CN102093433A - Method for separating out traditional Chinese medicine natural base by silicagel column chromatography - Google Patents

Abstract

The invention discloses a method for separating out traditional Chinese medicine natural base by silicagel column chromatography. The specific process steps are as follows: pre-treating raw materials, dissolving, extracting with chloroform, carrying out chromatography and drying. In the method, complicate and time-consuming instruments and equipment, such as a Soxhlet extraction device and the like, are prevented from being used; the method disclosed by the invention is high in efficiency, is simple and the like; and the silicagel column chromatography has the advantages of low cost and high repeatability, is convenient for operation, and the like.

Description

The alkaloidal method of silica gel column chromatography separating traditional Chinese medicine

Technical field:

The present invention relates to a kind of alkaloidal method of from Chinese medicine, extracting, especially relate to a kind of silica gel column chromatography that utilizes and from Chinese medicine, extract alkaloidal method.

Background technology:

Alkaloid is extensively to be present in the nitrogenous organic compound of a natural class, and great majority have complicated nitrogen heterocyclic structure, is alkalescence.Alkaloidal discovery started from for 19 beginnings of the century, be people study at most and also the natural organic compound of a class the earliest.Kind surplus the alkaloid of having found at present about 6000, and still with the speed increase of about kind more than 100 in every year.Because that most of alkaloids all have is anticancer, antitumor, the effect of anti-inflammatory, antiviral and cardiovascular disordeies such as anti-platelet aggregation, anti-arrhythmia and hypertension, therefore, it has huge application potential at aspects such as health, medicine.Because most of alkaloidal pharmacological actions still remain further research, this just need extraction from contain alkaloidal medicinal plant in a large number, separation, purifying biological alkali.How to use modern times separation means efficiently, easy and separate efficiently, be an importance of current alkaloid research.

According to alkaloidal basic structure, tens types can be divided into.Most of alkaloids are almost insoluble or be insoluble in water, can be dissolved in organic solvents such as chloroform, ether, alcohol, acetone, benzene, but behind alkaloid and the diluted acid effect generation salt, in the water soluble solution.Most alkaloids equal non-volatility when normal pressure, general aspects is more stable, and the separation and purification process does not need particular surroundings, except that lucifuge, does not need other special conditionss on storing.Being rich in alkaloid in many medicinal plants, is the integral part of effective components of Chinese medicinal.The isolation and purification of alkaloid effective constituent is the difficult point and key of tcm development, and Research on New and exploitation can improve the yield and the quality of chemical composition of Chinese materia medica greatly, save a large amount of time and the energy.

Summary of the invention:

The problem that prior art is higher to alkaloid extraction cost in the Chinese medicine in order to solve in the present invention, complicated operation, purification rate are low, be unfavorable for suitability for industrialized production and provide a kind of silica gel column chromatography separating traditional Chinese medicine alkaloidal method.

In order to solve above technical problem, technical scheme of the present invention is: the alkaloidal method of a kind of silica gel column chromatography separating traditional Chinese medicine, and concrete processing step is as follows:

(1). raw materials pretreatment

Chinese medicinal materials is cleaned, 60 ℃ down baking 8h pulverize the back with pulverizer and cross 60 mesh sieves to constant weight, herb powder is standby;

(2). dissolving

Adding concentration according to HCl and 1: 10 ratio of water is 0.04% HCl, fully stirs, and microwave oven was handled 2 minutes, with the centrifugal 20min of whizzer 6000r/min, got supernatant liquor; After getting supernatant liquor precipitation is repeated once according to 1 and 2 above-mentioned steps again; Merging supernatant liquor twice, is 10 with 10% ammoniacal liquor adjust pH;

(3). chloroform extraction

The chloroform extraction that adds supernatant liquor 1/5 volume, same chloroform extraction 3 times, the combined chloroform layer utilizes the Rotary Evaporators evaporated under reduced pressure;

(4). chromatography

According to the volume of chromatography column, to chromatography column 3/4 place, cross post with the chloroform of 3 times of chromatography column volumes with 300-400 order silica gel dry-packing; Liquid to be purified is added to the top of chromatography column, and add-on is no more than 1/10 of column volume, opens the post lower end, and liquid is immersed in the silica gel; With methyl alcohol and chloroform proportioning is 1: 1 eluent wash-out, collects since second the effusive liquid of cylinder, collects till the 6th the cylinder effluent liquid always;

(5). drying

To collect liquid Rotary Evaporators evaporated under reduced pressure.

Features such as the present invention has avoided using the plant and instrument of complicated and time consumption such as Soxhlet extraction element, has the efficient height, and is comparatively easy, advantage such as it is with low cost that silica gel column chromatography has, and repeatability is high, and is easy to operate.

