Characterization of extracellular polymeric substances from acidophilic microbial biofilms

Abstract

We examined the chemical composition of extracellular polymeric substances (EPS) extracted from two natural microbial pellicle biofilms growing on acid mine drainage (AMD) solutions. The EPS obtained from a mid-developmental-stage biofilm (DS1) and a mature biofilm (DS2) were qualitatively and quantitatively compared. More than twice as much EPS was derived from DS2 as from DS1 (approximately 340 and 150 mg of EPS per g [dry weight] for DS2 and DS1, respectively). Composition analyses indicated the presence of carbohydrates, metals, proteins, and minor quantities of DNA and lipids, although the relative concentrations of these components were different for the two EPS samples. EPS from DS2 contained higher concentrations of metals and carbohydrates than EPS from DS1. Fe was the most abundant metal in both samples, accounting for about 73% of the total metal content, followed by Al, Mg, and Zn. The relative concentration profile for these metals resembled that for the AMD solution in which the biofilms grew, except for Si, Mn, and Co. Glycosyl composition analysis indicated that both EPS samples were composed primarily of galactose, glucose, heptose, rhamnose, and mannose, while the relative amounts of individual sugars were substantially different in DS1 and DS2. Additionally, carbohydrate linkage analysis revealed multiply linked heptose, galactose, glucose, mannose, and rhamnose, with some of the glucose in a 4-linked form. These results indicate that the biochemical composition of the EPS from these acidic biofilms is dependent on maturity and is controlled by the microbial communities, as well as the local geochemical environment.

FIG. 1.

FTIR spectra of EPS from DS1 and DS2 biofilms. Peak 1, —CH— vibrations in lipids; peak 2, amide I in proteins; peak 3, amide II in proteins; peak 4, —COC— group vibrations in carbohydrates, DNA, and RNA.

FIG. 2.

Comparison of the biochemical compositions of EPS from DS1 and DS2 biofilms. The error bars indicate standard deviations (n = 3).

FIG. 3.

Solid-state ¹³C NMR spectra of EPS from DS1 and DS2 biofilms, revealing the distribution of bonding environments. The spectral regions with relevance to the present study include the regions for peptide-bonded carbon (peptide), polysaccharide glycosidic carbon and secondary alcohols (area in the rectangle), and saturated hydrocarbon (lipids). Differences in the spectral features in the carbohydrate region indicate substantial differences in the compositions of the polysaccharides in the two biofilms.