Unveiling the Power of Negative Ion Mode ESI-MS: Identifying Species with Remarkable Signal Intensity and Collisional Stability

26 September 2023, Version 2
This content is a preprint and has not undergone peer review at the time of posting.


Electrospray ionization mass spectrometry has long been the standard and most prevalent ionization method in mass spectrometry to detect and analyze molecules of low volatility that are relevant biologically, environmentally, and industrially. However, only a small number of analyses are conducted in negative ion mode, which has led to a dogmatic bias toward positive ion mode despite advantageous properties of the negative polarity, including lower back-ground noise and divergent tandem mass spectrometry behavior. We hypothesized that this bias was rooted in the relatively poor ionization efficiency of anionic functional groups seen in biochemistry; to explore this notion herein we evaluated 25 ions based on three criteria: (1) signal intensity relative to a sodium dodecylsulfate internal standard; (2) resistance to collision induced dissociation based on survival of the precursor ion; and (3) diagnostic tandem mass spectrometry behavior. Among these species, highly fluorous ions exhibiting weakly coordinating and hydro-phobic properties contributed to enhanced signal intensities. Trifluoromethanesulfonyl-containing ions proved to be unexpectedly labile, while tetrakis[3,5-bis(trifluoromethyl)phenyl]borate anion (23) and bis(nonafluoro-1-butane)sulfonimidate (25) were determined to be of optimal signal intensity with signal intensity ratios relative to sodium dodecylsulfate (12 + Na+) of 332.0% ± 25.0% and 939.0% ± 92.0%, respectively, as well as survival yields of 100.0% ± 0.0% and 72.6% ± 0.8% at –50 eV. To further emphasize their optimal signal intensity, ions tetrakis[3,5-bis(trifluoromethyl)phenyl]borate anion (23) and bis(nonafluoro-1-butane)sulfonimidate (25) were comparable in signal intensity across solvents of acetonitrile, methanol, isopropanol, water, and their respective 1:1 mixtures. Facile preparation of various salts of bis(nonafluoro-1-butane)sulfonimidate led to additional evaluation of cation effects where the signal intensity ratio ranged from 939.0% ± 92.0% to 3195.0% ± 145.0% across K+, NH4+, Na+, and H+ counter cations. The dogma of negative ion mode being less sensitive was then challenged by the analysis of the sig-nal intensity of ion 25 to tetra-n-butylammonium, tetra-n-butylphosphonium, and (4-methylphenyl)diphenylsulfonium cations. These experiments showed that 25 was more sensitive by between 136.2% ± 5.5% and 180.7% ± 13.8%, thereby successfully challenging the positive polarity bias.


negative ion mode
ionization efficiency
collision induced dissociation
survival yield

Supplementary materials

Negative Mode ESI Warnes Chihabi Manthorpe Supporting Info
Experimental procedures, spectral data


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