阅读说明：本技术 射干异黄酮苷元的制备方法 (Preparation method of blackberry lily isoflavone aglycone ) 是由 昝立峰 梁瑞娟 杨香瑜 于 2019-11-29 设计创作，主要内容包括：本发明涉及中药制备技术领域,具体公开一种射干异黄酮苷元的制备方法。所述射干异黄酮苷元的制备方法,是将射干须根的醇提液去醇处理后加入粗毛纤孔菌的发酵液中进行酶解,去除酶解液中的酶和菌体杂质,进行树脂吸附纯化,得到射干异黄酮苷元。本发明制备条件温和,方法简单,成本低,大大提高了从射干须根中提取和制备的射干异黄酮苷元的纯度和收率。(The invention relates to the technical field of traditional Chinese medicine preparation, and particularly discloses a preparation method of blackberry lily isoflavone aglycone. The preparation method of the blackberry lily isoflavone aglycone comprises the steps of carrying out alcohol removal treatment on an alcohol extract of blackberry lily fibrous roots, then adding the alcohol extract into a fermentation liquor of crude capillary fungus for enzymolysis, removing enzyme and thallus impurities in the enzymolysis liquor, and carrying out resin adsorption and purification to obtain the blackberry lily isoflavone aglycone. The method has the advantages of mild preparation conditions, simple method and low cost, and greatly improves the purity and yield of the blackberry lily isoflavone aglycone extracted and prepared from the blackberry lily fibrous root.)

1. A preparation method of blackberry lily isoflavone aglycone is characterized by comprising the following steps: removing alcohol from rhizoma Belamcandae fibrous root alcoholic extractive solution, adding into crude capillary fungus fermentation broth for enzymolysis, removing enzyme and thallus impurities in the enzymolysis solution, and performing resin adsorption and purification to obtain rhizoma Belamcandae isoflavone aglycone.

2. The method of claim 1, wherein: the alcoholic extract of the blackberry lily fibrous root is obtained by crushing the blackberry lily fibrous root, adding the crushed blackberry lily fibrous root into 60-80 vt% ethanol solution, and leaching at 50-70 ℃.

3. The method of claim 2, wherein: crushing the blackberry lily fibrous roots to 20-40 meshes of granularity; and/or

The leaching process is carried out by two times, in the first leaching process, the mass ratio of the blackberry lily fibrous root to the ethanol solution is 1:12-14, and the leaching time is 15-30min at the speed of 1000 r/min; in the second extraction process, the mass ratio of the blackberry lily fibrous root to the ethanol solution is 1:8-10, and the extraction is carried out for 15-30min at the speed of 1000r/min 800-.

4. The method of claim 1, wherein: the alcohol removing treatment process is to filter and evaporate and concentrate the alcohol extract until the alcohol content in the alcohol extract is less than or equal to 5vt percent.

5. The method of claim 1, wherein: the fermentation liquor of the crude capillary is homogenate enzyme liquid obtained by inoculating the crude capillary into a culture solution, culturing for 7-10 days at 28-30 ℃, and breaking the cell wall of the thallus in the culture solution; and/or

The adding volume of the fermentation liquor of the crude phellinus linteus is 5-10% of the volume of the concentrated solution after the alcohol removal treatment; and/or

The enzymolysis process comprises the following steps: performing enzymolysis at 40-55 deg.C and pH of 4-6 for 1-3 days.

6. The method of claim 5, wherein: the culture solution comprises 0.6-0.8% of malt extract, 0.07-0.08% of yeast extract, 0.8-1.2% of peptone and the balance of distilled water according to the mass ratio.

7. The method of claim 1, wherein: the process for removing enzyme and thallus impurities in the enzymolysis liquid comprises the following steps: and adding a flocculating agent into the enzymolysis liquid, precipitating for 2-3h, and carrying out centrifugal filtration to obtain a clear liquid.

