Eden L. Rue

Complexation of iron(III) by natural organic ligands in the Central North Pacific as determined by a new competitive ligand equilibration/adsorptive cathodic stripping voltammetric method

A highly sensitive voltammetric technique was developed to examine Fe speciation in seawater. The technique involves adding an Fe(III)-complexing ligand, salicylaldoxime, which competitively equilibrates with inorganic and organic Fe(III) species in ambient seawater. The Fe(III)-salicylaldoxime complex then is measured by adsorptive cathodic stripping voltammetry (ACSV). This new method revealed that 99.97% of the dissolved Fe(III) in central North Pacific surface waters is chelated by natural organic ligands. The total concentration of Fe-binding ligands is approximately 2 nM, a value greatly in excess of ambient dissolved iron concentrations. The titration data can be modeled as consisting of two classes of Fe-binding ligands, a strong ligand class (L1) with an average surface-water concentration equal to 0.44 nM with a conditional stability constant KL1Fe′cond = 1.2 × 1013 M−1, and a weaker ligand class (L2) with an average concentration equal to 1.5 nM with KL2Fe′cond = 3.0 × 1011 M−1. The low concentration of dissolved Fe present in surface waters (~ 0.2 nM), coupled with the excess of strong Fe-chelators, results in extremely low equilibrium concentrations of dissolved inorganic iron, [Fe′] ≈ 0.07 pM. In the deeper waters there is a 2 nM excess of Fe-binding ligands with a stability constant similar to that of the L2 class of ligands observed in surface waters, resulting in dissolved Fe(III) existing primarily in the chelated form in deep waters as well. The stability constants of the natural ligands are comparable to the model ligands desferal, a siderophore, and the prosthetic heme group, protoporphyrin-IX. The high degree of organic complexation of iron makes it critically important to reevaluate our perceptions of the marine biogeochemistry of iron and the mechanisms by which biota can access this chelated Fe.

Collection and detection of natural iron-binding ligands from seawater

Abstract Iron (Fe) is an essential element for the biochemical and physiological functioning of terrestrial and oceanic organisms, including phytoplankton, which are responsible for the primary productivity in the worlds oceans. However, due to the low solubility of Fe in seawater, phytoplankton are often limited by their inability to incorporate enough Fe to allow for optimal growth rates in regions with dissolved Fe concentrations below 1 nM. It has been postulated that certain phytoplankton may produce compounds to facilitate the uptake of Fe from seawater to overcome this limitation. Dissolved Fe in the oceans is overwhelmingly complexed (>99%) by strong organic ligands that may control the uptake of Fe by microbiota; however, the identity, origin, and chemical characteristics of these organic chelates are largely unknown. Although it has been implied that some components of natural Fe-binding ligands are siderophores, no direct analyses of such compounds from natural seawater have been conducted. Here, we present a simple solid-phase extraction technique employing Biobeads SM-2 and Amberlite XAD-16 resins for concentrating naturally occurring dissolved iron-binding compounds from large volumes (>200 l) of seawater. Additionally, we report on the first successful determination of molecular weight size classes and preliminary iron-binding functional group characterization within those size classes for isolates collected from the surface and below the photic zone (150 m) in the central California coastal upwelling system. Electrochemical analyses using competitive ligand equilibration/adsorptive cathodic stripping voltammetry (CLE-ACSV) showed that isolated compounds had conditional Fe-binding affinities (with respect to inorganic iron—Fe′) of K FeL,Fe′ cond =10 11.5 –10 11.9 M −1 , similar to purified marine siderophores produced in laboratory cultures and to the ambient Fe-binding ligands observed in seawater. In addition, 63% of the extracted compounds from surface-collected samples fall within the defined size range of siderophores (300–1000 Da). Hydroxamate or catecholate Fe-binding functional groups were present in each compound for which Fe binding was detected. These results illustrate that the functional groups previously shown to be present in marine and terrestrial siderophores extracted and purified from laboratory cultures are also present in the natural marine environment. These data provide evidence that a significant fraction of the organic Fe-binding compounds we collected contain Fe-binding functional groups consistent with biologically produced siderophores. These results provide further insight into characteristics of the Fe-binding ligands that are thought to be important in controlling the biological availability of Fe in the oceans.

