GC-TOF-MS Metabolite Identification Protocol

Chroma TOF 4.3X software and the LECO-Fiehn Rtx5 database (LECO Corporation, Benton Harbor, MI, USA) were used for raw peak extraction, database line filtering and calibration, peak alignment, deconvolution analysis, peak identification, and integration of peak area. Both mass spectrum match and retention index match were considered during metabolite identification. Additionally, peaks detected in less than 50% of the QC samples or for which relative standard deviation (RSD) greater than 30% in the QC samples were removed [50 (link),51 (link)]. Metabolites separated by GC-TOF-MS were identified using LECO ChromaTOF 4.3X software and the LECO/Fiehn Rtx5 metabolite mass spectral library by matching the mass spectrum and retention index.