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.