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Method for detecting insect in-vivo juvenile hormone JH II based on chromatography-mass spectrometry

A detection method, juvenile hormone technology, applied in the field of analytical chemistry, can solve the problem of low detection sensitivity of juvenile hormone JHII, and achieve the effect of ensuring quantitative accuracy, high sensitivity and good repeatability

Active Publication Date: 2017-05-24
DALIAN INST OF CHEM PHYSICS CHINESE ACAD OF SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] The present invention aims at the low detection sensitivity of juvenile hormone JH II in insects, and proposes to use liquid-liquid extraction and purification, solid phase extraction and purification, liquid chromatography separation and triple quadrupole mass spectrometry to detect juvenile hormone JH II in insects. This method can eliminate the interference of other substances in insects on the detection of juvenile hormone JH II. It has the characteristics of good repeatability and high sensitivity, and is suitable for the detection and research of juvenile hormone JH II in insects.

Method used

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  • Method for detecting insect in-vivo juvenile hormone JH II based on chromatography-mass spectrometry
  • Method for detecting insect in-vivo juvenile hormone JH II based on chromatography-mass spectrometry
  • Method for detecting insect in-vivo juvenile hormone JH II based on chromatography-mass spectrometry

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Embodiment 1

[0015] Example 1 Evaluation of the method for detecting juvenile hormone JH II in insects based on liquid chromatography-mass spectrometry

[0016] Concentration is 1mg / mL juvenile hormone JH II standard sample is diluted into a series of different concentration solutions (20ng / mL, 10ng / mL, 5ng / mL, 1ng / mL, 0.5ng / mL, 0.1ng / mL with volume concentration 60% acetonitrile / mL, 0.05ng / mL, 0.025ng / mL, 0.01ng / mL). Then take 200 μL of standard sample solutions with different concentrations, add 20 μL of internal standard methoprene solution (concentration: 50 ng / mL) and mix well, and wait for instrumental analysis. Adopt the above-mentioned standard solution of different concentrations that adds internal standard to carry out liquid chromatography mass spectrometry separation and analysis, investigate the linearity of instrument method, take the concentration of juvenile hormone JH II and the relative area of ​​peak as coordinates to draw linear standard curve, linear correlation coeff...

Embodiment 2

[0018] Embodiment two 4 instar cotton bollworm sample detection

[0019] Take 4 samples of the 4th instar cotton bollworm, put them into a 2mL Eppendorf tube, weigh 191.8mg, add grinding balls, add 250μL of acetonitrile, 250μL of 2% sodium chloride aqueous solution, 500μL of 1ng / mL internal standard Grind the n-hexane of methoprene for 2 minutes, centrifuge at 12000g for 5 minutes at 4°C, extract the supernatant n-hexane layer, then add 500 μL of pure n-hexane to the residue, grind, extract the supernatant, and repeat the extraction once with pure n-hexane , put the crude extract into a 2mL Eppendorf tube, vortex for 1min, centrifuge at low speed for 15s, blow dry slowly with nitrogen to obtain the crude extract; The solid-phase extraction column was pre-washed twice with 1 mL of methanol in advance, and eluted with 200 μL of acetonitrile for 3 times. The eluate in the collection box was taken out, and the collection box was washed once with 200 μL of acetonitrile. Centrifuge...

Embodiment 3

[0022] Embodiment three 3 instars cotton bollworm sample detection

[0023] Take 6 samples of the 3rd instar cotton bollworm, weigh 127.2 mg, add grinding balls, 250 μL of acetonitrile, 250 μL of 2% sodium chloride aqueous solution, 500 μL of n-hexane containing 1 ng / mL internal standard methoprene, and grind for 2 minutes , centrifuge at 12000g for 5min at 4°C, extract the supernatant n-hexane layer, repeat the extraction twice with 500μL pure n-hexane, combine the crude extract into a 2mL Eppendorf tube, vortex for 1min, centrifuge at low speed for 15s, blow dry with nitrogen; Add 200 μL of 60% acetonitrile to the dried crude extract, vortex for 1 min, then pass the reconstituted solution through a solid phase extraction column into a collection box, and elute with 200 μL acetonitrile for 3 times, take out the eluate in the collection box, and wash it with 200 μL Rinse the collection box once with acetonitrile, put it into a 2mL Eppendorf tube together with the eluent, centrif...

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Abstract

The invention discloses a method for detecting insect in-vivo juvenile hormone JH II based on chromatography-mass spectrometry and relates to application of analytical chemistry in entomology research. The method comprises the specific steps that an insect sample is taken, grinding balls, acetonitrile, a sodium chloride water solution and normal hexane are added for grinding, high-speed centrifugation is performed to obtain a normal-hexane-layer crude extract, then normal hexane is added to the sample for two times of grinding and extraction, the crude extract obtained through three times of grinding is added to a collecting bottle, and nitrogen blow-drying is performed; re-dissolution is performed with an acetonitrile-water solution, then the acetonitrile is eluted for three times by using a solid-phase extraction column, and eluant in a collecting box is taken out and subjected to nitrogen blow-drying; re-dissolution is performed with an acetonitrile-water solution, and analysis is performed by using a liquid chromatograph-mass spectrometer. The detecting method is high in sensitivity, strong in matrix interference resisting capability and good in repeatability and can provide help for research of in-vivo juvenile hormone JH II change laws of insects at all stages.

Description

technical field [0001] The invention belongs to the field of analytical chemistry, and specifically relates to a method for detecting juvenile hormone JHII in insects based on liquid chromatography-mass spectrometry, which uses extraction, purification pretreatment, liquid chromatography separation, and triple quadrupole mass spectrometry detection analysis A method for detecting the content of juvenile hormone JH II in insects by means of chemical technology. Background technique [0002] Insects are the most abundant animals in the world and play an important role in the biosphere. Studying the growth law and physiological functions of insects is of great significance to the control and management of insects. Juvenile hormone is a sesquiterpene compound synthesized by the pharyngeal body of insects and secreted into the hemolymph. It can regulate many physiological functions of insects, including growth, reproduction, polymorphism, diapause and pheromone production. In ...

Claims

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Application Information

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IPC IPC(8): G01N30/88G01N30/06
CPCG01N30/06G01N30/88
Inventor 王晓琳许国旺
Owner DALIAN INST OF CHEM PHYSICS CHINESE ACAD OF SCI
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