Identification method of periplaneta americana

A cockroach and enzymolysis technology, which is applied to instruments, measuring devices, scientific instruments, etc., can solve the problem that DNA identification cannot be applied to extracts and preparations, and can not quickly and accurately distinguish or identify cockroach and its pseudo-products. and other problems to achieve the effect of rapid identification

Active Publication Date: 2020-01-10
ZHEJIANG JINGXIN PHARMA +2
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0005] The technical problem solved by the present invention is to overcome the defects that the existing Periplaneta americana measurement or identification method cannot quickly and accurately distinguish or identify the Periplaneta americana and its counterfeit products, and DNA identification cannot be applied to extracts and preparations, etc. Provide a kind of identification method of Periplaneta americana

Method used

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  • Identification method of periplaneta americana
  • Identification method of periplaneta americana
  • Identification method of periplaneta americana

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0063] The cockroach control medicinal materials, American cockroach control medicinal materials, German cockroach medicinal materials, Dubia medicinal materials and wood beetle medicinal materials were tested, as follows:

[0064] Take about 2.5g of powder samples respectively, put them in a stoppered Erlenmeyer flask, add 50mL ether, sonicate for 30 minutes, filter, evaporate the residue to dry ether, add 50mL of 85% ethanol, sonicate for 30 minutes, centrifuge, and pour off the supernatant , the residue was dried in an oven at 60°C for 3 hours, taken out, added 20mL of 1% ammonium bicarbonate solution, ultrasonically treated for 30 minutes, filtered through a microporous membrane, and the total protein concentration was measured at 280nm ultraviolet light. Dilute until the absorbance of the solution is about 1.0, draw 100 μL of the above-mentioned filtrate into a micro-injection bottle, add 25 μL of trypsin (take trypsin for sequence analysis, add 1% ammonium bicarbonate sol...

Embodiment 2

[0071] Take 2.5g of medicinal material powder to be identified (batch number: Q20170802, place of origin: Weishan, Dali, Yunnan), put it in a stoppered Erlenmeyer flask, add 50mL of diethyl ether, sonicate for 30 minutes, filter, evaporate the residue to dry diethyl ether, add 50mL of 85% ethanol Sonicate for 30 minutes, centrifuge, pour off the supernatant, dry the residue in an oven at 60°C for 3 hours, take it out, add 1% ammonium bicarbonate solution 20mL, sonicate for 30 minutes, filter through a microporous membrane, absorb the above Put 100 μL of the filtrate into a micro-injection vial, add 30 μL of trypsin (take trypsin for sequence analysis, add 1% ammonium bicarbonate solution to make a solution containing 0.25 mg per 1 mL, and prepare it just before use), and place it in an oven at 37 °C to keep the temperature Enzymolysis for 24 hours, as the test solution.

[0072] Carry out LC-MS detection by the same method as in Example 1, take the test solution, and inject 4 ...

Embodiment 3

[0074] Take 1.5g of the medicinal material extract of the genus Blattella to be identified, add 20mL of 1% ammonium bicarbonate solution, ultrasonicate for 30 minutes, filter with a microporous membrane, absorb 100μL of the above filtrate and put it into a micro-injection bottle, add 25μL of trypsin (Take trypsin for sequence analysis, add 1% ammonium bicarbonate solution to make a solution containing 0.25mg per 1mL, and prepare it before use), put it in a 37°C oven for constant temperature enzymolysis for 24 hours, and use it as the test solution.

[0075] Carry out LC-MS detection by the same method as in Example 1, take the test solution, and inject 4 μL. The detected MRM spectrum is as Figure 8 As shown, the signal-to-noise ratio of the MRM chromatographic peaks is greater than 3:1, and the medicinal material extract is identified as the Periplaneta americana extract by the MRM spectrum.

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Abstract

The invention discloses an identification method of periplaneta americana, which comprises the specific steps of: performing enzymolysis treatment on a sample to be detected to prepare a test sample solution; then performing liquid chromatography-mass spectrometry detection on the test sample solution to determine whether the sample to be detected is the periplaneta americana or not, wherein, an electro-spray positive ion mode is adopted to perform multiple reaction monitoring, and MRM (Multiple Reaction Monitoring) scanning is performed by characteristic peptide ion pairs. The identificationmethod of the invention has the beneficial effects that: based on the found characteristic peptide, multiple reaction monitoring of mass spectrum is adopted, and the periplaneta americana and a counterfeit product thereof can be distinguished through detection of the characteristic ion pairs; the identification method has strong characteristics and high sensitivity, can identify various counterfeit varieties of the periplaneta americana, and can be suitable for identifying related extracts and preparations which cannot be subjected to DNA identification and detection.

Description

technical field [0001] The invention relates to the technical field of identification of Chinese medicinal materials, extracts and related preparations of Periplaneta americana. Background technique [0002] Periplaneta americana is an insect of the genus Periplaneta in the family Insecta Pteratae, commonly known as "cockroach". Accumulation, closed throat, internal cold and childless." The cockroach currently used clinically is mainly Periplaneta americana, and preparations such as Kangfuxin Liquid and Xinmailong Injection have been developed with this raw material, which are mainly used to treat cardiovascular diseases, wound healing, tumors and other diseases. [0003] Periplaneta americana is a commonly used medicinal material in clinical practice, and its fake products mainly include German cockroach, Dubia and wood beetle and other insects. Due to the little difference in the type and content of insect proteins, the current standard uses UV-Vis spectrophotometry to m...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N30/02G01N30/06G01N30/34G01N30/72G01N33/68
CPCG01N30/02G01N30/06G01N30/34G01N30/7266G01N33/6848G01N2030/027
Inventor 陈碧莲陈勇马若飞刘力于垂亮李尚蓉吕署一侯建
Owner ZHEJIANG JINGXIN PHARMA
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