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Method for measuring content of 4 sterols in fresh cordyceps sinensis simultaneously by HPLC-ELSD, and application thereof

A technology of Cordyceps sinensis and sterols, which is applied in the application field of identifying the authenticity of Cordyceps sinensis, can solve the problems of unstable baseline drift, high reagent toxicity, and small response value, and achieve good separation effect, simple method, and stable results

Active Publication Date: 2018-01-12
DONGGUAN HEC CORDYCEPS R&D CO LTD +1
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Simultaneously, at present, the identification of genuine and counterfeit products of Cordyceps sinensis is generally identified through physical and chemical properties in addition to microscopic, character, and genetic identification, such as the document "Cordyceps sinensis nuclear magnetic characteristic fingerprint establishment and identification research" ("World Science and Technology-Modernization of Traditional Chinese Medicine" , 2014,16(11)), "HPLC Fingerprint Study on Cordyceps and Its Confused Products and Identification of Common Components" ("Chinese Herbal Medicine", 2017,48(5):991-996), but currently the characteristic components in Cordyceps sinensis There are relatively few studies on sterol active substances. There are only a few literature reports using high performance liquid chromatography-ultraviolet detector (HPLC-UV) or HPLC-UV-MS, GC-MS to measure the content of ergosterol in Cordyceps sinensis. Simultaneous quantitative analysis of four sterols is rarely reported
As reported in the document "Analysis of Chemical Components of Cordyceps Sinensis" ("Acta Mycophyta Sinica", 2016, 35(4):476-490), the sample was derivatized by BSTFA / TMS method, although 7 sterols were identified by GC-MS similar substances, but the sample needs to be derivatized, which makes the sample processing process complicated, and the reagents used are highly toxic and time-consuming
In the document "High Performance Liquid Chromatography Determination of the Content of Ergosterol in Artificial Cordyceps" ("Journal of Shanxi Medical University", 2016,37(4):661-662), after the sample is saponified with saturated KOH solution and absolute ethanol, the Cyclohexane was extracted 3 times, and the extract was purified and concentrated with a distribution column and an adsorption column, and finally the amount of ergosterol was determined by HPLC-UV separation. UV has determined the content of ergosterol in the artificial Cordyceps; Similarly, the literature "Quantitative Analysis of Ergosterol in Cordyceps sinensis (Cordyceps sinensis) Mycelia Solid Fermentation Powder" ("Journal of Shenyang Normal University", 2005,23 (3): 293 -296) also adopts a similar method to prepare samples, and the result is only quantitative analysis of ergosterol, and if the sterol terminal absorption substances such as cholesterol and sitosterol in Cordyceps sinensis are measured, there will be unstable baseline drift and response in HPLC-UV detection. Disadvantages such as small value, low sensitivity and poor repeatability

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  • Method for measuring content of 4 sterols in fresh cordyceps sinensis simultaneously by HPLC-ELSD, and application thereof
  • Method for measuring content of 4 sterols in fresh cordyceps sinensis simultaneously by HPLC-ELSD, and application thereof
  • Method for measuring content of 4 sterols in fresh cordyceps sinensis simultaneously by HPLC-ELSD, and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0040] Embodiment 1 Methodological investigation

[0041] 1.1 Instrument

[0042] High performance liquid chromatography; evaporative light scattering instrument.

[0043] 1.2 Solution preparation

[0044] 1.2.1 Preparation of reference solution

[0045] Weigh 20.0mg each of ergosterol, cholesterol, stigmasterol and sitosterol, weigh them accurately, place them in a 100mL volumetric flask, add 0.5mol / L potassium hydroxide-ethanol solution to fully dissolve and dilute to the mark, then .

[0046] 1.2.2 Sample preparation: take 0.5g of Cordyceps sinensis sample (A7), add 10mL 0.5mol / L potassium hydroxide-ethanol solution for extraction, reflux at 85°C for 30min, centrifuge to obtain supernatant, filter through membrane, Obtain the test sample solution.

[0047] 1.3 HPLC-evaporative light scattering analysis conditions

[0048] Chromatographic column: Waters Xbridge C18 (4.6×250mm, 5μm); mobile phase A is water, mobile phase B is 0.005% formic acid-methanol solution; inject...

