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High-flux mass spectral method for detecting multiple amino acids in human body blood plasma

A technology for the detection of human body and amino acids, applied in the field of chemical analysis and detection, can solve the problems such as the inability to meet the needs of clinical large-scale sample detection and complexity, and achieve the effects of avoiding sample loss and human error, reducing labor costs, and improving detection efficiency.

Inactive Publication Date: 2016-03-16
GUANGDONG ZHONGKEKANGYI BIOTECH CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0003] At present, the sample pretreatment method for the detection of amino acids in plasma is mainly the traditional derivatization method, and the sample processing process is relatively complicated. The non-derivatization method has been more and more applied to the detection of blood in children because of its simple and easy pretreatment of specimens. The content of various amino acids in human body, and used to guide clinical diagnosis and treatment
However, as the amount of samples increases, the current sample pretreatment methods still cannot meet the detection needs of a large number of clinical samples, and there is an urgent need for a high-throughput, low-cost detection method in clinical practice.

Method used

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  • High-flux mass spectral method for detecting multiple amino acids in human body blood plasma
  • High-flux mass spectral method for detecting multiple amino acids in human body blood plasma
  • High-flux mass spectral method for detecting multiple amino acids in human body blood plasma

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Experimental program
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Effect test

Embodiment 1

[0020] 1. Plasma sample pretreatment steps:

[0021] In a 96-well plate, 50 μL of plasma sample to be tested, 5 μL of internal standard, 50 μL of water and 900 μL of acetonitrile were added to each well. The above mixed solution was vortexed for 3 minutes, then stood in a refrigerator at 4°C for 30 minutes, the treated solution was centrifuged at 3000r / min for 10 minutes, and the supernatant was transferred to a 96-well sample plate with a row gun. Among them, the 17 kinds of amino acids to be tested and their internal standards are shown in the following table:

[0022] serial number

amino acid

abbreviation

MW

isotopic internal standard

1

Methionine

met

150

D3

2

Leucine

Leu

137

D3

3

Asparagine

Asn

132

D3

4

Alanine

Ala

90.2

D4

5

Tyrosine

Tyr

181

13 C6

6

Phenylalanine

Phe

165.3

13 C6

7

threonine

Thr...

Embodiment 2

[0037] Determination of actual samples

[0038] According to the sample pretreatment method in Example 1, 96 normal human plasma samples (provided by Beijing Aviation General Hospital) were subjected to protein precipitation and amino acid extraction, and the amino acid extract was transferred to a 96-well sample plate, according to Example 1 Samples were analyzed and data processed under optimized mass spectrometry and liquid phase conditions, and the concentration of each amino acid was calculated according to the internal standard method, 5 of which are listed in the table below. figure 2 It is the high performance liquid chromatogram of 17 kinds of amino acids in sample 1.

[0039]

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Abstract

The invention discloses a high-flux mass spectral method for detecting multiple amino acids in human body blood plasma. The method comprises precipitating protein in blood plasma by employing an organic solvent, adding a stable isotope internal standard, performing high-speed centrifugation to extract amino acids contained in the supernatant, and directly injecting a sample to detect the multiple amino acids in the blood plasma. A 96-well plate sample pretreatment process is employed, protein precipitation, internal-standard addition and centrifugation are performed in a 96-well plate, then the 96-well plate is directly connected with an LC-MS automatic sample-injection device, 96 samples can be processed and analyzed at a time, the detection efficiency is substantially improved, also sample loss and personal error caused by repeated transfer of samples are avoided, and labor cost is reduced. At present, detection on amino acids is mainly applied to screening and diagnosis on inherited metabolic diseases, along with more and more attention of people, more and more clinic detection samples are provided. The method is a rapid high-flux detection method for detecting a large amount of clinic samples.

Description

technical field [0001] The invention belongs to the field of chemical analysis and detection, can perform fast and high-throughput sample pretreatment, greatly improves detection efficiency, and reduces labor costs, and specifically relates to a mass spectrometry method for high-throughput detection of various amino acids in human plasma. Background technique [0002] Inherited metabolic diseases refer to a group of diseases in which certain enzymes, carrier proteins, membrane receptors and other coding genes involved in the metabolism of the body undergo mutations, making the products encoded by them dysfunctional and causing metabolic disorders. Many genetic metabolic diseases are associated with abnormal amino acid metabolism, often with multiple amino acids. Amino acid genetic metabolic diseases not only have a large number of diseases and complex pathogenesis, but also have diverse and lack of specificity in clinical manifestations. Therefore, there is a great need for...

Claims

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Application Information

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IPC IPC(8): G01N30/02G01N30/06
CPCG01N30/02G01N30/06G01N2030/027
Inventor 李岩谢雯春李倩倩
Owner GUANGDONG ZHONGKEKANGYI BIOTECH CO LTD
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