Fig. 2.

Characterization of LNO₂ by ESI triple-quadrupole MS. (A) Synthetic LNO₂ positional isomers were separated by HPLC and detected by MS/MS in the MRM mode by monitoring the 324/277 collision-induced dissociation transition (graph 1). No other peaks were detected in the chromatogram. The same method was used to resolve LNO₂ isomers present in the total lipid extract from 1 ml of packed red blood cells (graph 2) and plasma (graph 3). (B) EPI analysis of Peak 1, revealing fragments unique to the C12 positional isomer of LNO₂, m/z = 196 and m/z =157, plus fragments common to all LNO₂ isomers (m/z = 233, 244, 277, 293, and 306) (graph 1). EPI analyses of Peak 1 for red cell (graph 2) and plasma (graph 3) lipid extracts gave similar fragmentation patterns. All identifying fragments from graph 1 are present in graphs 2 and 3. (C). In addition to the common fragment ions of LNO₂, peak 2 displayed unique fragments m/z = 228 and m/z = 168 (graph 1). These fragments, particularly m/z = 228, indicate nitration of the C10 of LNO₂. EPI spectra from Peak 2 of resolved red cell (graph 2) and plasma (graph 3) lipid extracts gave fragmentation patterns similar to the synthetic standard.