Embodiment:

Concrete processing step of the present invention is as follows:

1. raw materials pretreatment

Chinese medicinal materials (as lotus leaf, the coptis, Yanhusuo, the bark of eucommia etc., extracting respectively according to different needs) is cleaned, and 60 ℃ are dried by the fire 8h down to constant weight (constant weight: the weight that prolongs medicinal material time of drying does not change).Pulverize the back with pulverizer and cross 60 mesh sieves, it is standby to get herb powder.

2. dissolving

(HCl: it was 0.04% HCl that ratio water=1: 10) adds concentration, fully stirs according to HCl and water 1: 10; WD900B type microwave oven was handled 2 minutes, with the centrifugal 20min of whizzer 6000r/min, got supernatant liquor; After getting supernatant liquor precipitation is repeated once according to 1 and 2 above-mentioned steps again.Merging twice supernatant liquor (2 centrifugal supernatant liquors are merged), is 10 with 10% ammoniacal liquor adjust pH.

3, chloroform extraction

The chloroform extraction (chloroform extraction is exactly a kind of extracting process) that adds 1/5 volume (supernatant liquor after merging in the step 2 1/5), same chloroform extraction 3 times, the combined chloroform layer utilizes the Rotary Evaporators evaporated under reduced pressure.The anhydrous alcohol solution precipitation is treated column chromatography purification.

4, chromatography

(chromatography column is the main body in the chromatographic technique according to chromatography column, generally be Glass tubing or plexi-glass tubular) volume, (dry-packing: elder generation is filled to chromatography column 3/8 place with 300-400 order silica gel to chromatography column 3/4 place with 300-400 order silica gel dry-packing, then following piston is communicated with pump, make silica gel dress real by decompression, add the resolution of precipitate liquid in the step 3 of 3/8 volume again, soak into silica gel, carefully make it not produce bubble), cross post with the chloroform of 3 times of chromatography column volumes.Liquid to be purified is added to the top of chromatography column, and add-on is no more than 1/10 of column volume, opens the post lower end, and liquid is immersed in the silica gel; It with methyl alcohol and chloroform proportioning 1: 1 eluent wash-out, collect since second the effusive liquid of cylinder, till collecting the 6th cylinder effluent liquid always (with methyl alcohol and chloroform proportioning be 1: 1 be eluent, the flow velocity of 70ml/min carries out gradient elution, the applying pressure of post is 2-3kg/c, collect since second column volume effluent liquid, collect till the 6th the cylinder hydrops always.)。

5, drying

To collect liquid Rotary Evaporators evaporated under reduced pressure.

Microwave helps alkaloidal separating out, and the alkaloid crude product of separating out can be got to about 22%-23%.

Acidic solution helps alkaloid and other separating substances, and the refined product alkaloid that obtains after the separation reaches about 94%-95%.Identifying with high performance liquid chromatography, is example with the bark of eucommia: total biology total alkali separates each component and analyzes with HPLC in the bark of eucommia with high speed adverse current chromatogram.WatersC ₁₈Chromatographic column (260mmx4.6mm, 6um); The detection wavelength is 280nm; Moving phase is ethanol one acetate/sodium acetate buffer solution (60: 40, V/V, pH 5.0), flow velocity 1.0ml/min, sample size 10uL.The structure of each peak component of HSCCC is analyzed according to the data of ESI-MS-MS, 1H-NMR and 13C-NMR, draws total alkaloid content at 94-95%.

Claims (1)

1. alkaloidal method of silica gel column chromatography separating traditional Chinese medicine is characterized in that: concrete processing step is as follows:

(1). raw materials pretreatment

Chinese medicinal materials is cleaned, 60 ℃ down baking 8h pulverize the back with pulverizer and cross 60 mesh sieves to constant weight, herb powder is standby;

(2). dissolving

Adding concentration according to HCl and 1: 10 ratio of water is 0.04% HCl, fully stirs, and microwave oven was handled 2 minutes, with the centrifugal 20min of whizzer 6000r/min, got supernatant liquor; After getting supernatant liquor precipitation is repeated once according to 1 and 2 above-mentioned steps again; Merging supernatant liquor twice, is 10 with 10% ammoniacal liquor adjust pH;

(3). chloroform extraction

The chloroform extraction that adds supernatant liquor 1/5 volume, same chloroform extraction 3 times, the combined chloroform layer utilizes the Rotary Evaporators evaporated under reduced pressure;

(4). chromatography

According to the volume of chromatography column, to chromatography column 3/4 place, cross post with the chloroform of 3 times of chromatography column volumes with 300-400 order silica gel dry-packing; Liquid to be purified is added to the top of chromatography column, and add-on is no more than 1/10 of column volume, opens the post lower end, and liquid is immersed in the silica gel; With methyl alcohol and chloroform proportioning is 1: 1 eluent wash-out, collects since second the effusive liquid of cylinder, collects till the 6th the cylinder effluent liquid always;

(5). drying

To collect liquid Rotary Evaporators evaporated under reduced pressure.