Enhancing the Power of
Liquid Chromatography–Mass
Spectrometry-Based Urine Metabolomics in Negative Ion Mode by Optimization
of the Additive
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Abstract
Untargeted liquid chromatography–mass spectrometry
(LC-MS)-based
metabolomics studies are usually carried out in both positive and
negative ion modes; however, it is frequently ignored that the optimal
conditions in positive ion mode and negative ion mode are often not
the same. We carried out a systematic investigation on urine samples
to evaluate the additive effects in negative ion mode. It was found
that the widely used conditions, 0.1% formic acid (FA) and NH<sub>4</sub>Ac at different pH, are far from the optimum for untargeted
urine metabolomics studies. Compared to 0.1% FA, the use of 1 mM acetic
acid (HAc) resulted in almost three times as many detected peaks (401
vs 148) and around five times the size of the peak area (33.55 ×
10<sup>6</sup> vs 6.47 × 10<sup>6</sup>). The remarkable improvement
can be explained by two factors: (i) a significantly enhanced ionization
efficiency due to the combination of an appropriate pH at around 4.0–4.5,
the reducibility of H<sup>+</sup>, and the high gas-phase basicity
of Ac<sup>–</sup> and (ii) a reproducible LC separation due
to an acceptable buffering capacity. Our study revealed the importance
and necessity of additive optimization, which can be of benefit in
related metabolomics studies