Protein extraction from activated sludge: an analytical approach.
To investigate the efficiency of different methods on exopolymeric substance (EPS) extraction, mechanical and chemical treatments were applied on two activated sludges, regarding the yield of protein extraction as well as their compatibility with usual quantification methods. Mechanical disruption methods do not drastically affect protein measurements by both bicinchoninic acid (BCA) and modified Lowry methods. Chemical compounds such as cationic exchange resin and triton show high interference with modified Lowry method while the protein quantification by BCA method is not affected. In addition, inner sludge compounds were shown to interfere with both methods: BCA and modified Lowry measurement respectively overestimate and underestimate protein content. According to these data, BCA method was chosen in this study as the most appropriate protein quantification method in sludge extracts. Comparison of various extraction protocols, combining mechanical and/or chemical treatments, shows that efficiency can be increased by repeating the same method or by applying a prior mechanical treatment. Proteins are preferably extracted by triton treatments, indicating the importance of hydrophobic interactions linking proteins to the EPS matrix. The amount of extracted proteins reaches 182 and 148 mg eq.BSA g(-1)VSS using triton/triton and ultraturax/triton extractions, respectively. Protease activity/extracted protein ratios vary widely depending on extraction protocols. Protease seemed to be preferably extracted by ultrasound and triton treatments (150-220 U mg(-1)protein). This study underlines that the choice of a relevant coupled quantification/extraction method is of great importance for efficient EPS determination.
Ras
M
M
LBAE, Laboratoire de Biologie Appliquée à l'Agro-Alimentaire et à l'Environnement, Institut Universitaire de Technologie, Université Toulouse III, Auch, France.
Girbal-Neuhauser
E
E
Paul
E
E
Spérandio
M
M
Lefebvre
D
D
D002411:Cation Exchange Resins; D005954:Glucosephosphate Dehydrogenase / Q000302:isolation & purification; D008049:Lipase / Q000302:isolation & purification; D010447:Peptide Hydrolases / Q000302:isolation & purification; D011506:Proteins / Q000302:isolation & purification*; D012722:Sewage; D013501:Surface-Active Agents / Q000737:chemistry
D002411:Cation Exchange Resins; D011506:Proteins; D012722:Sewage; D013501:Surface-Active Agents; D005954:Glucosephosphate Dehydrogenase; D008049:Lipase; D010447:Peptide Hydrolases