Method for detecting the 3-hydroxymyristic acid component of the endotoxins of gram-negative bacteria in compost...

An analytical chemical method for the 3- or beta-hydroxymyristic (BHM) acid component of endotoxins has been developed for the lipopolysaccharide (LPS) of E. coli, intact E. coli, and sewage sludge compost. Endotoxins are the pyrogens associated with the outer membranes of many gram-negative bacteria, for example, E. coli and Salmonella. The BHM acid content was used as a chemical marker for endotoxin presence, since BHM acid is present in the molecular subunit (Lipid-A) responsible for the toxicity and is the most abundant saturated fatty acid in that subunit. BHM acid quantification thus complements the Limulus bioassay to detect gram-negative bacteria presence in such samples as cotton and other dusts, blood, water, compost and air samples. BHM acid was isolated after digestion, ether extraction, alkaline hydrolysis, and ether extraction. The free acid was quantitated as the methyl ester using two GC columns as a screening method or by GC/MS for confirmatory purposes. Recoveries were unreliable below 15 ng of BHM acid. By the use of the 2-column screening technique, the amounts of BHM equivalent in E. coli LPS, intact E. coli and compost in micrograms BHM acid/g substrate were (arithmetic mean +/- standard deviation), respectively: 56 120 +/- 2200; 2650 +/- 339; and 14.7 +/- 5.7.