Abstract

GUALTERO ESCOBAR, Diego Fernando et al. Purification and characterization of lipopolysaccharide from Eikenella corrodens 23834 and Porphyromonas gingivalis W83. Rev. colomb. biotecnol [online]. 2014, vol.16, n.1, pp.34-44. ISSN 0123-3475.  https://doi.org/10.15446/rev.colomb.biote.v16n1.44224.

Purification of lipopolysaccharide (LPS) or endotoxins and its characterization is an important aspect for studies aimed at clarify the role of these biomolecules from Gram negative bacteria present in the oral cavity and its relationship with periodontal and systemic diseases. This study describes an extraction, purification and characterization method of LPS from Eikenella corrodens 23834 and Porphyromonas gingivalis W83. LPS extraction was performed by using hot phenol-water; the purification was done with nuclease and protease enzymatic treatment, followed by size-exclusion chromatography (Sephacryl S-200 HR) with sodium deoxycholate as mobile phase. The characterization of the purified extracts was performed by spectrophotometric scanning, SDS-PAGE biochemical tests, Purpald assay and chromogenic LAL test. As control, commercial LPS from Escherichia coli, Salmonella typhimurium, P. gingivalis, and Rodobacter sphaeroides were used. The methodology mentioned above had allowed obtaining high purity LPS by identifying KDO or heptoses, a chemotype S-LPS (smooth) to E. corrodens; SR-LPS (semi-rough) for P. gingivalis W83. Both purified LPS showed endotoxic capacity at low concentrations. The methodology used in this study for purification and characterization of LPS from the whole bacteria was efficient when it was compared with commercial LPS.

Keywords : endotoxin; 2-keto-3-deoxyoctonate (KDO); Chromatography; Limulus test (LAL); periodontitis.

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