Preparation of endotoxin from pathogenic gram-negative bacteria

Endotoxins have been recognized for decades as important structural components of the outer cell wall/cell membrane complex of Gram-negative microorganisms. These chemically heterogeneous macromolecular structures were recognized very early on to consist of lipid, polysaccharide, and protein, and to have the capacity to induce deleterious pathophysiological changes when administered either systemically or locally to a wide variety of experimental laboratory animals. The recognition of the very significant disease-causing potential of these interesting microbial constituents provided a sound conceptual basis for studies directed at the isolation, purification, and detailed chemical characterization of the active constituent(s). It is perhaps not particularly surprising, therefore, that there are now numerous methods and modifications of methods, that have been published in the scientific literature describing various approaches that have been employed for the extraction and purification of endotoxin from bacteria. It would be beyond the scope of this chapter to describe in detail all of these various methods. Therefore, we shall provide only a brief historical perspective of the evolution of different methodologies. We will then focus upon a more detailed discussion of those that will ultimately serve the investigative purposes of most researchers interested in isolating and purifying endotoxins.