Determination of Sub-Nanomolar Levels of Low Molecular
Mass Thiols in Natural Waters by Liquid Chromatography Tandem Mass
Spectrometry after Derivatization with <i>p</i>‑(Hydroxymercuri)
Benzoate and Online Preconcentration
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Abstract
Low molecular mass (LMM) thiols is
a diverse group of compounds,
which play several important roles in aquatic ecosystems, even though
they typically occur at low concentrations. Comprehensive studies
of LMM thiols in natural waters have so far been hampered by selectivity
and limit of detection constraints of previous analytical methods.
Here, we describe a selective and robust method for the quantification
of 16 LMM thiols in natural waters. Thiols were derivatized with 4-(hydroxymercuri)benzoate
(PHMB) and preconcentrated online by solid-phase extraction (SPE)
before separation by liquid chromatography and determination by electrospray
ionization tandem mass spectrometry (LC-ESI-MS/MS). Their quantification
was performed by selective reaction monitoring (SRM), while the presence
of a product ion at <i>m</i>/<i>z</i> 355, specific
for thiols and common for the investigated compounds, also allows
to screen samples for unknown thiols by a precursor ion scan approach.
The robustness of the method was validated for aqueous matrices with
different pH, sulfide, and dissolved organic carbon (DOC) concentrations.
The limits of detection for the thiols were in the sub-nanomolar range
(0.06–0.5 nM) and the methodology allowed determination of
both reduced and total thiol concentrations (using tris(2-carboxyethyl)phosphine
(TCEP) as reducing agent). Six thiols (mercaptoacetic acid, cysteine,
homocysteine, <i>N</i>-acetyl-cysteine, mercaptoethane-sulfonate,
and glutathione) were detected with total concentrations of 7–153
nM in boreal lake or wetland pore waters while four thiols (mercaptoacetic
acid, cysteine, homocysteine, and <i>N</i>-acetyl-cysteine)
were detected in their reduced form at concentrations of 5–80
nM