| Description
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Extraction of metabolites
The freeze-dried samples were extracted with 50 μL of 80 % MeOH containing 2.5 μM 10-camphorsulfonic acid, 1.25 μM reserpine, 2.5 μM ampicillin, and 2.5 μM CHAPS as internal standards for calibration per mg dry weight using a mixer mill (MM300, Retsch) with zirconia beads for 7 min at 18 Hz and 4 °C. After 10 min of centrifugation, the supernatant was filtered with an HLB μElution plate (Waters). Extracts were routinely diluted with 80 % MeOH at a ratio of 1:1000 (extract: 80 % MeOH).
Direct infusion analysis
Ultra-high-resolution metabolome data were acquired using an FTICR-MS solariX 7.0 T (Bruker Daltonics) with an ESI source. The following settings were applied: MS Detection; Acquisition Mass Control (mass range, m/z 100–800; estimated resolution power, 530,000 at m/z 400; transient length, 3.9147 s); Data Storage (save full profile spectrum, on; save FID file, on); Accumulation (scan time, 100; source accumulation, 0.01 s; ion accumulation time, 0.1 s; ion cooling time, 0 s; time of flight, 0.45 s); API Source (source type, ESI; capillary, 4500 V; end plate offset, 1000 V); Source Gas Tune (nebulizer 1.0 bar; dry gas, 2.0 L/min; dry temperature, 200 °C); Source Optics (capillary exit, −220 V; deflector plate, −200 V; funnel 1, −150 V; skimmer 1, −25 V; funnel RF amplitude, 180 Vpp); Octopole (frequency, 5 MHz; RF amplitude, 350 Vpp); Collision Cell (collision voltage, 0.8 V; DC extract bias, −0.9 V; RF frequency, 2 MHz; collision RF amplitude, 850 Vpp); Transfer Optics (time of flight, 0.45 ms; frequency, 6 MHz; RF amplitude, 400 Vpp); Infinity Cell (transient exit lens, 20 V; analyzer entrance, 10 V; side kick, 0 V; side kick offset, 1.5 V; front trap plate, −0.5 V; back trap plate, −0.5 V; sweep excitation power, 20 %); Multiple Cell Accumulations (number of ICR cell fills, 1); Gated Trapping (gated trapping mode, off); Polarity, negative. MS spectra were internally calibrated using the MS spectra of internal standards.
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