1 research outputs found
Simple LC-MS Method for Differentiation of Isobaric Phosphatidylserines and Phosphatidylcholines with Deuterated Mobile Phase Additives
Lipids from different classes sometimes
can exhibit the same exact
mass upon electrospray ionization; this presents an analytical challenge
in lipidomics. In the negative ionization mode, for example, this
can occur with phosphatidylcholines (PCs) and phosphatidylserines
(PSs), making them indistinguishable in the absence of fragmentation
data. PSs are found at low concentrations in biological samples, making
MS/MS spectra difficult to obtain. Moreover, while PCs and PSs are
distinguishable in the positive mode, PSs do not ionize as well as
PCs, and their ionization is suppressed by the PCs. Here, we show
that, in the negative ionization mode, substituting protiated LC-MS
additives with their deuterated forms provides a way to distinguish
PCs and PSs without chemical derivatization. The method described
leverages the differential ionization mechanism of PCs and PSs. PCs
are ionized via adduction with salts, whereas PSs ionize via hydrogen
abstraction. Substituting the salts used for LC-MS with their deuterated
form shifts the mass of PCs by the number of deuterium atoms in the
salt, while the mass of PSs remains the same. This comparative shift
enables their direct differentiation. We demonstrate that the use
of deuterated formate shifts the mass of PCs and provides a direct
method to distinguish PCs and PSs, even at biologically relevant low
concentrations. The utility of the method was established and validated
in the simultaneous analysis of PCs and PSs in lipid extracts from
isolated liver mitochondria in two different rat strains. Thirteen
low concentration PSs were identified that would otherwise not have
been distinguishable from low concentration PCs