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Derivatization gas chromatography-mass spectrometry method for detecting ammonia content of biological sample

A gas chromatography and biological sample technology, applied in the field of biological material detection, can solve the problems that affect the accuracy of plasma ammonia measurement results and cannot be accurately measured, and achieve the effects of saving time and cost, improving sensitivity, and simple and fast operation

Active Publication Date: 2018-08-17
中国医科大学
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

In the case of hemolysis, it cannot be accurately determined
At the same time, LDH and AST in plasma can also use NADPH, which affects the accuracy of plasma ammonia determination results.

Method used

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  • Derivatization gas chromatography-mass spectrometry method for detecting ammonia content of biological sample
  • Derivatization gas chromatography-mass spectrometry method for detecting ammonia content of biological sample
  • Derivatization gas chromatography-mass spectrometry method for detecting ammonia content of biological sample

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Experimental program
Comparison scheme
Effect test

Embodiment 1

[0034] Example 1 Optimization of relevant conditions for the derivatization reaction.

[0035] 1. Investigation of the pH value of the derivatization reaction:

[0036] The reaction of acid chlorides and amines (ammonia) to form amides needs to be carried out under alkaline conditions, and hydrogen chloride is generated in the product. Adding an acid-binding agent can improve the reaction efficiency. This study investigates the alkaline conditions above pH=9. It is believed that a more basic condition contributes to the stability of the product.

[0037] The experimental process is as follows:

[0038] Accurately weigh 0.5ml of whole blood into a 10ml test tube, and add 0.5ml of boric acid buffer solution (adjust the pH of the buffer with NaOH) at pH 9.0, 10.0, 11.0, 12.0, 13.0, 14.0 respectively. Add 10 μl of heptafluorobutyryl chloride derivatization reagent. After vortexing for 30 s, microwave for 1 min. Add 2.0ml of ethyl acetate, shake for 1min, centrifuge at 3500rpm ...

Embodiment 2

[0064] A derivatization gas chromatography-mass spectrometry method for detecting ammonia content in biological samples, comprising the steps of: (1) detection conditions:

[0065] Gas spectrometer: adopt 7890B-5977A gas spectrometer;

[0066] Chromatographic column: HP-5ms capillary column, specification: 30m×0.25mm, 0.25μm;

[0067] Electron ion bombardment source (EI source, 70eV), full ion scanning, scanning mass range is 40amu~600amu;

[0068] The carrier gas is high-purity helium (99.9%), and the flow rate is 1.0ml / min;

[0069] Heating program: the initial column temperature is 60°C, keep it for 1min, raise the temperature to 180°C at 15°C / min, keep it for 8min, then raise the temperature to 260°C at 30°C / min, the temperature of the injection port is 265°C, and the temperature of the ion source is 230°C ℃.

[0070] (2) Sample processing:

[0071] Accurately weigh 0.5 g of whole blood from a deceased patient with liver disease into a 10 ml test tube, add 0.5 ml of 1M...

Embodiment 3

[0080] Accurately weigh 0.5g of the urine of the deceased with liver disease, and keep the other conditions unchanged. After calculation, the ammonia content in the urine is 5.2μg / ml.

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Abstract

The invention discloses a derivatization gas chromatographic-mass spectrometric method for detecting the ammonia content of a biological sample. The method includes the steps: (1) detection condition;(2) sample treatment; (3) atlas analysis; (4) standard blood ammonia sample preparation; (5) content calculation. By a pre-column derivatization gas chromatography-mass spectrometry method, the ammonia content of the biological sample can be detected in a targeted manner, the method supplements a clinical examination blood ammonia spectrometry method, and the influence of examination blood coagulation conditions and hemolysis on detection results can be effectively reduced. Detection sensitivity is improved, detection steps are greatly simplified, and time and cost are saved.

Description

technical field [0001] The invention belongs to the field of biological material detection, in particular to a derivatization gas chromatography-mass spectrometry method for detecting ammonia content in biological samples. Background technique [0002] Ammonia in biological samples represented by blood ammonia (BAN) is an important chemical form of non-protein nitrogen. The sources of blood ammonia include endogenous: the ammonia produced by metabolism in the body is called endogenous ammonia, which mainly comes from the deamination of amino acids; partly comes from the ammonia produced by the decomposition of glutamine in renal tubular epithelial cells; the decomposition of amines can also produce Ammonia; taking an amine drug. Exogenous: Ammonia absorbed by the human body from the digestive tract is called exogenous ammonia, such as undigested protein and unabsorbed amino acids in the intestinal tract, ammonia produced by intestinal bacteria; urea in the blood diffuses to...

Claims

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Application Information

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IPC IPC(8): G01N30/88G01N30/06
CPCG01N30/06G01N30/88
Inventor 袁慧雅刘俊亭
Owner 中国医科大学
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