Xcalibur 4.0 qual browser

Raw files generated by the Q Exactive HF instrument were analysed using the Compound Discoverer Software 3.1 (Thermo Fisher Scientific). Identified compounds were manually reviewed using Xcalibur 4.0 Qual browser and Freestyle (version 1.3.115.19) (both Thermo Fisher Scientific) and the obtained MS2 spectra were compared to reference spectra, which were retrieved from mzcloud (Copyright © 2013–2020 HighChem LLC, Slovakia). The match factor cut-off from mzcould was 80, while the mass tolerances were 5 and 10 ppm on MS1 and MS2 levels, respectively. Moreover, the identity of compounds suggested by Compound Discoverer was verified by analysing purchased standards using the same LC-MS method. The Tracefinder Software 4.1 (Thermo Fisher Scientific) was used for peak integration and calculation of peak areas. The generated batch table was exported and further processed with Microsoft Excel (version 1808), GraphPad Prism (version 6.07) and the Perseus software (version 1.6.12.0)^{70 (link)}, the letter being used for the principal component analysis. Untargeted metabolic profiling by mass spectrometry delivered more than 50000 reproducible sweat-specific features per analysis. Microsoft PowerPoint (version 1808) was used for creating figures.

Raw files generated by the Q Exactive HF were searched in the Compound Discoverer Software 3.1 (Thermo Fisher Scientific) applying a user-defined workflow. All identified compounds with a match factor ≥ 80 were manually reviewed using Xcalibur 4.0 Qual browser (Thermo Fisher Scientific) and the obtained MS2 spectra were compared to reference spectra taken from mzcloud (Copyright © 2013–2020 HighChem LLC, Slovakia). A maximum retention time shift of 0.1 min was allowed and the mass tolerance was restricted to 10 ppm for MS1 and MS2. Identified compounds were also verified using purchased analytical standards applying the same LC–MS method. The Tracefinder Software 4.1 (Thermo Fisher Scientific) was used for peak integration and calculation of peak areas. Generated batch tables were exported and further process with Microsoft Excel, GraphPad Prism (for two-tailed, paired t-tests) and the Perseus [45 (link)] (for principal component analysis) software. ACD/Labs’ ChemSketch (Freeware) 2020.1.1 was used to draw structures.