Enhancing Chimeric Fragmentation Spectra Deconvolution Using Direct Infusion-Tandem Mass Spectrometry Across...

Area of Science:

• Analytical Chemistry
• Mass Spectrometry
• Proteomics

Background:

• Direct infusion mass spectrometry (DI-MS) is rapid but suffers from co-fragmentation, creating complex chimeric spectra.
• Compound identification is challenging due to overlapping signals in DI-MS.
• DI-MS2, a DI-based tandem MS method, modulates precursor intensity to deconvolve chimeric spectra.

Purpose of the Study:

• Evaluate DI-MS2 performance on different high-resolution mass spectrometry platforms.
• Assess the impact of instrumental settings on DI-MS2 deconvolution accuracy.
• Provide optimization guidelines for DI-MS2.

Main Methods:

• Tested DI-MS2 on linear ion trap-Orbitrap (LIT-Orbitrap) and quadrupole-Orbitrap (Q-Orbitrap) instruments.
• Investigated six instrumental parameters: resolving power, isolation window, step size, microscans, collision energy, and AGC target.
• Analyzed isobaric mixtures with varying m/z differences.

Main Results:

• LIT-Orbitrap achieved consistent high-quality deconvolution (average similarity score 0.98).
• Q-Orbitrap offered faster measurements (4x) but showed variable results (similarity score 0.96 for m/z diff > 0.02, 0.56 for m/z diff = 0.006).
• DI-MS2 performance is dependent on m/z difference for Q-Orbitrap.

Conclusions:

• DI-MS2 is a robust and flexible method applicable across different mass spectrometry platforms.
• Q-Orbitrap is less suitable for highly complex samples with minimal m/z differences.
• DI-MS2 shows potential for structural elucidation in complex biological mixtures.
• Optimization guidelines are provided to enhance deconvolution and speed.