The diverse array
of chemical compounds present in tissue samples
results in many isobaric (i.e., same nominal mass) compounds in imaging
mass spectrometry experiments. Adequate separation and differentiation
of these compounds is necessary to ensure accurate analyte identification
and avoid composite images comprising multiple compounds. High-resolution
accurate mass (HRAM) measurements are able to resolve these compounds
in some instances, but HRAM measurements are not always feasible depending
on the instrument platform and the desired experimental time scale.
Alternatively, tandem mass spectrometry (MS/MS) can be used to perform
gas-phase transformations that improve molecular specificity. While
conventional MS/MS methods employ collision induced dissociation (CID)
to fragment compounds of interest and then analyze the product masses,
gas-phase ion/ion reactions can be used to instead selectively react
with desired classes of analytes. Herein, we have used gas-phase charge
inversion ion/ion reactions to selectively resolve phosphatidylcholines
(PCs) in isobaric lipid mixtures. A 1,4-phenylenedipropionic acid
(PDPA) reagent dianion readily reacts with [M + H]+, [M
+ Na]+, and [M + K]+ ion types to produce demethylated
product anions for each PC, [PC – CH3]−. These product anions are no longer isobaric and now differ in mass
by 22 Da (protonated versus sodiated) and 16 Da (sodiated versus potassiated),
respectively. This reaction has been used to differentiate isobaric
lipids in the imaging mass spectrometry analysis of rat brain tissue.