CN101130498B - Method for extracting and purifying salvianolic acid A - Google Patents

Abstract

The invention provides a method for extracting and purifying salvianolic acid A, which uses danshen decoction pieces or powder as raw materials, uses solvent extraction and concentration to obtain salvia miltiorrhiza concentrate, and obtains high-purity salvianolic acid A through protein adsorption and solvent extraction. The present invention utilizes gelatin or egg white protein to precipitate salvianolic acid A, dissolves and analyzes in acetone aqueous solution, and then extracts with an organic solvent to obtain high-purity salvianolic acid A. The final extract yield of the method of the invention is over 0.3%, the purity of the salvianolic acid A is over 90%, and the column chromatography step in the conventional method is avoided. The method of the invention has the advantages of reasonable design, high extraction rate, good purity, stable performance, simple process and low production cost, and is suitable for large-scale industrial production.

Description

A kind of extraction and purification method of salvianolic acid A

technical field

The invention belongs to compound preparation, and mainly relates to a method for extracting and purifying salvianolic acid A.

Background technique

Salvia miltiorrhiza is one of the most commonly used Chinese medicinal herbs. For the treatment of cardiovascular and cerebrovascular diseases such as angina pectoris, hypertension, coronary heart disease and stroke. There are many traditional Chinese medicine preparations that use Danshen decoction pieces or crude extracts of Danshen as medicine, such as compound Danshen tablets, compound Danshen dripping pills, Danshen injection, Xiangdan injection, etc. In addition, there are also pure compounds or their salts prepared from Danshen Prepared preparations, such as Danshensu Sodium Injection, Salvianolic Acid B Injection, etc.

In recent years, with the deepening of pharmacological research, various pharmacological activities of salvianolic acid A have been discovered. For example, Zhen Yongsu et al. found that salvianolic acid A has anti-tumor activity; Du Guanhua et al. found that salvianolic acid A has obvious protective effect on isolated myocardial ischemia-reperfusion injury in rats, and has a protective effect on brain injury caused by cerebral ischemia-reperfusion in mice. It has a protective effect on the memory acquisition disorder caused by anisodine or hyoscine in mice, etc.; Tao Wen (Tao Wen) found that salvianolic acid A has a protective effect on rat liver damage caused by carbon tetrachloride . Li Zhigang and others have also developed salvianolic acid A tablets and injections for the treatment of cardiovascular diseases.

Chinese patent CN1397276A disclosed in 2002 a method for preparing salvianolic acid A through water or alcohol extraction, resin chromatography, and purification by silica gel column chromatography. Chinese patent CN1513848A disclosed in 2003 a method for preparing salvianolic acid A by soaking in water, centrifuging, ultrafiltration, adjusting acid, extracting with organic solvent, and then reversed-phase column chromatography. Chinese patent CN1830947A and Chinese patent CN1887849A disclosed in 2006 and 2007, respectively, the use of water or alcohol to extract the medicinal material of Salvia miltiorrhiza, followed by resin or gel or reversed-phase column chromatography, and finally acid adjustment and organic solvent extraction to prepare salvianolic acid A Methods.

Since the content of salvianolic acid A in medicinal materials is very low, the production cost is high. In addition, the application of column chromatography in large-scale industrial production has certain limitations, and cannot well meet the needs of industrial production of salvianolic acid A preparations.

Contents of the invention

Aiming at the deficiencies in the prior art, the present invention provides a method for extracting and purifying salvianolic acid A. The technical means used in the present invention are: taking Danshen decoction pieces or powder as raw materials, extracting and concentrating with solvents to obtain the concentrated liquid of Salvia miltiorrhiza. Protein adsorption and solvent extraction methods to obtain high-purity salvianolic acid A, specifically through the following steps:

1. Obtain Danshen Concentrate: Use conventional solvent extraction and concentration methods to obtain Salvia Concentrate;

The Danshen decoction pieces or powder are extracted with water or different proportions of ethanol aqueous solution through ultrasonication, warm soaking or heating under reflux, and the extract is naturally settled, filtered or centrifuged to remove insoluble impurities. Concentrate under normal pressure or reduced pressure, naturally settle, filter or centrifuge to remove insoluble impurities, and make Danshen concentrate;

2. Purification of salvianolic acid A: use protein to adsorb salvianolic acid A in the concentrated solution of salvia miltiorrhiza, then analyze with acetone and extract with organic solvent to separate the organic solvent layer containing salvianolic acid A;

3. Concentrate and dry salvianolic acid A to obtain purified salvianolic acid A: Concentrate under reduced pressure or normal pressure, and dry by vacuum drying, spray drying or freeze-drying to obtain purified salvianolic acid A.

In step 1, Danshen herb powder is preferred for warm extraction or ultrasonic extraction, and Danshen decoction slices are preferred for heating and reflux extraction; the solvent for extraction is 0-50% ethanol; it is advisable to concentrate the Danshen extract to 1 mL, which is equivalent to 0.5g-2g of medicinal materials .

In step 2, the protein used for adsorption is gelatin or egg white, and the gelatin or egg white is dissolved in water to make a certain concentration of protein aqueous solution; then the gelatin or egg white protein aqueous solution is added to the Danshen concentrated solution until no precipitation occurs. Natural sedimentation, centrifugation or filtration to obtain salvianolic acid A and protein precipitation complex; dissolve the complex with acetone aqueous solution, natural sedimentation, centrifugation or filtration to obtain supernatant, or directly heat to dissolve the precipitate, and concentrate to Acetone free;

In step 2, when the selected protein is gelatin, its aqueous solution concentration is 1%-20%, preferably 5% gelatin aqueous solution; when the selected protein is egg white, its aqueous solution concentration is 1%-10%.

