concentration of polysaccharides and proteins in EPS of Pseudomonas putida and Aureobasidum pullulans as revealed by ¹³C CPMAS NMR spectroscopy

• Published in: Applied microbiology and biotechnology 2009-11, Vol.85 (1), p.197-206
• Main Authors: Metzger, Ulrich, Lankes, Ulrich, Fischpera, Kai, Frimmel, Fritz H
• Format: Article
• Language: English
• Subjects: Ascomycota - metabolism; Aureobasidium pullulans; Bacterial Proteins - analysis; Biofilms; Biological and medical sciences; Biotechnology; Carbohydrates; Carbon; cell aggregates; Cell culture; EDTA (chelating agent); Extraction processes; Formaldehyde; Fundamental and applied biological sciences. Psychology; Fungal Proteins - analysis; Life Sciences; Magnetic Resonance Spectroscopy - methods; Methods; Microbial Genetics and Genomics; Microbiology; nitrogen content; NMR; Nuclear magnetic resonance; nuclear magnetic resonance spectroscopy; Polymers; Polymers - chemistry; Polymers - isolation & purification; Polymers - metabolism; polysaccharides; Polysaccharides - analysis; protein content; Proteins; Pseudomonas putida; Pseudomonas putida - metabolism; Saccharides; Sodium hydroxide; Spectroscopy; Spectrum analysis; Stainless steel; Studies
• ISSN: 0175-7598; 1432-0614
• DOI: 10.1007/s00253-009-2218-y

Extracellular polymeric substances were extracted from the bacterial strain Pseudomonas putida and the fungal species Aureobasidium pullulans using three different methods (formaldehyde-NaOH, ethylenediaminetetraacetic acid (EDTA) and cation-exchange-resin). The composition of the extracellular polymeric substances (EPS) was analysed by biochemical and high-resolution solid state ¹³C nuclear magnetic resonance (NMR) spectroscopic methods. The EPS yield was strongly dependent on the extraction method, with the formaldehyde-NaOH method showing the best extraction efficiency. The NMR method revealed that when using the EDTA extraction method, about 40% of the EDTA accumulated in the EPS and that was responsible for the apparent high extraction yields. EPS protein content determined by the NMR method was up to 30% higher than the protein content determined using the biochemical (Lowry) method for P. putida and for A. pullulans. The average protein carbon content determined by the NMR method was approximately 70% of the total carbon content. NMR results could be supported by elemental analysis, which showed a high nitrogen content (~10%) in the EPS. The carbohydrate carbon content detected with both methods in the cell aggregates and the EPS was approximately 20% in each. In this study, quantitative ¹³C cross-polarisation magic angle spinning NMR spectroscopy was conducted on unlabeled cell strains, and EPS and could be used to quantify protein and carbohydrate of different samples.