Extending Calibration Anchors Resolves Low-m/z Shifts in Negative-Mode Orbitrap Workflows

Reliable mass accuracy is essential for the confident identification of diagnostic fragment ions in high-resolution mass spectrometry. During the analysis of perfluoroalkyl sulfonic acids (PFSAs) with a Q Exactive Orbitrap, we consistently observed the characteristic O₃Ṡ^- fragment, expected at a mass-to-charge ratio (m/z) of 79.9574, reported at m/z 79.9568, a systematic deviation of approximately -6 parts per million (ppm). Similar errors affected other ions below m/z 100, while precursors and higher-m/z fragments remained within the specification. Comparison with an Exploris 120 instrument, calibrated with anchors down to m/z 59, confirmed that the deviation is not intrinsic to the ion but originates from the limited calibration range of the Q Exactive standard calibration solution. We demonstrate that extending calibration to include additional low-m/z anchors generated in situ by in-source fragmentation fully corrects the error without affecting the accuracy at a higher m/z. This adjustment resolves systematic deviations for ions below m/z 100 in the Q Exactive instruments.