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Analysis method of glucose energy metabolism-related important metabolites in serum samples

An analytical method and energy metabolism technology, applied in the field of analytical chemistry and biochemistry, can solve problems such as unseen, complex extraction, and high solvent toxicity, and achieve the effects of low solvent toxicity, simple pretreatment, and high sensitivity

Active Publication Date: 2017-06-16
DALIAN INST OF CHEM PHYSICS CHINESE ACAD OF SCI
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  • Abstract
  • Description
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Problems solved by technology

The determination of glucose mainly includes enzymatic method, electrochemical method, photochemical method, etc.; the quantitative analysis of the intermediate products of the tricarboxylic acid cycle mostly uses the method of gas chromatography-mass spectrometry, such as Calderón-Santiago, etc., using chloroform-methanol-water dual-phase extraction, selected ion scanning mode detection , but there are problems such as complicated extraction and high solvent toxicity
Gas chromatography-triple quadrupole mass spectrometry, as a powerful tool for targeted analysis, through gas chromatography separation combined with selected ion scanning of two-stage mass spectrometry, can eliminate matrix interference to the greatest extent, and has the advantages of high sensitivity and wide linear range ; and there is no report on the analysis of important metabolites related to glucose energy metabolism based on gas chromatography-triple quadrupole mass spectrometry coupled with multiple reaction monitoring mode

Method used

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  • Analysis method of glucose energy metabolism-related important metabolites in serum samples
  • Analysis method of glucose energy metabolism-related important metabolites in serum samples
  • Analysis method of glucose energy metabolism-related important metabolites in serum samples

Examples

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Embodiment 1

[0017] Evaluation of a method based on gas chromatography-triple quadrupole mass spectrometry for the detection of important metabolites related to glucose energy metabolism in serum samples

[0018] Referring to the HMDB database (http: / / www.hmdb.ca / ), for example, the range of blood sugar levels in people of different ages is 1100-16440 μM, that is, 198.2-2961.8 μg / mL. Mixed standard solutions were prepared according to the reference concentration range of metabolites in the database. ), malic acid (800.0μg / mL), 2-hydroxyglutarate (800.0μg / mL), 2-ketoglutarate (800.0μg / mL), aconitic acid (800.0μg / mL), citric acid ( 800.0μg / mL), isocitric acid (800.0μg / mL), glucose (4000μg / mL), the mixed standard solution was diluted 1.33 times, 1.6 times, 2.67 times, 4 times, 8 times, 40 times, 80 times, 400 times, 800 times, 4000 times, 8000 times, 20000 times, 40000 times, 100000 times, 200000 times, 500000 times, 1000000 times, 2500000 times to get mixed standard solutions with different...

Embodiment 2

[0025] Detection of important metabolites related to glucose energy metabolism in normal human serum samples

[0026] Thaw normal human serum samples stored at -80°C, vortex for 10 seconds, pipette 50 μL accurately, add 200 μL of methanol solution containing internal standard, and vortex for 1 min. The concentrations of the internal standards in the methanol solution are: lactic acid-3,3,3-d 3 (100μg / mL), succinate-2,2,3,3-d 4 (0.2 μg / mL), fumarate-2,3-d 2 (0.1μg / mL), citrate-2,2,4,4-d 4 (5μg / mL), glucose- 13 C 6 ,1,2,3,4,5,6,6-d 7 (150 μg / mL). Centrifuge at 14,000 rpm for 15 min at 4°C, and take 200 μL of the supernatant solution to freeze-dry. Add 50 μL of methoxyamine solution (20 mg.ml, pyridine) to the freeze-dried sample, vortex for 30 s, sonicate for 1 min, and react in a 37°C water bath for 1.5 h; silylyl) trifluoroacetamide, vortexed for 10s, and reacted in a 37°C water bath for 1h; after the reaction was completed, centrifuged at 14000rpm for 10min at 4°C, an...

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Abstract

The invention discloses an analysis method of glucose energy metabolism-related important metabolites in serum samples. According to the analysis method, methanol is adopted for extraction, samples to be detected are subjected to freeze-drying and two-step derivatization, and then are subjected to analysis using a gas chromatography-triple quadrupole mass spectrometer; multi-reaction monitoring mode is adopted for detection, for low-content metabolites, ion pairs with the optimal ion abundance are obtained via optimization and are taken as quantification ion pairs so as to increase sensitivity; for high-content metabolites, ion pairs low in abundance obtained via optimization are selected as quantification ion pairs so as to widen the linear range and realize simultaneous analysis of high and low abundance metabolites. Pretreatment of the analysis method is simple; repeatability is high; sensitivity is high; and linear range is wide.

Description

technical field [0001] The invention relates to the fields of analytical chemistry and biochemistry, and is a method for analyzing important metabolites related to glucose energy metabolism in serum samples based on gas chromatography-triple quadrupole mass spectrometry combined with multiple reaction monitoring mode. Background technique [0002] Glucose is the transport form of sugar in the blood, occupies a major position in the body's glucose metabolism, and is an important substance for maintaining life activities. Glucose is transformed into pyruvate through glycolysis and produces energy, and pyruvate is completely oxidized to form CO through the tricarboxylic acid cycle and oxidative phosphorylation under aerobic conditions. 2 and H 2 O, producing a large amount of free energy; and converted into lactic acid under anaerobic conditions. Glycolysis to complete oxidation of glucose not only provides a large amount of free energy for the body, but also provides precurs...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N30/88
Inventor 许国旺周洋路鑫张俊杰
Owner DALIAN INST OF CHEM PHYSICS CHINESE ACAD OF SCI
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