and sodium carbonate; gelatin, sodium carboxymethylcellulose, pectin, sorbitol solution; creatinine, glycine; sodium sulfite, sodium bisulfite, sodium pyrophosphite, sodium thiosulfate , benzyl alcohol, butyl p-hydroxybenzoate and ethyl phenol, chlorobutanol; lidocaine; cephalin, soybean lecithin, etc. Typical forms of injection preparations are powder injections, suspension injections, etc.

In tablets, chlorogenic acid can be mixed with a disintegrant composed of sodium bicarbonate and tartaric acid or citric acid to make an effervescent tablet. When it meets water, carbon dioxide gas is generated to make the tablet disintegrate rapidly.

Chlorogenic acid can also be made into aerosols, for example, polyvinyl chloride paste resins such as dibutyl phthalate, dioctyl phthalate, calcium stearate, zinc stearate, etc. are used to prepare aerosols.

Detailed ways

Decoct honeysuckle with water twice, concentrate the decoction to 1:2 under reduced pressure, add 20% lime milk suspension to adjust the pH to about 10, centrifuge the precipitate, grind the precipitate with 2 times the amount of ethanol, and add dropwise under stirring 50% sulfuric acid, adjust the solution to pH3, stir well, and centrifuge to remove solids. The filtrate was adjusted to pH 6 with 2N NaOH, the ethanol was recovered, and dried under reduced pressure to obtain a crude extract. The crude extract was dissolved in a small amount of water, separated by polyamide column chromatography, eluted with gradient ethanol-water, the eluate was detected by polyamide thin layer, the fraction containing chlorogenic acid was combined, and dried under reduced pressure. Recrystallize with water to obtain pure chlorogenic acid.

                    Anti-SARS virus activity results of chlorogenic acid

Test item: Screening of chlorogenic acid anti-SARS virus activity (virus-cell level model)

Test principle: Vero-E6 cells are used as virus host cells (susceptible cells), and the protective effect of samples on virus-infected cells is tested. The detection indicators are cell degeneration (CPE) and the protection rate of infected cells.

Test materials and methods: Chlorogenic acid

Virus strain: BJ-01

Sample treatment: Dissolve the sample in culture medium or DMSO, make an appropriate initial concentration, 5-fold dilution, 7 dilutions each.

Test method: Inoculate Vero-E6 cells in a 96-well culture plate, culture in a 37°C, 5% CO2 incubator, add SARS virus and samples of different dilution concentrations, and set virus control, cell control and sample control. Observe the results under the microscope every day, record the CPE, and measure the OD value with neutral red staining, and calculate and evaluate the anti-SARS virus activity of the sample with reference to the control.

Test Results

Concentration (ug/ml) CPE Protection rate of infected cells

100 - 3.01

20 - 2.54

4 - 2.65

0.8 - 2.77

0.16 - 2.38

0.032 - 2.33

0.0064 + 1.21

Cytopathic pathology (CPE): The degree of cytopathic pathology is indicated by a plus sign, <25%+, 25%-50%++, 50%-75+++, >75%++++.

Infected cell protection rate: By comparing the OD values of the virus control, cell control and sample control, the protective activity of the sample on the virus-infected cells was calculated. The protection rate was greater than 1.5 times. It was preliminarily considered that the sample had a certain protective activity on the virus-infected cells.

*Sample cytotoxicity: if the ratio of the cytotoxicity of the sample to the cell control is >50%, the CPE evaluation and protection rate calculation will not be performed.

Above-mentioned result shows that this compound has obvious anti-SARS virus active effect. The LD ₅₀ of chlorogenic acid is greater than 1g/Kg, and the intraperitoneal injection greater than 0.25g/Kg proves that the toxicity is low.

Embodiment one

Select the following dosage of chlorogenic acid and additives: (compound: vitamin = 1: 1w/w)

Chlorogenic Acid 400g

Vitamin C 400g

Hydroxypropyl vitamin 20 grams

Micropowder silica gel 60g

120g pregelatinized starch

Magnesium Stearate Appropriate amount

Made into 1000 capsules

Mix chlorogenic acid and additives evenly, use 10% pregelatinized starch slurry as a binder, wet granulate, dry, add appropriate amount of magnesium stearate, mix, and press into tablets.

