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Preparation method of antitussive, expectorant, anti-inflammatory, and antimicrobial extract from earthworm

An extract and earthworm technology, which is applied in the field of earthworm extract and its preparation, can solve the problems of insignificant separation effect, long and complicated process, time-consuming and labor-consuming earthworm extract, etc.

Active Publication Date: 2019-01-25
首都儿科研究所
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

The applicant has carried out a research report on the antitussive, expectorant, antimicrobial, anti-inflammatory effects of the earthworm extract and its preparation method. In the previous report (see literature Zhu Z, Yang Q, Yu B, et al. Purification and characterization of a novel earthworm DNase.PrepBiochem Biotechnol.2015; 45(4):365-79.), the preparation method of earthworm extract is time-consuming and labor-intensive, the process is lengthy and complicated, and the obtained product still contains a large amount of impurities. not obvious

Method used

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  • Preparation method of antitussive, expectorant, anti-inflammatory, and antimicrobial extract from earthworm
  • Preparation method of antitussive, expectorant, anti-inflammatory, and antimicrobial extract from earthworm
  • Preparation method of antitussive, expectorant, anti-inflammatory, and antimicrobial extract from earthworm

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Effect test

Embodiment 1

[0096] Embodiment 1, the preparation method of earthworm biological expectorant extract

[0097] (1) Preparation process of crude extract

[0098] The specific preparation process of the earthworm crude extract provided in this embodiment is as follows: figure 2 Shown, wherein, the concentration percent of described acetone is volume percent content.

[0099] (2) Purification process of crude extract

[0100] The crude extract prepared in the above step (1) is subjected to ion-exchange chromatography and affinity chromatography to obtain earthworm extract, which is one of the raw materials for preparing cough-relieving, expectorant, antimicrobial and anti-inflammatory drugs

[0101] 1 Prepare the sample: the crude extract is fully dissolved in NaAC buffer solution (A liquid: 0.02M NaAC+0.02M NaCl) with pH 4.8-5.2, and then centrifuged at 15000g for 10 minutes. The precipitate was discarded, and the supernatant was filtered through a 22-micron filter membrane and used for c...

Embodiment 2

[0108] Embodiment 2, the product yield of the extract that the present invention prepares and the removal rate of impurity

[0109] The expectorant active ingredient of earthworm extract is calculated by deoxyribonuclease activity: definition of activity unit: under the conditions of 25°C and pH 5.2, the amount of enzyme required to cause the 260nm light absorption value of the substrate to increase by 0.001 is one unit.

[0110] The product yield calculation method or formula is: Yield = total activity of enzyme after purification ÷ total activity of enzyme before purification

[0111] Protein concentration measurement method: Bradford method

[0112] The calculation method of purification multiple is: purification multiple = specific activity after purification ÷ specific activity of homogenate

[0113] Calculated by the above method or formula:

[0114] The purification factor of the earthworm antitussive, expectorant, antimicrobial and anti-inflammation extract prepared ...

Embodiment 3

[0122] Embodiment 3, the deoxyribonuclease activity assay experiment of earthworm extract

[0123] The method for measuring deoxyribonuclease activity conventional in the art (see literature Zhu Z, Yang Q, YuB, et al. Purification and characterization of a novel earthworm DNase. PrepBiochem Biotechnol. 2015; 45 (4): 365-79. ), measure the lysozyme activity of earthworm homogenate, crude extract, and earthworm extract respectively, with the existing general expectorant preparation pancreas deoxyribonuclease as contrast, the bovine pancreatic DNase I of 1mg / mL and the earthworm crude extract Dilute, use the plasmid as a substrate, and detect its enzyme activity, the results are as follows figure 1 As shown, after diluting pancreatic deoxyribonuclease I by 100 times, there is almost no degradation effect, while the earthworm crude extract diluted to 500 times can still completely degrade the plasmid DNA.

[0124] The effective deoxyribonuclease concentration of the crude extract p...

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Abstract

The invention provides a preparation method of an antitussive, expectorant, anti-inflammatory and anti-microbial extract from earthworms. The extraction method provided by the invention mainly extracts the earthwormhomogenate through NaAC buffer and acetone of pH 4.8-5.4. The invention optimizes the extraction process provided by the applicant through repeated experiments, greatly shortens the preparation time of earthworm extract, and reduces the preparation time of earthworm extract from 42to 45 hours to 15 to18 hours; the yield was increased from 73% to 82%, and the recovery of total activity was increased from 1141 to 4443, and the specific activity was increased by 3.9 times. The purification times increased from 41 times to 158 times, and the purification times of the active component of deoxyribonuclease increased nearly fourfold. The preparation method provided by the invention is simple in process, low in cost and easy to be popularized and applied in industrial production. The purity and the activity of the product prepared by the preparation method provided by the invention are significantly higher than those of the related similar products prepared by the prior art.

Description

technical field [0001] The invention belongs to the field of pharmacy, and in particular relates to an antiphlegm and anti-inflammatory extract of earthworm and a preparation method thereof. Background technique [0002] Earthworm, commonly known as earthworm, is a traditional Chinese medicinal material in my country. It contains a variety of biologically active ingredients in its body. It has the effects of calming the liver and calming wind, clearing heat and relieving spasm, dredging collaterals, relieving asthma, and diuresis. It is often used in some asthmatic diseases. Studies have shown that it has antihypertensive, antithrombotic, antiasthmatic, antipyretic and analgesic, anticoagulant, antibacterial, and other biological activities such as immune regulation. There are active ingredients in its body that can relieve cough and reduce phlegm; the alcohol extract has anti-inflammatory effect. The applicant has carried out a research report on the antitussive, expectora...

Claims

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Application Information

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IPC IPC(8): A61K35/62A61P11/10A61P11/00A61P31/04A61P31/12A61P29/00
CPCA61K35/62
Inventor 朱智强牛勃孙春荣王秀伟官臻
Owner 首都儿科研究所
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