Combined detection method for multiple vitamin D metabolites and detection kit and application thereof

A technology for metabolites and vitamins, applied in the field of analysis and detection, to achieve the effects of saving sample pretreatment time, high sensitivity, and avoiding interference

Active Publication Date: 2021-08-27
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

Most detection methods in the prior art either rely on sample derivation or are only able to detect individual metabolites

Method used

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  • Combined detection method for multiple vitamin D metabolites and detection kit and application thereof
  • Combined detection method for multiple vitamin D metabolites and detection kit and application thereof
  • Combined detection method for multiple vitamin D metabolites and detection kit and application thereof

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Experimental program
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Embodiment 1

[0045] This embodiment provides a method for detecting vitamin D metabolites, which is detected by liquid chromatography tandem mass spectrometry, specifically as follows:

[0046] 1. Sample pretreatment

[0047] Add 200 μL of serum samples, standards or quality control products in sequence to a 5 mL glass tube, then add 50 μL of mixed internal standard solution and 200 μL of methanol in sequence, and mix thoroughly; then add 500 μL of 0.1M zinc sulfate aqueous solution, 1 mL of n-hexane and ethyl acetate (The volume ratio of n-hexane and ethyl acetate is 8:2), vortex, mix well, centrifuge at 12000g for 5min, transfer 750μL of the supernatant to a 2mL 96-well plate, and dry it with nitrogen (37℃) ; Add 100 μL methanol aqueous solution to redissolve (V:V=5:5) to obtain the sample to be tested. Take 40 μL of the sample to be tested for LC-MS / MS analysis and detection.

[0048] Mixed internal standard solution: the concentration of each metabolite is: 25OHD 3 -d 3 (50ng / mL), ...

Embodiment 2

[0070] This embodiment provides a method for detecting vitamin D metabolites. The method is detected by liquid chromatography tandem mass spectrometry. The difference from Example 1 is that in mobile phase A and mobile phase B, the concentration of lithium acetate is 0.22mM.

experiment example 1

[0071] Experimental example 1 Detection of separation effect of vitamin D metabolites

[0072] The detection method of Example 1 is used to carry out liquid phase separation of vitamin D metabolites and their interfering structural analogs, and the results are as follows figure 1 and figure 2 As shown, the results show that the detection method of the present invention can well 25-hydroxyvitamin D3, 25-hydroxyvitamin D2, 1,25 dihydroxyvitamin D3, 1,25 dihydroxyvitamin D2, 24,25 dihydroxyvitamin Six vitamin metabolites, D3, 24,25 dihydroxyvitamin D2, were separated and separated from the interfering substance 3-epi 25(OH)D 3 , 3-epi 24,25(OH) 2 D. 3 、23R,25(OH)D 3 , 4β,25(OH) 2 D is well separated.

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Abstract

The invention relates to the technical field of analysis and detection, in particular to a combined detection method for multiple vitamin D metabolites and a detection kit and application thereof. According to the method for detecting the vitamin D metabolites, liquid chromatography-tandem mass spectrometry is adopted for detection, a mobile phase of liquid chromatography comprises a mobile phase A and a mobile phase B, the mobile phase A is an aqueous solution containing lithium acetate, and the mobile phase B is a methanol solution containing lithium acetate. According to the method, the simultaneous detection of 25OHD3, 25OHD2, 1, 25 (OH) 2D3, 1, 25 (OH) 2D2, 24, 25 (OH) 2D3 and 24, 25 (OH) 2D2 can be realized, and various isomers and interferences can be distinguished at the same time; and the method has high sensitivity and specificity, does not need derivatization treatment, saves sample pretreatment time, and provides convenience for high-throughput detection of vitamin D metabolites.

Description

technical field [0001] The invention relates to the technical field of analysis and detection, in particular to a combined detection method for multiple vitamin D metabolites, a detection kit and applications thereof. Background technique [0002] Vitamin D (vitaminD) is a fat-soluble, steroidal vitamin necessary for the human body. Vitamin D2 (ergocalciferol) and vitamin D3 (cholecalciferol) are the most important members of the vitamin D family and have a wide range of physiological and metabolic activities. . Vitamin D is metabolized by the liver and kidneys to produce a series of metabolites, including 25-hydroxyvitamin D3, 25-hydroxyvitamin D 2 , 1,25 dihydroxyvitamin D 3 , 1,25 dihydroxyvitamin D 2 , 24,25 dihydroxyvitamin D 3 , 24,25 dihydroxyvitamin D 2 Wait. [0003] The content of vitamin D metabolites in the body is relatively low, and different metabolites are relatively similar in structure and properties, and there are different isomers, which makes it di...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N30/02G01N30/06G01N30/34G01N30/36G01N30/72
CPCG01N30/02G01N30/06G01N30/34G01N30/36G01N30/7266
Inventor 邱玲王丹晨禹松林张琪
Owner PEKING UNION MEDICAL COLLEGE HOSPITAL CHINESE ACAD OF MEDICAL SCI
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