Agilent masshunter profinder

The GC-MS data were processed using an in-house MATLAB script, R2016a (The MathWorks, Inc., Natick, MA, USA). The LC-MS data were processed using Agilent Masshunter Profinder version B.08.00 (Agilent Technologies Inc., Santa Clara, CA, USA). Putative metabolites were extracted using unique mass channels, retention indices (GC-MS data), MS-MS spectra (LC-MS dat) and matched to our in-house mass spectral library at the Swedish Metabolomics Centre (www.swedishmetabolomicscentre.se, 10 October 2021). Labelled internal standards were used for alignment and normalization. Extensive filtering was performed to remove noise and entities with poor quality (e.g., suffering from peak broadening, ion suppression, or non-Gaussian peaks), and only unique spectral profiles with a relative standard deviation (RSD) < 40% calculated from QC samples were included in sample comparison modelling. Criteria set by the Human Metabolome Database (www.HMDB.ca, 15 August 2021) were used to assign extracted components to different compound classes (e.g., amino acids and derivatives, BCAAs, carbohydrates, lipid subtypes, or no class). All lipids were annotated according to standard lipid nomenclature set by Lipid Maps Lipidomics Gateway (lipidmaps.org).

To illustrate the benefit of counting with an extensive lipid database of a particular biological matrix, we investigated the lipid alterations induced by the Mtb infection at two different time points (4 weeks, Mtb⁺4w and 12 weeks, Mtb⁺12w) post-infection. The previously obtained database, including molecular formulas, mass, and RT information, was imported into the Agilent MassHunter Profinder software (B.10.0.2, Agilent Technologies, Santa Clara, CA, USA) using the Batch Targeted Feature Extraction mode to perform the feature extraction and time alignment. Features were built as the sum of coeluting ions that are related by charge-state envelope, isotopologue pattern, and/or the presence of different adducts and dimers in the analyzed samples. To detect coeluting adducts of the same feature, the following adducts were selected: [M + H]⁺, [M+Na]⁺, [M + K]⁺, [M + NH₄]⁺ and [M + C₂H₆N₂ + H]⁺ in LC-ESI(+)-MS; [M-H]⁻, [M+Cl]⁻, [M + CH₃COOH-H]⁻, and [M + CH₃COONa-H]⁻ in LC-ESI(-)-MS.