Abstract

GUALTERO, DIEGO; CASTELLANOS, JAIME E; PEREZ, GERARDO  and  LAFAURIE, GLORIA I. Purification of Porphyromonas gingivalis polysaccharide free lipopolysaccharide using Sephacryl S-200 high resolution chromatography. Acta biol.Colomb. [online]. 2008, vol.13, n.3, pp.147-160. ISSN 0120-548X.

The aim of this work was to improve a standard methodology to purify Porphyromonas gingivalis lipopolysaccharide (LPS) using a protocol of extraction, enzymatic digestion and high resolution chromatography. P. gingivalis bacteria was cultured in anaerobiosis, their membranes were extracted using the phenol-water method, then subjected to DNAse, RNAse and protease digestion and finally, the extract was separated by chromatography using Sephacryl S-200. The purified extract was characterized by silver staining after polyacrylamide gel electrophoresis and 2-keto-3-deoxioctanoic acid (KDO) was detected using the Purpald’s method. A preparation free of nucleic acid-, protein-or polysaccharides was obtained. The chromatographic separation showed high resolution since there was two discrete peaks with different components. The purification protocol allowed us to obtain a highly purified P. gingivalis LPS which could be used in future tests to evaluate its behavior in vitro and in vivo and elucidate its function, as well as to obtain LPS from other periodontopatic bacteria to address the association of periodontal disease with cardiovascular diseases.

Keywords : Periodontitis; lipopolysaccharide; endotoxin; chromatography.

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