In step 2, the rotational speed of the aqueous protein solution of gelatin or egg white is more than 3000 rpm during centrifugation.

In step 2, the acetone aqueous solution used for the compound of salvianolic acid A and protein precipitation is 10% to 90% acetone aqueous solution, preferably 50% acetone aqueous solution, and the rotation speed during centrifugation is above 8000 rpm, and when directly heating to dissolve the precipitate, the temperature is 40 to 100 ℃.

The organic solvent used in step 2 is acetate solvents such as ethyl acetate, propyl acetate, butyl acetate.

According to the extraction and purification method of salvianolic acid A of the present invention, salvianolic acid A has the property of being able to combine with protein to form a precipitate, after using gelatin or egg white protein to precipitate salvianolic acid A, the precipitate is then dissolved in acetone aqueous solution, and the salvianolic acid A is resolved from protein precipitation, and high-purity salvianolic acid A is obtained through organic solvent extraction, so as to achieve the purpose of separation and purification of salvianolic acid A. The final extract yield of the method is over 0.3%, the purity of the salvianolic acid A is over 90%, and the column chromatography step in the previous method is avoided. The method of the invention has the advantages of reasonable design, high extraction rate, good purity, stable performance, simple process and low production cost, and is suitable for large-scale industrial production.

Description of drawings

Fig. 1 is the chromatogram of checking the purity of salvianolic acid A.

Detailed ways

The present invention will be further described in conjunction with drawings and embodiments.

Embodiment one

The extraction and purification method of salvianolic acid A, its processing steps are:

1. Take the medicinal material of Danshen, add 8 times the amount of water, decoct and reflux 3 times, 1 hour each time, filter, concentrate the filtrate under reduced pressure to 1ml equivalent to 2g of medicinal material, and filter;

2. Add 5% gelatin aqueous solution until no precipitation occurs, and centrifuge at 4000 rpm for 10 minutes; add acetone aqueous solution equivalent to 50% of the volume of the concentrated solution in step 2 to the precipitation, and heat at 60°C to dissolve the precipitation; after decompression recovery to remove acetone, the aqueous solution Extract 3 times with an equal volume of ethyl acetate, and combine the organic phases;

3. Concentrate the extract under reduced pressure to extract, and freeze-dry to obtain a light yellow powder, which is salvianolic acid A.

See Figure 1 for the chromatogram of salvianolic acid A purity check.

Embodiment two

It is basically the same as Example 1, except that the extraction solvent is 50% aqueous ethanol.

Embodiment three

It is basically the same as in Example 1, except that the concentrated solution in step 1 is concentrated to 1 ml, which is equivalent to 0.5 g of medicinal materials.

Embodiment Four

It is basically the same as Example 1, except that the gelatin aqueous solution used in step 2 has a concentration of 20%.

Embodiment five

It is basically the same as Example 1, except that the concentration of the acetone aqueous solution added in step 2 is 80% acetone aqueous solution.

Embodiment six

It is basically the same as Example 1, except that after adding acetone aqueous solution in step 2, it is stirred and centrifuged (10000rpm).

Embodiment seven

It is basically the same as Example 1, except that the extraction solvent in step 2 is butyl acetate.

Embodiment Eight

It is basically the same as in Example 1, except that the freeze-drying in step 3 is changed to vacuum drying at 50° C. for 10 h.

Embodiment nine

It is basically the same as Example 1, except that the protein solution used in step 2 is 1% egg white aqueous solution.

Claims (4)

1. the extracting and purifying method of a salvianolic acid A is a raw material with salvia piece or powder, adopts solvent extraction, concentrates and obtain red sage root liquid concentrator, obtains the salvianolic acid A of purifying through protein adsorption, solvent extration, it is characterized in that:

(1) adopt conventional solvent extraction, method of enrichment to obtain red sage root liquid concentrator;

(2) salvianolic acid A purifying:,, separate and contain the salvianolic acid A organic solvent layer again through acetone parsing, organic solvent extraction with the salvianolic acid A in the protein adsorption red sage root liquid concentrator;

The organic solvent layer that (3) will contain salvianolic acid A concentrates, and drying is obtained the salvianolic acid A of purifying;

Wherein: protein is selected gelatin or egg white for use, and the organic solvent of extraction is selected ETHYLE ACETATE, propyl acetate, butylacetate for use;

In the step (2), gelatin or egg white is soluble in water, process protein solution; Protein solution with gelatin or egg white joins in the red sage root liquid concentrator again, to no longer producing deposition, and natural subsidence, centrifugal or filter and obtain salvianolic acid A and protein precipitation mixture; Dissolve mixture with aqueous acetone solution; Natural subsidence, centrifugal or filter and to obtain supernatant, or direct heating dissolution precipitation are concentrated into no acetone under normal pressure or the decompression.

2. the extracting and purifying method of a kind of salvianolic acid A as claimed in claim 1 is characterized in that: in the said step (1), warm lixiviate is got or is selected the red rooted salvia powder for use during supersound extraction, selects salvia piece during heating and refluxing extraction for use; Extracting and using solvent is 0～50% ethanol; Radix Salviae Miltiorrhizae extract is concentrated into 1 milliliter and is equivalent to 0.5g～2g medicinal material.

3. the extracting and purifying method of a kind of salvianolic acid A as claimed in claim 1 is characterized in that: resolving used in the step (2) is 10%～90% aqueous acetone solution.

4. the extracting and purifying method of a kind of salvianolic acid A as claimed in claim 1 is characterized in that: concentrate described in the step (3) and adopt concentrating under reduced pressure or normal pressure to concentrate, dry employing vacuum-drying, spraying drying or lyophilize.