8. The method of claim 7, wherein: the flocculant is one or a combination of chitosan, gelatin and A + B clarifying agent; the addition amount of the flocculating agent is 3-5% of the mass of the enzymolysis liquid.

9. The method of claim 1, wherein: the resin is macroporous adsorption resin.

10. The method of claim 9, wherein: the resin adsorption purification process comprises the following steps: diluting the enzymolysis solution by 1-1.5 times, adding into macroporous adsorption resin, controlling the adsorption flow rate at 1-1.5BV/h, eluting with 20-40 vt% ethanol solution to remove impurities, eluting with 60-80 vt% ethanol solution, concentrating and drying.

Technical Field

The invention relates to the technical field of traditional Chinese medicine preparation, in particular to a preparation method of blackberry lily isoflavone aglycone.

Background

Belamcanda chinensis is the dry rhizome of Iridaceae plant, is bitter in taste and cold in nature, enters liver and lung channels, has the effects of clearing heat and removing toxicity, relieving sore throat and eliminating phlegm, is mainly used for treating throat lung carbuncle, phlegm cough and asthma and the like, and is the essential medicine for treating pharyngitis and sore throat.

The research shows that the tectorigenin and other aglycone substances can obviously inhibit the transcription and translation levels of tumor immunocytes TNF- α, IL-1 β and IL-6, the tectorigenin and other aglycone substances can also prevent the accumulation of α -synuclein, slow down the degenerative change of dopaminergic neurons caused by hydroxypolyamide, enhance the food sensitivity and prolong the service life, the compound also has the treatment effect on potential gold-induced diseases, the activity of the isoflavone substances such as the tectorigenin, the wild tectorigenin and other isoflavone substances and the like does not waste a large amount of isoflavone resources in extraction, and the extraction cost is low, so that the extraction cost is low, and the extraction cost is low.

Disclosure of Invention

Aiming at the problems of low extraction rate and low product purity of the existing method for separating and extracting isoflavone aglycone from the fibrous root of the blackberry lily medicinal material, the invention provides a preparation method of the blackberry lily isoflavone aglycone.

In order to achieve the purpose of the invention, the embodiment of the invention adopts the following technical scheme:

a method for preparing rhizoma Belamcandae isoflavone aglycone comprises removing alcohol from rhizoma Belamcandae fibrous root alcoholic extractive solution, adding into crude Fibrouromycetes fermentation broth for enzymolysis, removing enzyme and thallus impurities in the enzymolysis solution, and performing resin adsorption and purification to obtain rhizoma Belamcandae isoflavone aglycone.

Wherein, the crude capillary is medicinal fungus crude capillary [ Inonotus hispidus (Bull.: Fr.) P.Karst. ], the name Xanthochrous fuscus (Xanthochrous hispidus) is superior to Basidiomycota (Basidiomycota), Agaricaceae (Agaricamycetes), Hymenochaetales (Hymenochaetales), Inonochaetaceae (Hymenochaetaceae), and Microchaetaceae (Inonotus); the strain is mainly used for treating various cancers, arthritis, gout, diabetes and other diseases and various stomach diseases caused by dyspepsia and the like, and the crude capillary fungus mainly contains polyphenol, triterpene, sterol and derivatives thereof, nucleoside, polysaccharide and other active ingredients, wherein polyphenol compounds Hispidin and Hispolon have various pharmacological actions of resisting virus, resisting tumor, resisting oxidation and the like.

Compared with the prior art, in the preparation method of blackberry lily isoflavone aglycone, isoflavone glycoside substances extracted from blackberry lily fibrous root alcohol extract are hydrolyzed by using various degrading enzymes generated in the fermentation process of crude capillary, so that various isoflavone glycoside substances (mangiferin, tectoridin, wild tectoridin and the like) which are difficult to degrade and have low biological activity in the alcohol extract can be fully hydrolyzed and converted to form isoflavone aglycone, the yield of the isoflavone aglycone extracted and prepared from blackberry lily fibrous root is greatly increased, the alcohol extract is subjected to resin adsorption and purification after enzymolysis, high-purity isoflavone aglycone can be obtained, the preparation condition is mild, the preparation method is simple, the cost is low, and the method is suitable for industrial production.

