Classification of Bacillus sp. by Determination of Volatile Compounds Produced in the Culture

Abstract

The strains of the genus Bacillus are well known as a causative organism of food spoilage. In order to classify Bacillus sp., gas chromatography was applied for determination of volatile compounds produced in the culture medium and statistical analysis employing cluster analysis were made. The conditions were determined as optimum for producing the volatile compounds as follows; the medium contained 0.5% yeast extract, 0.5% glucose, 1.0% tryptone, 3.66% K₂HPO₄ and 0.54% KH₂PO₄, and the cultures were grown in a 500ml volume shaker flask containing 200ml of the medium at 30°C for 24 hours with reciprocal shaking. The concentrate of volatile compounds from the culture was prepared by passing a centrifuged supernatant (200ml) through a porapak R column and eluting adsorbed compounds with ethyl ether (1.5ml). About 40-80 compounds from 3 to 300 ppb were determined by GC analysis. By employing cluster analysis, 27 samples of 9 strains in the genus Bacillus were clearly classified corresponding to their strains. Further, 6 varieties of B. subtilis were also able to be classified correctly.