Method for preparing timosaponin BIII and uses thereof

A technology of timosaponin and mother saponin, which is applied in the field of preparation of plant extracts, can solve problems such as not being able to meet industrial applications, and achieve good anti-diabetic activity and simple preparation method

Inactive Publication Date: 2008-11-19
SHANGHAI INST OF MATERIA MEDICA CHINESE ACAD OF SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, the HPLC separation method is only suitable for laboratories and cannot be satisfied with industrial applications.

Method used

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  • Method for preparing timosaponin BIII and uses thereof
  • Method for preparing timosaponin BIII and uses thereof
  • Method for preparing timosaponin BIII and uses thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0034] Embodiment 1: Extraction and separation method one of timosaponin BIII

[0035] Take 5.0 kg of Anemarrhena medicinal material, degrease by gasoline percolation, and dry. Reflux extraction with 25 liters of 85% ethanol three times for 2 hours each time, filter, and combine the filtrates. Recover the solvent under reduced pressure, concentrate until it has no alcohol smell, and obtain 1 kg of mother extract.

[0036] Suspend the extract with 6 liters of water, and extract four times with 4 liters of n-butanol saturated with water. The n-butanol layer was recovered and dried to obtain 135 grams of extract. The sample was mixed with methanol for silica gel column (2700 g, 160-200 mesh) chromatography. First elute with chloroform until the color is light; then elute with chloroform-n-butanol (1:1). TLC detection, the developing solvent is n-butanol-ethyl acetate-water (4:1:5) upper layer. Using 10% phosphomolybdic acid ethanol at 110oC as the developer, the fractions co...

Embodiment 2

[0041] Embodiment 2: Extraction and separation method two of timosaponin BIII

[0042] Take 5.0 kg of Anemarrhena medicinal material, degrease by petroleum ether percolation, and dry. Reflux extraction with 40 liters of methanol three times for 2 hours each time, filter, and combine the filtrates. The solvent was recovered under reduced pressure, concentrated until there was no alcohol smell, and 1.2 kg of the mother extract was obtained.

[0043] Suspend the extract with 10 liters of water, and extract four times with 10 liters of n-pentanol saturated with water. The n-pentanol layer was recovered and dried to obtain 150 grams of extract. The sample was mixed with methanol for silica gel column (2700 g, 160-200 mesh) chromatography. First eluted with chloroform until the color became light; then eluted with chloroform-n-butanol (5:1). TLC detection, the developing solvent is n-butanol-ethyl acetate-water (4:1:5) upper layer. Using 10% ethanol phosphomolybdic acid as the ...

Embodiment 3

[0045] Embodiment 3: In vitro inhibition of α-glucosidase activity test

[0046] a) Reagents and instruments:

[0047] α-glucosidase (EC 232-604-7), purchased from Sigma Company, USA,

[0048] pNPG (4 nitrophenyl-α-D-glucopyranoside) (EC 223-189-3), purchased from Sigma

[0049] Anhydrous sodium carbonate, phosphate, etc., are analytically pure.

[0050] Microplate reader: BIO-TECK Instruments, produced in the United States.

[0051] b) Reagent preparation

[0052] Phosphate buffer (67mM, PH6.8): Weigh an appropriate amount of K 2 HPO 4 ·3H 2 O solution, adjusted to pH 6.8 with phosphoric acid, stored at 4°C until use.

[0053] Enzyme solution: Weigh an appropriate amount of α-glucosidase freeze-dried powder and dilute it with 67mM, pH 6.8 phosphate buffer solution at a concentration of 0.5mg / ml, aliquot 0.5ml into one tube, and freeze at -20°C.

[0054] Substrate PNPG: Prepare 29mM α-PNPG with phosphate buffer (67mM, pH 6.8), aliquot and store at -20°C.

[0055] Reac...

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Abstract

The invention relates to a method for preparing timosaponin BIII. The method separates an effective constituent of timosaponin BIII from a Chinese traditional medicine anemarrhenae in a chemical way. Simultaneously in-vivo and in-vitro pharmacological tests verify that: as the compound can remarkably inhibit the activity of alpha-glucosaccharase, and effectively reduce animal blood sugar, the compound can be applicable to the preparation of alpha-glucosaccharase inhibitors and medicines for curing diabetes. The method for preparing the timosaponin BIII replaces HPLC preparation with processes such as crystallization with simple operation, thereby facilitating industrialized production more effectively. Simultaneously, the compound displays good anti-diabetes activity in vivo and vitro, thereby not only widening the application field of the compound, but also becoming the effective medicine for curing diabetes in future.

Description

technical field [0001] The present invention relates to the preparation method and application of the plant extract, more specifically to the method for extracting and separating the active ingredient timosaponin BIII from the traditional Chinese medicine Zhimu, and the application of timosaponin BIII in the preparation of medicines for treating diabetes. Background technique [0002] Diabetes is a common clinical disease caused by the absolute or relative lack of insulin in the human body. The concentration of glucose in the blood increases, and then a large amount of sugar is excreted from the urine, resulting in polydipsia, polyuria, polyphagia, weight loss, dizziness, and fatigue. and other symptoms. Further development then causes various serious acute and chronic complications throughout the body, threatening health. Among them, patients with type I diabetes have completely lost the function of producing insulin, and insulin-dependent treatment must be used for life; ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07J71/00A61K31/7048A61K36/8964A61P3/10
Inventor 黄成钢马春辉唐意红李志雄赵维民范明松朱海燕孙兆林施松
Owner SHANGHAI INST OF MATERIA MEDICA CHINESE ACAD OF SCI
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