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RESEARCH ARTICLE
Contents Vol 60(12)

Optimising extraction of extracellular polymeric substances (EPS) from benthic diatoms: comparison of the efficiency of six EPS extraction methods

Eri Takahashi A , Jérôme Ledauphin B , Didier Goux C and Francis Orvain A D
+ Author Affiliations
- Author Affiliations

A UMR 100 IFREMER-Université de Caen Basse-Normandie ‘Physiologie et Ecophysiologie des Mollusques Marins (PE2M)’, Laboratoire de Biologie et Biotechnologies Marines, Esplanade de la Paix, B.P. 5186, 14032 Caen Cedex, France.

B Laboratoire ERPCB-EA 3914, Boulevard Maréchal juin, Campus 2, 14032 Caen Cedex, France.

C Centre de microscopie appliquée à la biologie, Université de Caen Basse-Normandie, IFR 146, Campus 1, Esplanade de la Paix, B.P. 5186, 14032 Caen Cedex, France.

D Corresponding author. Email: francis.orvain@unicaen.fr

Marine and Freshwater Research 60(12) 1201-1210 https://doi.org/10.1071/MF08258
Submitted: 9 September 2008  Accepted: 18 March 2009   Published: 17 December 2009

Abstract

There is no universal method that can be applied to extract bound extracellular polymeric substances (EPS) from benthic diatoms of intertidal sediments without causing cell lysis. Six extraction methods were tested on a diatom culture of Navicula jeffreyi to establish the best compromise between high yields of carbohydrate extraction and minimum cell lysis. Extraction with distilled water provoked cell lysis (as already known). The five other extraction methods (dowex resin, artificial seawater of half salinity and extractions after pretreatment with gluteraldehyde by three methods: water, dowex water and dowex buffer) did not provoke cell lysis as shown by transmission electronic microscopy. This result was confirmed by the minimum release of internal compounds (protein, ATP) and by the low proportions of glucose in dowex-extracted EPS compared with the water-extracted EPS, from which the high glucose content must be inferred as contamination by the chrysolaminaran. The extraction with dowex resin resulted in the second-highest concentration of carbohydrate after the water extraction and the EPS were especially rich in deoxy sugars, hence increasing the hydrophobic feature of these substances. For these reasons, we recommend extraction with dowex, which is also the best method for extracting bound EPS from other biofilms such as in activated sludges.

Additional keywords: biofilm, cell lysis, dowex, GCMS analysis, immunostaining, immuno-staining, intertidal sediments, Navicula jeffreyi, protein, transmission electron microscopy.


Acknowledgements

The authors thank the two anonymous referees for their helpful comments and Dr Anthony Chiovitti for his initial guidance in the study about immuno-staining. We are very grateful to Professor M. R. Gretz for his advice about monosaccharide analysis using the GC/MS. We thank Dr Benoit Véron and Mr Bertrand Le Roy of Algobank for supplying us with the diatoms. We also thank Dr Pascal Claquin and Dr Sébastien Lefebvre for their advice during cell culturing, and Florent Pierre, a student who helped during the preparation of the study. Funding was provided by the National Research Agency (ANR) and this study is a part of the VASIREMI project (Trophic significance of benthic microorganisms in intertidal mudflats) led by Dr Christine Dupuy.


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