Domoic acid binds iron and copper: a possible role for the toxin produced by the marine diatom Pseudo-nitzschia

Toxigenic species of the pennate diatom Pseudo-nitzschia can produce domoic acid (DA), an analog of the excitatory neurotransmitter glutamate and known to cause the human illness amnesic shellfish poisoning (ASP). Although the trophic transfer of this phycotoxin resulting in mass marine bird and mammal mortality has recently been demonstrated, the physiological role of domoic acid to the causative organism is still unknown. Domoic acid is a small tricarboxylate amino acid whose structure resembles that of known iron-complexing agents produced by terrestrial plants, such as mugineic acid. This similarity in chemical structure of domoic acid to other phytosiderophores suggests a role for domoic acid as a trace metal chelator. Using a highly sensitive adsorptive cathodic stripping voltammetric technique, we investigated the iron and copper-binding characteristics of domoic acid revealing it does form chelates with both iron and copper and with the following conditional stability constants: KFeDA,Fe(III)′cond=108.7±0.5 M−1 and KCuDA, Cu(II)′cond=109.0±0.2 M−1 (KFeDA,Fe3+cond=1018.7±0.5 M−1 and KCuDA, Cu2+cond=1010.3±0.2 M−1). Certain Pseudo-nitzschia species may therefore produce domoic acid to selectively bind trace metals in order to either increase the availability of an essential micronutrient, such as in the case of iron, or to decrease the availability of a potentially toxic trace metal, such as in the case of copper. The strength with which domoic acid binds iron and copper in seawater combined with concentrations of dissolved domoic acid potentially produced and released during toxic bloom conditions render dissolved iron and copper sufficiently bound to domoic acid in seawater so as to affect their chemical speciation and thus their biological availability. In addition, domoic acid may be particularly important in solubilizing particulate iron suspended in these coastal waters where Pseudo-nitzschia blooms tend to occur. Thus, possible physiological roles for domoic acid with respect to the harmful algal species Pseudo-nitzschia may include the acquisition (iron) or detoxification (copper) of trace metals in seawater.

Short-term biogeochemical influence of a diatom bloom on the nutrient and trace metal concentrations in South San Francisco Bay microcosm experiments

Two laboratory microcosm experiments were conducted to mimic an annual spring diatom bloom in South San Francisco Bay by isolating the phytoplankton community from the benthic grazing pressure to induce a phytoplankton bloom. The purpose of these experiments was to isolate the impact of a spring diatom bloom on the nutrient and trace metal geochemical cycling. Microcosms were created in 2.5 L incubation bottles and subjected to one of 4 treatments (control, copper [Cu] addition, manganese [Mn] addition, and both Cu and Mn addition) to investigate the toxicity of Cu on the resident plankton and the potential antagonistic effects of Mn on reducing Cu toxicity. Dissolved macronutrient (nitrate + nitrite, phosphate, and silicate), and dissolved and particulate trace metal (Cu, Ni, Mn) concentrations were monitored in the grow-out incubations on a daily basis. Chlorophylla concentrations were also monitored over the course of the experiment and used to calculate diatom-specific growth rates. In the experiments containing ambient South San Francisco Bay surface waters, average specific growth rates were on the order of 1.1 d−1. The induced diatom blooms resulted in significant removal of macronutrients from the microcosms over the course of the experiments. Our research supports previous suggestions that dissolved Ni and Cu concentrations in South San Francisco Bay have a very low biological availability as a result of organic chelation. Ni(EDTA)2− has been found to be the dominant dissolved Ni species by other researchers and Cu speciation analyses from this study and others indicate that > 99% of the dissolved Cu in South San Francisco Bay is strongly chelated as CuL1. The free cupric ion concentration was on the order of 10−12 M. Marked removal of dissolved Mn was observed in the control treatments, well exceeding expected dissolved Mn removal by diatom uptake. Additions of 375 nM Cu resulted in the complete titration of the chelating ligand (L1) concentrations. The elevated [Cu2+] (≈10−8MM) appeared to have a toxic effect on the diatom community observed in the significant decreases in their specific growth rates (μ=0.4 d−1). The suppression of dissolved Mn removal from solution was also observed in treatments spiked with high levels of dissolved Cu, providing support that Mn precipitation was due to biologically mediated oxidation not phytoplankton assimilation. The observed geochemical behavior in the concurrent Cu and Mn addition treatments provide evidence in support of Mn alleviation of Cu toxicity. The biological role in the ambient short-term biogeochemical cycling of Cu and Ni in South San Francisco Bay appears to be minimal due to the inert character of the organic ligand-metal complexes. A significant portion of the annual macronutrient and Mn cycling occurs as a result of spring diatom blooms in South San Francisco Bay.