Lc ms grade water

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.

We extracted 150 mg of frozen powder in 450 µL of LC-MS grade methanol (Honeywell, Seezle, Germany). The mixture was vortexed for 30 s and sonicated at 40 kHz in a Sonica Ultrasonic Cleaner ultrasonic bath (SOLTEC, Milano, Italy) for 10 min before two rounds of centrifugation at 14,000× g for 15 min each. For the untargeted metabolomics analysis, the methanol extracts were diluted 1:3 (v/v) with LC-MS grade water (Honeywell), passed through Minisart RC4 filters with 0.2 µm pores (Sartorius, Göttingen, Germany) and 5 µL was injected into the UPLC device. For targeted metabolomics analysis, tryptamine and serotonin were quantified by diluting the samples 1:20 (v/v) with LC-MS grade water including the authentic commercial standards d₄-tryptamine and d₄-serotonin (Sigma, Darmstadt, Germany). The hydro-alcoholic mixtures containing the deuterated molecules at final concentrations of 5 pg/µL were passed through Minisart RC4 filters (0.2 µm pores) and 1 µL was injected into the UPLC device.