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Method for measuring six aldehyde-DNA adducts in saliva

A determination method and adduct technology, applied in the direction of measuring devices, instruments, scientific instruments, etc., to reduce consumption, improve selectivity, accuracy and sensitivity, and improve the effect of chromatographic separation process

Inactive Publication Date: 2016-06-01
CHINA NAT TOBACCO QUALITY SUPERVISION & TEST CENT
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Currently, high-throughput analysis methods for HOMe-dA, HOMe-dG, Ethylidene-dG, CdG, α-Acr-dG, and γ-Acr-dG in saliva have not been reported

Method used

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  • Method for measuring six aldehyde-DNA adducts in saliva
  • Method for measuring six aldehyde-DNA adducts in saliva
  • Method for measuring six aldehyde-DNA adducts in saliva

Examples

Experimental program
Comparison scheme
Effect test

example 1

[0036] 1. Instruments and reagents:

[0037] ABSSCIEX triple quadrupole tandem mass spectrometer (Applied Biosystems, California, USA), Agilent1200 high performance liquid chromatography (Agilent, USA), NanoDrop2000 ultra-trace spectrophotometer (Thermo Scientific, USA), constant temperature incubator (Thermo Scientific, USA) Flying Technology Co., Ltd.), nitrogen concentration dryer (Sweden Baitaqi company), Milli-Q water purification system (Germany Merck Group), Analyst1.5.1 data acquisition and processing software.

[0038] Deoxyribonuclease I, alkaline phosphatase (New England Biotechnology, USA), phosphodiesterase I, ammonium acetate, formic acid, and calf thymus DNA (Sigma, USA), NaBH 3 CN (Shanghai Aladdin Reagent Co., Ltd.), methanol (Toksan Reagent Co., Ltd., Korea), Strata-X polymer cartridge (33μm, 30mg / 1mL, Guangdong Philomena Scientific Instrument Co., Ltd.), Oragene DNA saliva collector (OG -500, DNAGenotek, Canada). Standards Me-dA, Me-dG, Et-dG, CdG, α-Acr-d...

example 2

[0044] Example 2: As described in Example 1, aldehyde-DNA adducts were detected in 5 saliva samples using the method established in this study (each sample was detected in parallel three times). Calculate the concentration of DNA adducts in each sample based on the standard curve and peak area, according to the formula

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Abstract

The invention discloses a method for measuring six aldehyde-DNA adducts in saliva, namely a method for measuring HOMe-dA, HOMe-dG, Ethylidene-dG, CdG, alpha-Acr-dG and gamma-Acr-dG in saliva. The method comprises the following steps: collecting saliva by an OG-500 saliva collecting tube, performing DNA extraction on the saliva, performing enzyme hydrolysis on a DNA solution, and sequentially adding stable isotope internal standards, a reducing agent, NaBH3CN and DNA hydrolase; and performing solid-phase extraction on hydrolysate via Strata-X columella, collecting an eluant, nitrogen-blowing at room temperature till the product is dried, re-dissolving into a water solution of methyl alcohol, and introducing LC-MS / MS system analysis, thereby accurately detecting the content levels of the six substances. With stable isotope as internal standard quantitative analyte, errors caused in pretreatment of samples can be reduced, and the selectivity, accuracy and sensitivity of the method are better improved by LC-MS / MS. By selecting and optimizing chromatographic columns and elution conditions, the chromatographic separation process is better improved, the time for analysis is shortened, and the consumption of a solvent is reduced.

Description

technical field [0001] The invention belongs to the technical field of physical and chemical testing of saliva samples, and mainly relates to N in saliva 6 -Hydroxymethyl-adenine (HOMe-dA), N 2 -Hydroxymethyl-guanine (HOMe-dG), N 2 -Ethylidene-dG, 1,N 2 -Propanol-guanine (CdG), 6-hydroxy-1,N 2 -Propanol-2'-guanine (α-Acr-dG) and 8-hydroxy-1,N 2 - The technical field of determination of propanol-2'-guanine (γ-Acr-dG) DNA adducts, specifically a liquid chromatography-tandem mass spectrometer (LC-MS / MS) method for the determination of six species in saliva Methods for Aldehyde-DNA Adducts. Background technique [0002] Formaldehyde, acetaldehyde, acrolein, and crotonaldehyde are a class of pollutants that are widely present in environmental matrices. They are mainly produced from the incomplete combustion of organic matter, such as industrial production, cigarette smoke, motor vehicle exhaust and cooking fumes. Active aldehyde groups can directly attack nucleophilic grou...

Claims

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Application Information

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IPC IPC(8): G01N30/88
CPCG01N30/88G01N2030/8813
Inventor 侯宏卫胡清源陈欢刘鲁娟陈建张小涛张森
Owner CHINA NAT TOBACCO QUALITY SUPERVISION & TEST CENT
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