DESI/PI-MSI
The experimental procedure for DESI/PI-MSI was previously described (Liu et al., 2021; Liu et al., 2019). Samples were prepared as described before. The metabolite distribution was conducted on a modified Agilent 6224 Accurate-Mass TOF Mass Spectrometer (Agilent, USA). The data storage was synchronized with the activation of stepper motor for every line scanning by a custom-developed software based on LabVIEW (Bitter, 2006) For imaging, by using the open source ProteoWizard converters, the collected Agilent Chemstation.D files were converted to “.mzML” format files at first, which were then converted into one “.imzML” format file for image reconstruction with imzMLConverter (Race et al., 2012) All ion image reconstructions, background subtraction and the generation of an average mass spectrum in a region of the sections were performed using the freely available standalone version of the MSiReader software with the following parameters: X and Y pixel size 200 μm; raster speed 370 μm/s; spray solvent 70%. MeOH, 30% Toluene, 1% Formic acid, and flow rate at 3 μl/min; MS at positive polarity, 3.5 kV capillary voltage, 80 V cone voltage, 1.2MPa pressure of nitrogen as carrier gas, and mass range m/z 20-1000. Imaging mass spectra of available standards (atractylodin & atractylone) are shown in Figure S6.