Lipopolysaccharide (LPS) of Brucella abortus strain 45/20 was purified using a novel method. Monophosphoryl lipid A (MPLA) was prepared from this LPS, methylated, and purified by high performance liquid chromatography. Chemical, mass spectral, and nuclear magnetic resonance analyses showed that MPLA consists of heptaacyl lipid As with molecular weights of 2095, 2123, 2151 and 2179. They contained the β-1,6-linked 2,3-diamino-2,3,-dideoxy-glucose disaccharide backbone and a phosphate group at the 4' position. Bisphosphoryl lipid A was also prepared and completely O-deacylated. It contained an additional phosphate group, and either 2 hydroxyhexadecanoic, 1 hydroxytetradecanoic, 1 hydroxydodecanoic acids or 2 hydroxyhexadecanoic and 2 hydroxydodecanoic acids, all in amide linkage. The predominant ester-linked fatty acyl group in acyloxyacyl linkage was hexadecanoate. The purified LPS, bisphosphoryl lipid A, and MPLA from B. abortus showed about 14%, 3% and 1%, respectively, of the B cell mitogen activity of ReLPS from Escherichia coli at 1.0 μg/ml.
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