Preferably, the blackberry lily fibrous root ethanol extract is obtained by crushing blackberry lily fibrous roots, adding the crushed blackberry lily fibrous roots into 60-80 vt% ethanol solution, and leaching at 50-70 ℃.

Preferably, the blackberry lily roots are crushed to the granularity of 20-40 meshes.

Preferably, the leaching process is carried out by two times, in the first leaching process, the mass ratio of the blackberry lily fibrous root to the ethanol solution is 1:12-14, and the leaching time is 15-30min at 1000r/min of 800-; in the second leaching process, the mass ratio of the blackberry lily fibrous root to the ethanol solution is 1:8-10, and the leaching is carried out for 15-30min at the speed of 1000r/min of 800-; and combining the two leaching solutions to obtain the alcoholic leaching solution of the blackberry lily fibrous roots.

The above alcohol extraction process can extract isoflavone glycoside contained in rhizoma Belamcandae.

Preferably, the dealcoholization treatment process is to filter the alcoholic extract of the dried fibrous roots and concentrate the alcoholic extract by a thin film evaporator under reduced pressure until the alcohol content is less than or equal to 5vt percent.

The extraction liquid is subjected to alcohol removal treatment, so that the influence of alcohol on the hydrolysis efficiency of enzyme in the subsequent enzymolysis process can be avoided.

Preferably, the fermentation liquid of the crude capillary is a homogenate enzyme liquid obtained by inoculating the crude capillary into a culture solution, culturing for 7-10 days at 28-30 ℃, and breaking the cell wall of the culture solution by using a homogenizer.

The wall breaking treatment can fully release intracellular enzymes in the thalli, increase the concentration and the variety of the enzymes and improve the enzymolysis efficiency.

Preferably, the addition volume of the fermentation liquid of the crude phellinus linteus is 5 to 10% of the volume of the concentrated liquid after the dealcoholization treatment.

Preferably, the enzymolysis process is as follows: performing enzymolysis at 40-55 deg.C and pH of 4-6 for 1-3 days.

Under the enzymolysis condition, the hydrolysis efficiency of the enzyme in the enzymolysis liquid to the isoflavone glycoside substances can be improved.

Preferably, the culture solution comprises 0.6-0.8% of malt extract, 0.07-0.08% of yeast extract, 0.8-1.2% of peptone and the balance of distilled water by mass ratio.

Preferably, the process for removing the enzyme and the bacterial impurities in the enzymatic hydrolysate comprises the following steps: and adding a flocculating agent into the enzymolysis liquid, precipitating for 2-3h, and carrying out centrifugal filtration to obtain a clear liquid.

The impurity removal method can fully remove enzyme, other macromolecular protein and thallus fragments in the enzymolysis liquid, and ensures the smooth proceeding of the subsequent resin adsorption treatment.

Preferably, the flocculant is one or more of chitosan, gelatin and A + B clarifying agent; the addition amount of the flocculating agent is 3-5% of the mass of the enzymolysis liquid.

Preferably, the resin is macroporous adsorption resin, and specifically one of HPD700, LX-17 and LK-17 macroporous resin can be selected.

Preferably, the resin adsorption purification process is as follows: diluting the enzymolysis solution by 1-1.5 times, adding into macroporous adsorption resin, controlling the adsorption flow rate at 1-1.5BV/h, eluting with 20-40 vt% ethanol solution to remove impurities, eluting with 60-80 vt% ethanol solution, concentrating and drying.

Detailed Description

In order to make the objects, technical solutions and advantages of the present invention more apparent, the present invention is further described in detail with reference to the following embodiments. It should be understood that the specific embodiments described herein are merely illustrative of the invention and are not intended to limit the invention.

The following examples are provided to better illustrate the embodiments of the present invention.