Metal- Pseudomonas veronii 2E Interactions as Strategies for Innovative Process Developments in Environmental Biotechnology

Abstract

The increase of industrial discharges is the first cause of the contamination of water bodies. The bacterial survival strategies contribute to the equilibrium restoration of ecosystems being useful tools for the development of innovative environmental biotechnologies. The aim of this work was to study the Cu(II) and Cd(II) biosensing, removal and recovery, mediated by whole cells, exopolymeric substances (EPS) and biosurfactants of the indigenous and non-pathogenic Pseudomonas veronii 2E to be applied in the development of wastewater biotreatments. An electrochemical biosensor was developed using P. veronii 2E biosorption mechanism mediated by the cell surface associated to bound exopolymeric substances. A Carbon Paste Electrode modified with P. veronii 2E (CPEM) was built using mineral oil, pre-washed graphite power and 24 h-dried cells. For Cd(II) quantification the CPEM was immersed in Cd(II) (1-25 μM), detected by Square Wave Voltammetry. A similar procedure was used for 1-50 μM Cu(II). Regarding Cd(II), removal mediated by immobilized EPS was tested in a 50 ml bioreactor with 0.13 mM Cd(II), pH 7.5. A 54% metal retention by EPS was achieved after 7 h of continuous operation, while a 40% was removed by a control resin. In addition, surfactants produced by P. veronii 2E were studied for recovery of Cd(II) adsorbed on diatomite, obtaining a 36% desorption efficiency at pH 6.5. Cu(II) adsorption from a 1 mM solution was tested using P. veronii 2E purified soluble EPS in 50 mL- batch reactors (pH = 5.5, 32°C). An 80% of the initial Cu(II) was retained using 1.04 g immobilized EPS. Focusing on metal recovery, Cu nanoparticles (NPs) biosynthesis by P. veronii 2E was carried out in Cu(II)-PYG Broth at 25°C for 5 days. Extracellular CuNPs were characterized by UV-Vis spectral analysis while both extracellular and intracellular NPs were analyzed by SEM and TEM techniques. Responses of P. veronii 2E and its products as biosurfactants, bound and soluble EPS allowed Cu(II) and Cd(II) removal, recovery and biosensing resulting in a multiple and versatile tool for sustainable wastewater biotreatments.

Keywords: Cu nanoparticles; Pseudomonas veronii; biosensor; biosurfactant; exopolymeric substances; metal removal and recovery.

FIGURE 1

Continuous (A) and batch (B) reactors designed for metal biosorption by EPS.

FIGURE 2

Chemical structure of secreted Pseudomonas veronii 2E biosurfactant.

FIGURE 3

Kinetics of copper biosorption (A) and desorption cycles (B) mediated by EPS in a batch reactor.

FIGURE 4

Percentage of remaining Cd(II) in time in a continuous flow bioreactor, using the EPS as biosorbent and Chelex^® 100 resin as reference.

FIGURE 5

Visual color changes and absorption spectra of P. veronii 2E culture supernatants after 5 days-growth in presence of CuSO₄ (A). SEM Microphotographs for extracellular Cu-NPs detection and characterization from culture supernatants (B). TEM Microphotographs of aggregates belonging to cell-associated Cu-NPs: images I and II correspond to Cu(II) free cultures, while images III, IV, V, and VI show cells grown in 1–1.5 mM CuSO₄ (C).

FIGURE 6

Effect of temperature on peak current after 10 μM Cd(II) exposition (A). Effect of preconcentration potential on Cd(II) peak current. Insert graphic: SWV in 0.1 M NaNO₃ for preconcentration potential –0.9 to –1.2 V (B). Effect of preconcentration time on peak current. Insert graphic shows SWV for preconcentration times: 60 s (blue), 40 s (red), and 20 s (green) (C).

FIGURE 7

Comparison of the responses to Cd(II) exposure of CPE and CPEM in 0.1 M NaNO₃. CPE, exposed to 102.5 μM Cd(II) (red) and baseline (green); CPEM, exposed to 50 μM Cd(II) (blue) and baseline (violet) (A). Comparison of the responses to Cu(II) exposure of CPE and CPEM and effect of supporting electrolyte. CPEM exposed to 15 μM Cu(II) in 0.003 M HNO₃ (green) and in 0.1 M NaNO₃ (red). Baselines for CPEM in 0.1 M NaNO₃ (black) and 0.003 M HNO₃ (blue). CPE exposed to 100 μM Cu(II) in 0.003 M HNO₃ (violet) and baseline (pink) (B).

FIGURE 8

Calibration curve for Cd(II) under optimal experimental conditions. Insert graphic shows linear range between 0.6 and 9 μM (y = 0.4279x – 0.1121; R² = 0.9902) (A). Calibration curve for Cu(II) and linear range between 1 and 25 μM (y = 0.14536x – 0.7377; R² = 0.99794) (B).