For Pe extraction a protocol used by Marcati et al., 2014 was followed, involving a two-step membrane PD123319 process. First an ultrafiltration by 1.5 * 106 Da membrane, then the resulting permeate filtered by 10 * 103 Daltons membrane to recover from the retentate the Pe. Finally the Pe was dried (24 h) using a freeze drier.
2.6. Fourier Transformed Infra-red (FTIR) analysis
FTIR attenuated total reflectance (ATR) spectra was collected on a PerkinElmer Spectrum Two instrument equipped with a diamond crystal ATR reflectance cell with a DTGS detector scanning over the wavenumber range of 4000–450 cm−1 at a resolution of 4 cm−1. Approximately 3–5 mg of finely powdered freeze-dried biomass, Pe or EPS extract was prepared in the same manner as for conventional biochemical analysis (described in Sections 2.2 and 2.4) and was applied to the surface of the crystal and then pressed onto the crystal head. Three replicates (each consisting of an melanin average of 10 scans) for each sample were taken and the results averaged. Background correction scans of ambient air were made prior to each sample scan. Scans were recorded using the spectroscopic software Spectrum (version 10. PerkinElmer, Germany). Spectra were background corrected for ambient air. Ethanol (70%) was used to clean the diamond ATR between samples.