Agilent 7200 accurate mass q tof gc ms

The GC/MS-based metabolite measurements were performed as described by Fiehn, (2007) , using ribitol as an internal standard. Leaf samples were collected by rapid freeze-quenching from the custom gas exchange chamber describe above. Freezequenched tissue was ground into a fine powder in liquid nitrogen using a mortar and pestle. Extracted metabolites were injected into a gas chromatograph (Agilent 7890B
GC System, Agilent Technologies, USA) that was in line with a mass spectrometer (Agilent 7200 Accurate-Mass Q-TOF GC/MS, Agilent Technologies, USA). Metabolite peaks were evaluated using Mass Hunter Software (Agilent Technologies, UAS). The relative amount of each metabolite was calculated from the peak area, taking into account both the initial fresh weight used for extraction and the internal standard.

All solvents were purified by standard procedures. Chemicals were obtained from commercial sources (Merck KGaA, Darmstadt, Germany) and were used without further purification. 1 H, 13 C, and 19 F NMR spectra were recorded on Bruker spectrometers (Bruker Corporation, Billerica, Massachusetts, USA) Avance-300 (300.13, 282.37, and 75.47 MHz for 1 H, 19 F, and 13 C, respectively), Avance-400 (400.13 and 100.61 MHz for 1 H and 13 C, respectively), DRX-500 and Avance III 500 (500. 13 MHz for 1 For 19 F spectra, C 6 F 6 (δ 19 F = -162.9 ppm with respect to CFCl 3 ) was used as an external reference. 13 C NMR spectra were registered with C-H spin decoupling. Masses of molecular ions were determined by high-resolution mass spectrometry (HRMS) on a DFS Thermo Scientific instrument at an ionization energy of 70 eV (Thermo Fisher Scientific, Waltham, MA, USA) and by means of Agilent 7200 Accurate Mass Q-TOF GC/MS at an ionization energy of 70 eV (Agilent, Santa Clara, CA, USA). Melting points were registered on a Mettler-Toledo FP900 Thermosystem apparatus (Mettler-Toledo, Greifensee, Switzerland). IR spectra were recorded on a Bruker Vector-22 spectrometer for samples pelleted with KBr (0.25%). UV/Vis spectra were acquired by means of a Cary 5000 instrument (Agilent).