A detection and analysis method of trace triptolide in a biological sample

A technology of triptolide and biological samples, which is applied in the field of detection and analysis of trace amounts of triptolide in biological samples, can solve the problems of low content, matrix interference, accuracy and mass spectrometry ionization efficiency, etc., and achieve the recovery rate High, reduce matrix interference, overcome the effect of long derivatization reaction time

Active Publication Date: 2020-12-01
QUFU NORMAL UNIV +1
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, due to the extremely low content of active pharmaceutical ingredients in biological samples, there are limitations such as serious matrix interference, so the sensitivity, accuracy and ionization efficiency of mass spectrometry need to be improved urgently.

Method used

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  • A detection and analysis method of trace triptolide in a biological sample
  • A detection and analysis method of trace triptolide in a biological sample
  • A detection and analysis method of trace triptolide in a biological sample

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Experimental program
Comparison scheme
Effect test

Embodiment 1

[0040] Chromatographic separation and mass spectrometry qualitative and quantitative analysis methods of triptolide:

[0041] In the range of 3-5000 pg / mL, prepare 7 different concentrations d 0 - MCR6G labeled triptolide standard solution (20 pg / mL, 400 pg / mL, 800 pg / mL, 1500 pg / mL, 2000 pg / mL, 3000 pg / mL, 4000pg / mL), where d 3 -MCR6G-labeled standard (2500 pg / mL) was used as fixed internal standard. The specific derivatization process is as follows: 1 part of the above-mentioned standard solution with different concentration levels, 25 μL CMPI (7 wt%) and 25 μL DMAP (10.5 wt%) acetonitrile solution, 150 μL d 0 -MCR6G or d 3 - MCR6G acetonitrile solution, add to a 1.5 mL centrifuge tube one by one, vortex for 10 seconds. Seal and react in a microwave reactor (840 W) at 56 °C for 6.5 min. 7 levels of concentration d 0 -MCR6G standard derivative and d 3 -MCR6G-labeled immobilized standard derivatives 1:1 respectively ( V / V ) to mix and shake well. The magnetic di...

Embodiment 2

[0046]The extraction of triptolide in the brain microdialysate comprises the following steps:

[0047] Take 200 μL of rat brain microdialysate into a centrifuge tube, blow dry with nitrogen, and redissolve the residue in 200 μL of acetonitrile for stable isotope labeling derivatization. Take 50 µL brain microdialysate acetonitrile reconstitution solution or standard solution, 25 µL CMPI (8 wt%) and 25 µL DMAP (15 wt%) acetonitrile solution, 150 µL d 0 -MCR6G or d 3 - MCR6G acetonitrile solution, add to a 1.5 mL centrifuge tube one by one, vortex for 10 seconds. Seal and react in a microwave reactor (850 W) at 60 °C for 6.5 min. d 0 -MCR6G-labeled brain microdialysate and d 3 -MCR6G labeled standard 1:1 ( V / V ) to mix and shake well. Disperse 10 mg of magnetic molecularly imprinted polymer into the above mixed solution and shake for 60 minutes. Magnetic separation, the adsorbed d 0 -MCR6G- triptolide derivative desorbed down. Magnetically separated and blown dry w...

Embodiment 3

[0049] The extraction of triptolide in the hemodialysate comprises the following steps:

[0050] Take 200 μL of rat blood microdialysate into a centrifuge tube, blow dry with nitrogen gas, and redissolve the residue in 200 μL of acetonitrile for stable isotope labeling derivatization. Take 50 µL hemodialysate acetonitrile reconstituted solution or standard solution, 25 µL CMPI (7 wt%) and 25 µL DMAP (14 wt%) acetonitrile solution, 100 µL d 0 -MCR6G or d 3 -MCR6G acetonitrile solution, add to 1.5mL centrifuge tube in turn, vortex for 10 seconds. Seal and react in a microwave reactor (820 W) at 57 °C for 6 min. d 0 -MCR6G-labeled hemodialysate and d 3 -MCR6G labeled standard 1:1 ( V / V ) to mix and shake well. Disperse 12 mg of magnetic molecularly imprinted polymer into the above mixed solution and shake for 60 minutes. Magnetic separation, the adsorbed d 0 -MCR6G- triptolide derivative desorbed down. Magnetically separated and blown dry with nitrogen. The residue...

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Abstract

The invention belongs to the field of analytical chemistry and in particular relates to a detection and analysis method for the content of trace triptolide in a biological sample. The method specifically comprises the following steps: carrying out stable isotopic labeling derivatization reaction on the triptolide and a derivatization reagent d0- / d3-3-N-methyl-2'-carboxyrhodamine 6G; after filtering a derived product, which is obtained by specifically extracting through a magnetic molecularly imprinted polymer, through a filtering membrane, detecting by utilizing an ultra performance liquid chromatography triple quadrupole tandem mass spectrometry analysis system. According to the detection and analysis method, the trace triptolide is labeled and derived by utilizing a stable isotopic labeling derivatization reagent with permanent positive charge d0- / d3-MCR6G; reaction is moderate and rapid and the efficiency is high; the chromatographic separation degree and the mass spectrum ionization efficiency of an analyte are remarkably improved. The magnetic molecularly imprinted polymer provided by the invention is used for selectively identifying and enriching a target object in a complicated sample and reducing matrix interference; the magnetic molecularly imprinted polymer has the advantages of simplicity in preparation, good stability and high selectivity and recycling rate.

Description

technical field [0001] The invention belongs to the field of analytical chemistry, and in particular relates to a detection and analysis method for the content of trace triptolide in biological samples, in particular, to a method using d 0 - / d 3 -3- N -Methyl-2'-carboxyrhodamine 6G ( d 0 - / d 3 -MCR6G) as a derivatization reagent, microwave-assisted-stable isotope labeling derivatization-magnetic dispersion solid-phase extraction technology combined with ultra-high performance liquid chromatography triple quadrupole tandem mass spectrometry detection analysis method. Background technique [0002] Triptolide is an active diterpene lactone compound extracted from the traditional Chinese herbal medicine Tripterygium wilfordii. It has a wide range of pharmacological properties, including anti-inflammatory, neuroprotective, immunosuppressive and anti-tumor activities. It is also the cause of toxic and side effects of triptolide. The main ingredient has attracted widespre...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): G01N30/02G01N30/06
CPCG01N30/02G01N30/06G01N2030/062G01N2030/067
Inventor 朱树芸汪鑫赵先恩徐燕秋白玉刘虎威
Owner QUFU NORMAL UNIV
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