Figure 4.

Relative quantification of nitrated rat brain peptides using dimethylation with heavy and light formaldehyde. Results from experiments comparing the relative quantities of nitrated peptides extracted from undiluted vs. diluted samples of nitrated rat brain homogenate. Panel (A) depicts the observed vs. expected ratios in each dilution group of the 10 most abundant nitrated peptides. Each observed ratio value is the mean value from three replicate experiments. The average observed ratios are 0.502 for the 1:1 dilution, 0.202 for the 1:4 dilution, 0.096 for the 1:9 dilution, and 0.058 for the 1:19 dilution. In Panel (B), the chromatograms from the purified heavy and light versions of the nitrated rat brain peptide GHyTEGAELVDSVLDVVR are shown. As this peptide is triply charged and contains just one dimethylation site (the N-terminus), the m/z difference between the heavy and light peptides is +2 (6 Da/3 charges). Though these equivalent peptides co-elute, they clearly separate from one another on the m/z axis.