Metabolite Profiling by UPLC-MS

For the UPLC-MS analysis, the methanol extracts were diluted 1:1 (v/v) with LC-MS grade water (Honeywell, Seezle, Germany), whereas a sample of concentrated wine, resuspended in 1 mL of LC-MS grade water, was diluted 1:10 (v/v). The diluted samples were passed through Minisart RC4 filters with 0.2 µm pores (Sartorius, Göttingen, Germany), and then 1, 3, and 5 µL of the cell extracts and 1 µL of the resuspended wine were injected into the UPLC-MS system. The analysis was performed with an ACQUITY I CLASS UPLC system (Waters Corporation, Milford, Massachusetts ), connected to a Xevo G2-XS qTOF mass spectrometer (Waters) equipped with an electrospray ionization (ESI) source operating in either positive or negative ionization modes. The chromatographic conditions and mass spectrometer parameters were set as described in Commisso et al. [40 (link)]. The raw data were processed with Progenesys QI (Nonlinear dynamics, UK). The metabolite identification was performed using an “in house” library of mass spectra, through the m/z value, and, where possible, isotopic similarity and fragmentation patterns.
The data matrix obtained by using Progenesys QI was submitted to principal component analysis (PCA) through SIMCA 13.0 (Umetrics, Sartorius, Gottingen, Germany), after Pareto scaling and centering.