I’m currently working on LC-QTOF-MS/MS profiling of phenolic compounds, and like you, I detect some metabolites only in ESI−, some only in ESI+, and a few in both modes. I understand that ionization efficiency and detector response vary significantly between modes, so I calculated the % peak area separately for each mode, using:

% Peak Area = (compound response / total response in that mode) × 100

However, I still wonder, is this the correct approach, or would it be acceptable to combine response values from both modes and calculate % peak area from the total (ESI+ + ESI−)? Or is that not recommended due to the different ionization behavior?

When preparing data for heatmaps or multivariate analysis, if a compound was detected in both modes, I selected only one (usually the stronger or more consistent signal) to represent that compound. So the heatmap shows one value per compound, but the underlying % peak area was still calculated within its original mode.

I’d appreciate any insight on:

Whether combining modes during peak area calculation is valid in any cases

If selecting one mode per compound for visualizations like heatmaps is an acceptable simplification

Or whether mode-specific data should always remain completely separate, even for untargeted studies

Looking forward to hearing how others handle this thanks again!