Embodiment 2

[0065] The HPLC-ELSD chromatogram of embodiment 2 fresh Cordyceps sinensis

[0066] 2.1 Preparation of fresh Cordyceps sinensis samples

[0067] Accurately weigh 1.5g of fresh Cordyceps sinensis (A4, A5, A6), add 10mL 0.5mol / L potassium hydroxide-ethanol solution for extraction, reflux at 85°C for 30min, centrifuge to obtain the supernatant, filter through a filter membrane to obtain The test sample solution.

[0068] 2.2 High-performance liquid phase-evaporative light scattering (HPLC-ELSD) analysis conditions: the same as in the item "1.3" of Example 1.

[0069] 2.3 Data processing and analysis

[0070] Each test sample solution was injected into the HPLC-ELSD coupler for detection. The retention time and integration of the chromatograms of each sample were automatically completed by the workstation, and the peak position of the reference substance was used as a reference to identify the chromatographic peaks of ergosterol, cholesterol, stigmasterol and sitosterol in fres...

Embodiment 3

[0073] Embodiment 3 Application of HPLC-ELSD detection method

[0074] 3.1 Preparation of the test solution

[0075] Accurately weigh 0.5g of each sample powder of Cordyceps sinensis (wild Cordyceps sinensis A1-A3, Cordyceps sinensis breeding product A7-A9), Cordyceps sinensis counterfeit product (A10-A13), Bailing capsule (A14) and Jinshuibao capsule (A15) sample powder, respectively placed in Into a 50mL glass bottle, add 10mL of 0.5mol / L potassium hydroxide-ethanol solution, and reflux at 85°C for 30min. Centrifuge to obtain supernatant. Filter through a filter membrane to obtain the test sample solution.

[0076] 3.2 High-performance liquid phase-evaporative light scattering (HPLC-ELSD) analysis conditions: the same as in the item "1.3" of Example 1.

[0077] 3.3 Data processing and analysis

[0078] Each test sample solution was injected into the HPLC-ELSD coupler for detection. The retention time and integration of the chromatograms of each sample are automatically ...

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Abstract

The invention belongs to the field of a method for performing quantitative analysis and detection on special ingredients in traditional Chinese medicinal materials, and particularly discloses a methodfor measuring the content of four sterols in fresh cordyceps sinensis simultaneously by HPLC-ELSD, and application of the method to true and false identification of cordyceps sinensis. The method comprises the following steps: (1) preparing a test product solution; (2) preparing a reference solution; and (3) performing HPLC-ELSD measurement and detecting four sterol substances in the cordyceps sinensis. The sample treatment method provided by the invention is simple and low in time consumption; furthermore the analysis method provided by the invention can fulfill the aim of quantifying the four sterol components simultaneously, and is stable, accurate and reliable in result; and by the detection method provided by the invention, the quality standard of the cordyceps sinensis taking the content of the four sterol components in the cordyceps sinensis as indexes can be established, and the method can be applied to true and false identification of cordyceps sinensis traditional Chinese medicine materials.

Description

technical field [0001] The invention belongs to the field of quantitative analysis and detection methods for specific components in Chinese medicinal materials, and in particular relates to a method for simultaneously measuring the contents of four sterols in Cordyceps sinensis by using HPLC-ELSD, and the application of the method in identifying genuine and counterfeit products of Cordyceps sinensis. Background technique [0002] Cordyceps sinensis is a complex of larvae and larvae of Cordyceps sinensis (BerK.) Sacc. It has the effects of invigorating the lungs and kidneys, secreting essence and nourishing qi, etc. It is mainly used for chronic cough, dyspnea, hemoptysis, impotence and nocturnal emission, and soreness of the waist and knees. It is a traditional Chinese medicinal material in my country. Modern pharmacological studies have shown that Cordyceps is beneficial to the human immune system, circulatory system, and cardiovascular system. However, wild Cordyceps sine...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N30/02G01N30/06
Inventor 李文佳孙敏甜李文庆钱正明邱健健
Owner DONGGUAN HEC CORDYCEPS R&D CO LTD