Estimation of Reactive Thiol Concentrations in Dissolved Organic Matter and Bacterial Cell Membranes in Aquatic Systems

Abstract

Organic thiols are highly reactive ligands and play an important role in the speciation of several metals and organic pollutants in the environment. Although small thiols can be isolated and their concentrations can be estimated using chromatographic and derivatization techniques, estimating concentrations of thiols associated with biomacromolecules and humic substances has been difficult. Here we present a fluorescence-spectroscopy-based method for estimating thiol concentrations in biomacromolecules and cell membranes using one of the soluble bromobimanes, monobromo(trimethylammonio)bimane (qBBr). The fluorescence of this molecule increases significantly when it binds to a thiol. The change in the sample fluorescence due to thiols reacting with qBBr is used to determine thiol concentration in a sample. Using this method, small thiols such as cysteine and glutathione can be detected in clean solutions down to ~50 nM without their separation and prior concentration. Thiols associated with dissolved organic matter (DOM) can be detected down to low micromolar concentration, depending on the DOM background fluorescence. The charge on qBBr prevents its rapid diffusion across cell membranes, so qBBr is ideal for estimating thiol concentration at the cell membrane-water interface. This method was successfully used to determine the thiol concentration on the cell envelope of intact Bacillus subtilis to nanomolar concentration without any special sample preparation. Among the chemical species tested for potential interferences (other reduced sulfides methionine and cystine, carboxylate, salt (MgCl(2))), carboxylates significantly influenced the absolute fluorescence signal of the thiol-qBBr complex. However, this does not affect the detection of thiols in heterogeneous mixtures using the presented method.

Characterization of dissolved organic matter from a restored urban marsh and its role in the mobilization of trace metals

“…Organic thiol functional groups are important DOM complexing ligands for soft Lewis acids, such as Cd and Pb. Their presence influences metal speciation and solubility in natural waters (Joe-Wong et al, 2012). Thiols affinity for metals like Cd and Pb would result in the higher measured binding capacity values observed in the measured complexation constants for natural waters (see Table 1).…”

“…One such functional group not taken into account by the MINTEQ model is the thiol group. These organic sulfide groups (R-SH) are ubiquitous in wetlands (Zhang and Wang, 2004) and are produced by plants mediating interactions with metals (Joe-Wong et al, 2012). Organic thiol functional groups are important DOM complexing ligands for soft Lewis acids, such as Cd and Pb.…”

Thiol-Based Selective Extraction Assay to Comparatively Assess Bioavailable Mercury in Sediments

“…Moreover, the reactivity of dissolved and particulate Hg sulfides with thiols correlated with differences in net methylation potential by methylating microorganisms (Zhang et al, 2012). Likewise, a number of recent studies point to thiols as the site for mercury binding to the cell envelope of methylating bacteria ( Joe-Wong et al, 2012;Hu et al, 2013;Dunham-Cheatham et al, 2014, 2015. While the interactions between Hg and thiols in these studies and the mechanism of Hg uptake by the cell remain unknown, we hypothesize that a simple thiolbased selective extraction could be used to approximate the bioavailable fraction of Hg in anaerobic settings.…”

The effect of natural organic matter on the adsorption of mercury to bacterial cells

“…Heavy metals, such as Hg, adsorb to proton-active functional groups on bacterial cell envelopes (e.g., Beveridge and Murray, 1976;Fortin and Beveridge, 1997;Daughney et al, 2002;Fein, 2006;Kenney and Fein, 2011), affecting the speciation and distribution of these metals in environmental systems. Recent studies (e.g., Guiné et al, 2006;Mishra et al 2007;2009;2010;Joe-Wong et al, 2012;Pokrovsky et al, 2012;Song et al, 2012;Colombo et al, 2013;Yu et al, 2014) have shown that at least some bacterial cell envelopes contain protonactive sulfhydryl functional groups. Because Hg binds readily and strongly to sulfur compounds (Compeau and Bartha, 1987;Winfrey and Rudd, 1990;Benoit et al, 1999), bacterial adsorption of Hg may dramatically affect the distribution, transport and fate of Hg in geologic systems.…”