Chemical characterization of Pseudomonas veronii 2E soluble exopolymer as Cd(II) ligand for the biotreatment of electroplating wastes

MARIA L. FERREIRA; ADRIANA CASABUONO; ALICIA S COUTO; SILVANA A.M. RAMIREZ; DIANA L. VULLO

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Simposio; 15th. International Symposium on Microbial Ecology, ISME 15; 2014

International Society for Microbial Ecology (ISME)

Wastewater treatments mediated by bacteria or bacterial components are an innovative technology available for metal containing industrial discharges. Some bacteria are adapted to metal polluted environments because of the development of different survival strategies. Pseudomonas veronii 2E is an indigenous bacterium from the highly polluted Reconquista River (Buenos Aires Metropolitan Area) that produces exopolymers (ES) with potential metal-binding ability. The target of this study was to characterize the P.veronii 2E exopolysaccharide (EPS), a major component of the secreted exopolymer, and study Cd(II) complexation capacity for future applications in waste treatments. ES was produced in a minimal medium M9-2% (v/v) glycerol, precipitated with ethanol and purified by both dialysis (MW>12,400Da) and enzymatic treatments (DNase I and Proteinase K). The biochemical composition was assessed using colorimetric methods: protein content (Bradford); neutral sugars (anthrone assay); phosphate (molybdenum blue assay) and uronic acids (sulfamate/m-hydroxydiphenyl assay). Products of total acid hydrolysis of ES (with TFA, trifluoroacetic acid) were analyzed by High Performance Anion Exchange Chromatography with Pulsed Amperometric Detection (HPAEC-PAD), showing the exclusive presence of fucose, galactosamine, glucosamine, galactose, glucose, mannose and glucuronic acid. A pyruvilated sugar was revealed after treatment with oxalic acid following TFA hydrolysis. Deoxy-cholate-polyacrylamide gel electrophoresis (PAGE) and silver/alcian blue stain of the purified ES showed both extracellular polymeric substances EPS and lipopolysaccharide LPS. Monosaccharide analysis of the isolated EPS showed no difference with the whole ES fraction, suggesting that the oligosaccharide linked to the Lipid A moiety of the LPS would have a similar structure. The ability of ES to complex Cd(II) was determined by an electrochemically monitored titration (ASV) at pH 5.5, 6.2, 7.5 or 8.1 (25±1°C). One family of moderate strength binding sites was observed at all pHs evaluated with a range of Log K´ (conditional formation constant) 5.69±0.08, while ligand concentration (moles of binding sites/g dry weight ES) was in the 74±7 mM range. Future electrochemically monitored titrations of EPS and LPS fractions combined with structure characterization by Mass Spectrometry will help understand the ES complexing capacity responsible for the interaction with metals. ES represents a useful tool for metal retention from electroplating wastes.