A simplified LC-MS-based method for sensitive analysis of DNA adducts utilizing accessible in vitro metabolism models

Area of Science:

• Biochemistry
• Toxicology
• Analytical Chemistry

Background:

• DNA modifications significantly impact cellular function and integrity.
• Current DNA adduct analysis methods lack structural information and are often complex.
• Identifying suitable biological systems for reliable DNA adduct formation is challenging.

Purpose of the Study:

• To develop a fast, simple, and sensitive workflow for DNA adduct analysis.
• To compare different biological systems for their metabolic activation capabilities.
• To provide structural insights into DNA adducts using instrumental techniques.

Main Methods:

• Utilized HepG2 cells and liver S9 fractions for metabolic activation of carcinogens.
• Employed high-performance liquid chromatography coupled to mass spectrometry (HPLC-MS) for adduct detection.
• Compared biological systems using six known carcinogens and an aryl hydrocarbon receptor agonist.

Main Results:

• HepG2 cells and liver S9 fractions showed effective metabolic activation for DNA adduct formation.
• Successfully detected DNA adducts from aflatoxin B1, benzo[a]pyrene, methyleugenol, α-asarone, and β-asarone.
• Documented phenylpropanoid adduct formation in a non-transfected cancer cell line for the first time.

Conclusions:

• The developed workflow is efficient for sensitive DNA adduct analysis.
• HepG2 cells and S9 fractions are suitable biological systems for studying DNA adducts.
• The method enables the detection of novel DNA adducts and provides valuable structural information.