Studies on an endotoxin produced by Pseudomonas aeruginosa

O'Neill, George J (1963) Studies on an endotoxin produced by Pseudomonas aeruginosa. PhD thesis, University of Glasgow.

Abstract

The Introduction to the thesis is devoted to a review of the principal methods which have been used in the extraction of the extraction of the endotoxins or somatic antigens of Gram negative bacteria, and the chemical nature of those substance. The part played by the lipid portion of the complex in the biological activity is discussed along with the effect on the biological activity of chemical modification of endotoxins. The experimental work is in two main sections. In the first of these, the isolation, purification and properties of the ondotoxin from Pseudomonas aeruginosa are described and the results discussed. The ondotoxin was isolated as a lipopolysaccharide by a procedure which involved heating acetone-dried bacterial cells with aqueous phenol, followed by dialysis and high speed centrifugation of the aqueous extract. The lipopolysaccharide was shown to contain galactose, glucose, galactosamine, glucosamine and an aldohoptooe. The lipid portion obtained after short acid hydrolysis contained other-soluble and chloroform-soluble fractions in both of which glucosamine, several amino acids and a series of fatty acids were detected. The lipopolysaccharide was shown to be antigenic and highly pyrogenic when injected into rabbits, and to be toxic to rabbits and mice. The second experimental section is concerned with the effect of a combination of trichloracetic acid and phenol extraction on the Pseoudomonas aerpginosa cells. Cold trichloracetic acid extraction by the Boivin technique gave a low yield of endotoxin, but it was found that the rosidue after trichloracetic acid extraction could be treated with hot aqueous phenol to give appreciable amounts of an endotoxic material which was similar to the ondotoxin obtained by direct treatment of the cells with hot phenol, but which was free from contaminating nucleic acid. By the use of the combined extraction technique, nucleic acid-free lipopolysaucharides were also obtained in good yield from proteus vulgaris and Escherichia coli. It was also found that the maintenance of low temperature during the trichloracotic acid extraction was not essential when using the double extraction method to obtain nucleic acid-free lipopolysaccharides. A farther series of experiments showed that even after multiple extraction with cold trichioracetic acid, a considerable amount of endotoxin remained unextracted and could be obtained from the residue by treatment with hot phenol.

• Item Type: Thesis (PhD)
• Qualification Level: Doctoral
• Additional Information: Adviser: J P Todd
• Keywords: Biochemistry, Microbiology
• Date of Award: 1963
• Depositing User: Enlighten Team
• Unique ID: glathesis:1963-73105
• Copyright: Copyright of this thesis is held by the author.
• Date Deposited: 14 Jun 2019 08:56
• Last Modified: 14 Jun 2019 08:56
• URI: https://theses.gla.ac.uk/id/eprint/73105