Using E. coli to Generate Internal Standards for LC-MSbased Metabolomics (

Abstract

Metabolomics is a large scale study of all metabolites which are small molecules that are contained within cells, biofluid, tissues and organisms (1). Much progress in metabolomics has been made in recent years and metabolomics has been applied in multiple fields to make new discoveries and confirm hypotheses. However, substantial challenges in the field remain. Factors such as sample extraction efficiency, instrumentation status, or metabolite stability lead to variations of metabolomics quantification results. To reduce such unwanted variations and improve metabolomic analysis accuracy, internal standards (IS) are often used during sample preparations. Internal standards (IS) usually have similar chemical and physical properties to the analyte in real samples but are not expected to be present in real samples. IS behave similarly to experimental analytes during sample preparation and data acquisition and can be used to correct the variations of analyte signals. An internal standard can be a structural isomer of the analyte or a stable isotope-labeled analyte. This study demonstrates the utilization of stable isotope-labeled internal standards in a high-performance liquid chromatography coupled to mass spectrometry (HPLC-MS)-based metabolomics study. Stable isotope-labeled internal standards can be very costly; thus we sought an alternative method of IS production. In this research, Escherichia coli (E. coli) was utilized as a biological product generating system to produce a stable isotope labeled internal standard. This method which is an inexpensive, labor friendly, fast, and an effective way to generate a large amount of IS to be used in multiple metabolomics-based research projects.