Lipopolysaccharides (LPS) from the type strains of the anaerobic beer spoilage bacteria Pectinatus cerevisiiphilus and P. frisingensis were extracted with the 5:5:8 volume ratio modification of the phenolchloroform-petroleum ether method (H. Brade and C. Galanos, Eur. J. Biochem. 122:233-237, 1982). Sequential precipitations of LPS with water and acetone from the phenol phase yielded LPS which differed in that water-precipitable material (LPS-H2O; 0.1 to 0.4% of the dry weight of the cells) was rough-type LPS, whereas acetone-precipitable material (LPS-Ac; 4.6 to 5.8% of the dry weight) contained both rough-type LPS and high-molecular-weight material resembling smooth LPS. The LPS were chemically characterized, and they contained D-glucosamine, 4-amino-4-deoxy-L-arabinose, 3-deoxy-D-manno-2-octulosonic acid, D-fucose, D-galactose, D-glucose, D-mannose, and phosphate. D-Fucose was present mostly in LPS-Ac, suggesting that it is a constituent of the O antigen. The major fatty acids were ester- and amide-linked (R)-3-hydroxytridecanoic and ester-linked undecanoic acids, with minor amounts of ester-linked tridecanoic and (R)-3-hydroxyundecanoic acids. The chemical compositions of LPS-H2O and LPS-Ac suggested that they differ not only in their smooth or rough nature but also in the structure of their core regions. This may explain their different precipitabilities from the extraction mixture. The extraction method was also shown to be applicable to the isolation of smooth-type LPS from Salmonella enterica serovar Typhimurium. Extraction of two Typhimurium strains carrying chemically different O antigens resulted in high yields (8% of the dry weight) of LPS. Strain SH2183, which contains the relatively hydrophobic O-4,5,12 antigen yielded almost exclusively LPS-Ac, whereas the LPS of strain SH5770, which has a hydrophilic O-6,7 antigen, was exclusively LPS-H2O. No fractionation to smooth and rough LPS occurred with the Typhimurium strains.