Lyophilized, 48-hr log-phase vegetative cells were extracted with chloroformmethanol (2:1, v/v) and ethanol-ether (3:1, v/v) and then saponified with methanolic KOH. Gas-liquid chromatography of the methyl esters of extractable fatty acids revealed distinctive "pattern profiles" of Clostridium botulinum toxigenic types "A," "B," and "E." C. perfringens type "A" and Escherichia coli strain "B" were also studied in a similar manner and were found to give pattern profiles which were distinct even from those obtained for the C. botulinum microorganisms. Amino sugar content of the five microorganisms was determined by using a Beckman amino acid analyzer. The molar ratio of glucosamine to that of galactosamine was found to be of further assistance in distinguishing the individual microorganisms.

Lyophilized, 48-hr log-phase vegetative cells were extracted with chloroform-methanol (2:1, v/v) and ethanol-ether (3:1, v/v) and then saponified with methanolic KOH. Gas-liquid chromatography of the methyl esters of extractable fatty acids revealed distinctive “pattern profiles” of Clostridium botulinum toxigenic types “A,” “B,” and “E.” C. perfringens type “A” and Escherichia coli strain “B” were also studied in a similar manner and were found to give pattern profiles which were distinct even from those obtained for the C. botulinum microorganisms. Amino sugar content of the five microorganisms was determined by using a Beckman amino acid analyzer. The molar ratio of glucosamine to that of galactosamine was found to be of further assistance in distinguishing the individual microorganisms.

One hundred and two strains of aerobic, gram-negative bacteria, isolated mainly from drugs and cosmetics, were tested for catabolic products after incubation in liquid media containing single amino acids. Breakdown products from L-arginine, lysine, and ornithine were identified as their N-heptafluoryl butyl esters by gas-liquid chromatographic techniques. A Finnigan model 1015 D gas chromatograph/mass spectrometer/PDP-8E data system was used to identify trace amounts of catabolites after amino acid substrates were inoculated with bacteria. The derivatization reaction was monitored in like manner. The Finnigan data system may be used to identify catabolic compounds rapidly and thus provide a means for bacterial characterization and identification.The arginine dihydrolase test is particularly valuable in differentiating between various species of pseudomonads and other carbohydrate-oxidizing bacteria (11,13,15,18). Combination of this test with procedures reported by Moss et al. (15) and Lambert and Moss (12) for determining catabolic products of amino acids by gas-liquid chromatography (GLC) has resulted in a practical means of identifying bacteria. GLC has been utilized to differentiate the members of the family Enterobacteriaceae from the fermentative and oxidative bacteria. However, the decarboxylase reactions of the pseudomonads have not been studied as extensively as have those of the members of the family Enterobacteriaceae. We have evaluated the use of the GLC rapid test for differentiation of 102 strains of gram-negative bacteria. Both the saccharolytic and weakly saccharolytic, nonfermentative strains of Pseudomonas species as well as nonsaccharolytic nonfermentative bacteria, e.g. , Acinetobacter (16), were examined.The objectives of this study were to determine five catabolic metabolites produced by 102 various strains of gram-negative bacteria representing 12 species of Pseudomonas and 20 species of Enterobacteriaceae and to investigate the use of N-heptafluorobutyric anhydride as a GLC derivatizing reagent for the catabolites. This study was performed with the aid of a. gas chromatograph-mass spectrometer-data system. MATERIALS AND METHODSBacterial strains. The strains used in this study were obtained from either the culture collection of the Drug Microbiology Branch, U.S. Food and Drug Administration, Washington, D. C., or the American Type Culture Collection, Rockville, Md. The strains used are listed in Table 1.Methods. Identification of the bacteria was confirmed by established culture and biochemical procedures (4-6,8,9). The cultures were grown on brain heart infusion agar (Baltimore Biological Laboratories) slants for 16 to 24 h and then transferred to triple sugar iron agar (Difco) slants for an additional 36 t o 48 h at 35°C. Growth from the top half of the triple sugar iron agar slants was used t o inoculate all media. Cultures were tested for L-arginine dihydrolase and L-ornithine and L-lysine decarboxylase activities in two media: Moeller medium as described by Edwards and Ewing (5) and ...