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

Area of Science:

• Analytical Chemistry
• Mass Spectrometry

Background:

• Accurate mass measurements are critical for identifying fragment ions in high-resolution mass spectrometry.
• Perfluoroalkyl sulfonic acids (PFSAs) analysis on Q Exactive Orbitrap instruments showed systematic mass errors for low-mass ions (below m/z 100).

Purpose of the Study:

• To investigate the cause of systematic mass deviations observed for low-mass ions in Q Exactive Orbitrap instruments.
• To develop and validate a method for correcting these mass accuracy errors.

Main Methods:

• Comparative analysis of perfluoroalkyl sulfonic acids (PFSAs) using Q Exactive Orbitrap and Exploris 120 mass spectrometers.
• Evaluation of standard calibration solutions versus extended calibration with in-situ generated low-mass anchors.

Main Results:

• A systematic deviation of approximately -6 ppm was observed for the O3S- fragment (m/z 79.9574) on the Q Exactive, while higher m/z ions were accurate.
• The deviation was traced to the limited calibration range of the standard Q Exactive calibration solution, not an intrinsic ion property.
• Extending calibration with additional low-m/z anchors generated in-situ fully corrected the systematic errors for ions below m/z 100.

Conclusions:

• The limited calibration range of standard solutions is the source of systematic mass errors for low-m/z ions in Q Exactive instruments.
• In-situ generation of low-m/z calibration anchors effectively resolves these errors without compromising accuracy at higher m/z.
• This calibration strategy enhances the reliability of high-resolution mass spectrometry analysis for compounds with low-mass fragments.