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Method for simultaneously detecting trimethylamine oxide and phenylacetyl glutamine as well as detection kit and application thereof

A technology for oxidizing trimethylamine and glutamine, which is applied in the field of analysis and detection, and achieves the effects of easy popularization and application, less sample volume, and improved accuracy

Pending Publication Date: 2021-07-27
PEKING UNION MEDICAL COLLEGE HOSPITAL CHINESE ACAD OF MEDICAL SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

[0003] At present, there is only a mass spectrometry method for the determination of TMAO in blood or urine (Wang Z, et al, Measurement oftrimethylamine-N-oxide by stable isotope dilution liquid chromatographytandem mass spectrometry, Anal Biochem, 2014, 455:35-40; Qiu W, et al,A faster and simpler UPLC-MS / MS method for the simultaneous determination oftrimethylamine N-oxide,trimethylamine and dimethylamine in different types of biological samples,Food Funct,2019,10,6484-6491), or PAGln in blood and urine Determination of mass spectrometry (Laryea MD, et al, Simultaneous LC-MS / MS determination ofphenylbutyrate, phenylacetate benzoate and their corresponding metabolitesphenylacetylglutamine and hippurate in blood and urine.J Inherit MetabDis.2010Dec; 33Suppl 3:S321-8.), but not yet There is no method for simultaneous detection of TMAO and PAGln

Method used

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  • Method for simultaneously detecting trimethylamine oxide and phenylacetyl glutamine as well as detection kit and application thereof
  • Method for simultaneously detecting trimethylamine oxide and phenylacetyl glutamine as well as detection kit and application thereof
  • Method for simultaneously detecting trimethylamine oxide and phenylacetyl glutamine as well as detection kit and application thereof

Examples

Experimental program
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Effect test

Embodiment 1

[0052] This embodiment provides a method for simultaneously detecting trimethylamine oxide and phenylacetylglutamine, which is detected by liquid chromatography tandem mass spectrometry, as follows:

[0053] 1. Sample pretreatment

[0054] Take 50 μL serum or 20-fold diluted urine or mixed standard solution into a 2ml plastic tube or glass tube, add 20 μL isotope internal standard, mix and shake, add 130 μL precipitant, mix and shake, centrifuge at 13000 rpm for 10 minutes, take 100 μL of supernatant, add 200 μL mobile phase initial concentration mixture, waiting for the test on the machine, injection volume: 2 μL.

[0055] Mixed standard solution: Dissolve the standard products of TMAO and PAGln in distilled water to prepare a mother liquor, mix the mother liquor of TMAO and the mother liquor of PAGln, use a constant volume bottle to make accurate volume, and perform a series of dilutions with acetonitrile so that the concentration of TMAO and PAGln is 0.02 ,0.05,0.1,0.2,0.5...

Embodiment 2

[0071] This embodiment provides a method for simultaneously detecting trimethylamine oxide and phenylacetylglutamine. The method is detected by liquid chromatography tandem mass spectrometry. The difference from Example 1 is that the concentration of ammonium acetate in the precipitant is 10mM. The liquid chromatogram under this condition is shown in figure 2 .

experiment example 1

[0074] Precision Evaluation of Experimental Example 1 Detection

[0075] Adopt the method of embodiment 1 to carry out the simultaneous detection of trimethylamine oxide and phenylacetylglutamine, evaluate the precision of detection method, specific method is as follows: use the method of embodiment 1 to measure the target in three different serums and diluted urine For the concentration of metabolites, each sample was measured 5 times, and five parallel tubes were set up each time, and each sample preparation and measurement were completed on different working days. Average within-assay and total imprecision (CV) were calculated. , The evaluation results are shown in Table 3, and the results show that the precision of the detection method of the present invention is good.

[0076] Table 3 Precision evaluation results

[0077]

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Abstract

The invention relates to the technical field of analysis and detection, in particular to a method for simultaneously detecting trimethylamine oxide and phenylacetylglutamine as well as a detection kit and application thereof. According to the detection method provided by the invention, a to-be-detected sample is subjected to sample pretreatment, then detection is performed by adopting a liquid chromatography-tandem mass spectrometry method, the sample pretreatment comprises the following steps: the to-be-detected sample is subjected to precipitation treatment by adopting a precipitant to obtain separated supernate, and the precipitant is an acetonitrile and methanol mixed solution containing 10-100mM ammonium acetate. The method has the advantages of simple sample reservation, simple sample treatment, small sample use amount and fast detection speed, can reduce the problems of two-time sample reservation, time consumption, cost increase and the like due to respective determination, and is easy to popularize and apply; the detection method has high precision and low matrix effect, and can significantly improve the accuracy of trimethylamine oxide and phenylacetyl glutamine detection.

Description

technical field [0001] The invention relates to the technical field of analysis and detection, in particular to a method for simultaneously detecting trimethylamine oxide and phenylacetylglutamine, a detection kit and application thereof. Background technique [0002] Trimethylamine-N-oxide (TMAO) and phenylacetylglutamine (PAGln) are the metabolites of intestinal microorganisms. PAGln is the metabolite produced by the liver after the unabsorbed phenylalanine in the intestine is metabolized into phenylacetic acid by microorganisms. [0003] At present, there is only a mass spectrometry method for the determination of TMAO in blood or urine (Wang Z, et al, Measurement oftrimethylamine-N-oxide by stable isotope dilution liquid chromatographytandem mass spectrometry, Anal Biochem, 2014, 455:35-40; Qiu W, et al,A faster and simpler UPLC-MS / MS method for the simultaneous determination oftrimethylamine N-oxide,trimethylamine and dimethylamine in different types of biological samp...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N30/06G01N30/88
CPCG01N30/06G01N30/88
Inventor 禹松林邱玲唐月明王丹晨
Owner PEKING UNION MEDICAL COLLEGE HOSPITAL CHINESE ACAD OF MEDICAL SCI
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