Esi q tof ms ms 6520

Tissue samples (50–70 mg) were homogenized using 20 volumes of cold methanol as previously described [14 (link)]. Metabolite extracts were subjected to mass spectrometry using an HPLC 1290 series coupled to an ESI-Q-TOF MS/MS 6520 (Agilent Technologies, Santa Clara, CA, USA) as previously described [14 (link)]. Multivariate statistic analyses were performed using Metaboanalyst platform [19 ]. The preliminary identification of differential metabolites (Student’s T-Test, Benjamini Hochberg False Discovery Rate, p<0.05) was performed using the PCDL database from Agilent (Agilent Technologies, Barcelona, Spain), which accounts retention times in a standardized chromatographic system, exact mass and isotope distribution as an orthogonal searchable parameters to complement accurate mass data (AMRT approach) according to previously published works [14 (link)]. The version of the PCDL database used had retention times and accurate mass data for 679 compounds. To complete the identification process we searched for unidentified metabolites in Metlin Database (https://metlin.scripps.edu/index.php) which includes accurate masses and MS/MS spectrum for 961.829 molecules.