Application of Raman Spectroscopy to the Biooxidation Analysis of Sulfide Minerals

We report the application of confocal laser scanning microscopy CLSM and Raman spectroscopy on the (bio)chemical oxidation of pyrite and chalcopyrite, in order to understand how surface sulfur species (S𝑛2−/S0) affects biofilm evolution during mineral colonization by Acidithiobacillus thiooxidans. We found that cells attachment occurs as cells clusters and monolayered biofilms within the first 12 h. Longer times resulted in the formation of micro- and macrocolonies with variable cell density and higher epifluorescence signal of the extracellular polymeric substances (EPS), indicating double dynamic activity of A. thiooxidans: sulfur biooxidation and biofilm formation. Raman spectra indicated S𝑛2−/S0 consumption modification during biofilm evolution. Hence, cell density increase was primarily associated with the presence of S0; the presence of refractory sulfur species on the mineral surfaces does not to affect biofilm evolution. The EPS of the biofilms was mainly composed of extracellular hydrophobic compounds (vr. gr. lipids) and a minor content of hydrophilic exopolysaccharides, suggesting a hydrophobic interaction between attached cells and the altered pyrite and chalcopyrite.