Embodiment two

Select the following dosage of chlorogenic acid and additives: (compound: vitamin=1:9)

Chlorogenic acid 80g

Vitamin C 720g

Hydroxypropyl vitamin 20 grams

Micronized silica gel 48g

Alcohol Appropriate amount

Magnesium Stearate Appropriate amount

Made into 900 capsules

Mix chlorogenic acid with hydroxypropyl cellulose and micropowder silica gel, use an appropriate amount of 70% ethanol as a wetting agent, wet granulate, sieve through 40 meshes to form granules, dry, add magnesium stearate, mix, and pack into hard capsules.

Embodiment three

Select the following dosages of chlorogenic acid and additives: (compound: vitamin = 2: 1)

Chlorogenic acid 800g

Vitamin C 400g

sucrose 40 grams

Preservatives Appropriate amount

Appropriate amount of distilled water

Made 20000ml

Take an appropriate amount of distilled water, add chlorogenic acid, stir while heating to dissolve, and filter. In addition, sucrose and preservatives are heated and dissolved with distilled water, slowly added to the above solution under stirring, added distilled water to the full amount, mixed, refrigerated, filtered, potted and sterilized to obtain an oral liquid.

Embodiment four

Select the following dosage of chlorogenic acid and additives: (compound: vitamin = 6: 1)

Chlorogenic acid 2400g

Vitamin C 400g

Tween 80 Appropriate amount

Water for Injection Appropriate amount

Made 45000ml

Take an appropriate amount of water for injection, add chlorogenic acid, pharmaceutical excipients and excipients, heat, stir to fully dissolve, add 4% Tween 80, stir well, filter, add hydrochloric acid to adjust the pH to 6.8, add water for injection to a sufficient volume, hollow fiber ultrafiltration, fine filtration, potting, and sterilization to obtain an injection.

Claims (6)

1. A medical application of chlorogenic acid with the following structure, characterized in that it is used in the preparation of medicines for preventing and treating diseases caused by SARS virus

2. The medical use of chlorogenic acid according to claim 1, characterized in that chlorogenic acid is formulated with pharmaceutical excipients, carriers or diluents. 3. The medical use of chlorogenic acid according to claim 2, characterized in that the oral excipients, carriers or diluents are made into different pharmaceutical dosage forms, and chlorogenic acid, vitamin C and typical oral preparations Drug carriers and excipients are made into oral preparations, and drug carriers and excipients include lactose; starch and its derivatives include corn starch and sodium carboxymethyl starch; cellulose derivatives include methyl and ethyl cellulose, gelatin ; Inorganic compounds include light magnesium oxide and talcum powder; oils include vegetable oil and sesame oil; inclusion agents and dibasic alcohol oral preparations can be made into tablets, capsules, emulsions, solutions, and can also be used in solid dispersible tablets, lipids, etc. Plastid formulations, targeted formulations and controlled release formulations. 4. The medical use of chlorogenic acid according to claim 2, characterized in that the excipient carrier or diluent made into injections is prepared into injections including isotonic saline, 5% glucose aqueous solution, water for injection, tea oil, soybean oil , sesame oil, ethanol, glycerin, propylene glycol, polyethylene glycol, benzyl benzoate; Tween 80, lecithin, sodium deoxycholate, egg yolk phospholipids, polyvinylpyrrolidone; acetic acid, sodium acetate, citric acid and sodium salt, Lactic acid, tartaric acid and its sodium salts, disodium hydrogen phosphate, sodium dihydrogen phosphate, sodium bicarbonate and sodium carbonate; gelatin, sodium carboxymethylcellulose, pectin, sorbitol solution; creatinine, glycine; sodium sulfite, sodium bisulfite , sodium pyrophosphite, sodium thiosulfate, benzyl alcohol, butyl p-hydroxybenzoate and ethyl phenol, chlorobutanol; lidocaine; cephalin, soybean lecithin Suspension injection. 5. The medical use of chlorogenic acid according to claim 2, characterized in that the oral tablets are made of sodium bicarbonate tartaric acid and citric acid to make effervescent tablets. 6. The medical use of chlorogenic acid according to claim 2, characterized in that the formula uses polyvinyl chloride paste resin, dibutyl phthalate, dioctyl phthalate, calcium stearate, hard Zinc fatty acid is formulated as an aerosol.