Bacterial Exudates Influences on Trace Metal Chemistry in Aquatic Systems. The Cd2+, Cu2+, Ferrihydrite and Bacterial Exudate Systems

Abstract

Understanding trace metal speciation in aquatic systems is an important prerequisite to understanding water resources. A diverse range of organic ligands are important in trace metal speciation in many aquatic systems. In this work the chemistry of the trace metals Cd2+ and Cu2+ is explored with the organic ligands present in a bacterial exudate obtained from suspensions of Bacillus subtilis. The system is extremely complex and heterogeneous and several approaches were used for the investigation. Acid-base potentiometric titrations of the exudate could be well described using a simplified approach in the modelling with a single diprotic ligand (H2Ex) with a concentration of 1.2 ± 0.2 mM and pKA’s of 3.9 ± 0.2 and 8.3 ± 0.1. The exudate titration curves were shifted by the presence of Cd2+ or Cu2+ and this could be described by including a metal-ligand complex. In the case of Cu2+ the titration curve could be reproduced using a single Cu-ligand complex with stoichiometry CuOH-Ex-. In the case of Cd two Cd-ligand complexes with stochiometries Cd-Ex0 and CdOH-Ex- were needed. Polarograms of Cu2+ in exudate solutions had two peaks. One peak at ≈ -0.2 V is due to a copper-exudate complex and a second peak at ≈ 0.0 V due to free Cu2+. As Cu2+ was added in increments to the 10 times diluted exudate at pH 5.2 the peak at -0.2 V grew and the free Cu2+ peak only appeared with > 6 μM added Cu2+. This extent of Cu2+ complexation is much greater than that predicted by the parameters derived from the acid-base titration curve modeling and suggests heterogeneity of Cu2+ binding sites in the exudate with a low concentration of high affinity sites. Fluorescence excitation-emission matrix spectra of the exudate indicated the presence of 4 classes of fluorescent compounds which, based on their spectral positions, are broadly attributed to proteins with aromatic amino acids, microbial byproduct like compounds, humic acid like and fulvic acid like compounds. The reaction of the exudate with the iron oxide ferrihydrite was probed by in situ attenuated total reflection infrared (ATRIR) spectroscopy and using fluorescence spectroscopy and polarography. The ATRIR revealed linkages to the ferrihydrite via phospho-groups with several strong peaks at ≈ 1100 cm-1. In addition amide peaks in the ATRIR indicated protein association with the surface which was apparently independent of the phospho-groups. The microbial by-product like feature in the fluorescence spectra was the only feature definitively reduced after exudate reaction with ferrihydrite and the extent of reduction (≈ 50%) was almost independent of pH. Similarly the Cu2+ binding capacity as assessed by polarography was also reduced by approximately 50 % after exudate exposure to ferrihydrite and this too was almost independent of pH. Finally the effect of exudate on Cu2+ sorption by ferrihydrite was measured as a function of pH in a system with 1 mM ferrihydrite and 23 μM Cu2+. The presence of exudate enhanced Cu2+ sorption by up to 20 % at pH < 6 but at pH > 6 the opposite was true with the exudate causing up to ≈ 20 % less Cu2+ to be sorbed. This indicates a competing influence of the formation of solution Cu-exudate complexes and ternary Cu-exudate ferrihydrite complexes and is quite similar to the effect of phthalate on Cu2+ sorption by ferrihydrite under some conditions. Overall the exudate demonstrates a very rich and complex range of chemistries with a high level of heterogeneity. The current study deployed a range of methods and each provided insights into the reactions occurring but the study is very much a “broad brush” approach to developing initial insights into this system. Substantially more work would be needed to make definitive conclusions about the system in particular that link the different aspects probed in the current study.