Comparison of Lysis and Detachment Sample Preparation Methods for Cultured Triple-Negative Breast Cancer Cells Using UHPLC–HRMS-Based Metabolomics

Abstract

Dysregulation of cellular metabolism is now a well-recognized hallmark of cancer. Studies investigating the metabolic features of cancer cells have shed new light onto processes in cancer cell biology and have identified many potential novel treatment options. The advancement of mass spectrometry-based metabolomics has improved the ability to monitor multiple metabolic pathways simultaneously in various experimental settings. However, questions still remain as to how certain steps in the metabolite extraction process affect the metabolic profiles of cancer cells. Here, we use ultra-high-performance liquid chromatography–high-resolution mass spectrometry (UHPLC–HRMS) untargeted metabolomics to investigate the effects of different detachment and lysis methods on the types and abundances of metabolites extracted from MDA-MB-231 cells through the use of in-house standards libraries and pathway analysis software. Results indicate that detachment methods (trypsinization vs. scraping) had the greatest effect on metabolic profiles whereas lysis methods (homogenizer beads vs. freeze–thaw cycling) had a lesser, though still significant, effect. No singular method was clearly superior over others, with certain metabolite classes giving higher abundances or lower variation for each detachment–lysis combination. These results indicate the importance of carefully selecting sample preparation methods for cell-based metabolomics to optimize the extraction performance for certain compound classes.

Keywords

• cell metabolomics
• mass spectrometry
• breast cancer
• pathway analysis
• detachment
• lysis