A Comparative Study of Pentafluorophenyl and Octadecylsilane Columns in High-throughput Profiling of Biological Fluids

In high-throughput metabolomic profiling, chromatographic separation is crucial because a well-performed chromatographic separation may reduce signal suppression from complex biological matrices and improve the discoverability of low-abundance metabolites. We compared the performance of pentafluorophenyl (PFP)- and octadecylsilane (ODS)-based columns in profiling biological fluids. Peak resolutions and consistencies were acquired using several reversed-phase columns and were evaluated. Total and extracted ion chromatograms demonstrated that the PFP column achieved better analyte separations than the ODS column. Low relative standard deviations on peak areas and retention times (<10.2 and <0.9%, respectively) acquired using the PFP column evidenced the high reproducibility and consistency of this column. In our study, a PFP column was used for profiling metabolomes extracted from urine and serum samples. Metabolomic study revealed a metabolome difference in normal and overweight participants. In total, 26 lipid species were significantly perturbed and further identified. Choline-containing lipids were the most abundant perturbed lipidome in overweight participants, followed by sphingolipids and various phospholipids. We recommend the use of PFP columns in high-throughput metabolomic analysis to promote the development of basic biological and clinical research in the future.