Lipopolysaccharides spectrometry

Li, J. Thibault, P. Martin, A. Richards, J. C. Wakarchuk, W. W. der Wilp, W. Development of an on-line preconcentration method for the analysis of pathogenic lipopolysaccharides using capillary electrophoresis-electrospray mass spectrometry—Application to small colony isolates. J. Chromatogr. A 1998, 817, 325-336. 

B. Lindner, U. Zahringer, E. Th. Rietschel, and U. Seydel, in A. Fox, S. L. Morgan, L. Larsson, and G. Odham (Eds.), Analytical Microbiology Methods Structural Elucidation of Lipopolysaccharides and Their Lipid A Component Application of Soft Ionization Mass Spectrometry, p. 149. Plenum, New York/London, 1990. 

Li J, Purves RW, Richards JC. 2004a. Coupling capillary electrophoresis and high-field asymmetric waveform ion mobility spectrometry mass spectrometry for the analysis of complex lipopolysaccharides. Anal Chem 76 4676. 

Total fatty acids were liberated by subjecting Salmonella minnesota Re lipopolysaccharide (or free lipid A) to acidic (4 N HC1, 5 h, 100°C) followed by alkaline (1 N NaOH, 1 h, 100°C) hydrolysis. After extraction (chloroform), the free fatty acids were converted into their methyl esters (diazomethane) and analysed by combined gas-liquid chromatography/mass spectrometry. Alternatively, the fatty acids of lipid A are transesterified by treatment of lipopolysaccharide with methanolic HC1 (2 N HC1 in water-free CHaOH, 18 h, 85°C). By these procedures the following fatty acids were identified (in approximate amounts relative to 2 moles glucosamine) dodecanoic (12 0, 1.1 mole), tetradecanoic (14 0, 0.8 mole), hexadecanoic (16 0, 0.9 mole), 2-hydroxytetradecanoic (2-OH-l4 0, 0.1 mole), and 3-hydroxytetradecanoic acid (3-OH-14 0, 4 moles). In total, therefore, approximately 7 moles of fatty acids are present per mole of lipid A backbone. The stereochemistry of the hydroxylated fatty acids was determined by gas-liquid chromatography of their diastereomeric methoxyacyl-L-phenylethylamide derivatives (24). It was found that 2-hydroxyte-tradecanoic acid possesses the-Ts), and the predominating 3-hydroxytetradecanoic acid the (R) configuration. 

A. Gamian, E. Romanowska, A. Romanowska, C. Lugowski, J. Dabrowski, and K. Trauner, Citrobacter lipopolysaccharides Structure elucidation of the O-specific polysaccharide from strain PCM 1487 by mass spectrometry, one-dimensional and two-dimensional H NMR spectroscopy and methylation analysis, Eur. J. Biochem., 146 (1985) 641-647. 

Li, J., Cox, A.D., Hood, D., Moxon, E.R., Richards, J.C. Application of capillary electrophoresis-electrospray-mass spectrometry to the separation and characterization of isomeric lipopolysaccharides of Neisseria meningitidis. Electrophoresis 25 (2004) 2017-2025. 

Note. ELISA = enzyme-linked immunosorbent assay, FACS = fluorescence-activated cell sorting, HPLC = high-performance liquid chromatography, IHC = immunohistochemistry, LC/MS = liquid chromatography/MS = mass spectrometry, LPS = lipopolysaccharide (endotoxin), NA = not applicable, PAHA = (nonhuman) primate anti-human antibodies. 

Electrospray mass spectrometry has developed into a well-established method of wide scope and potential over the past 15 years. The softness of electrospray ionization has made this technique an indispensable tool for biochemical and biomedical research. Electrospray ionization has revolutionized the analysis of labile biopolymers, with applications ranging from the analysis of DNA, RNA, oligonucleotides, proteins as well as glycoproteins to carbohydrates, lipids, gly-colipids, and lipopolysaccharides, often in combination with state-of-the-art separation techniques like liquid chromatography or capillary electrophoresis [1,2]. Beyond mere analytical applications, electrospray ionization mass spectrometry (ESMS) has proven to be a powerful tool for collision-induced dissociation (CID) and multiple-stage mass spectrometric (MSn) analysis, and - beyond the elucidation of primary structures - even for the study of noncovalent macromolecular complexes [3]. 

Sonesson A., Larsson L., Fox A., Westerdahl G. and Odham G. (1988) Determination of environmental levels of peptidoglycan and lipopolysaccharide using gas chromatography-mass spectrometry utilizing bacterial amino acids and hydroxy fatty acids as biomarkers. J. Chromatogr. Biomed. Appl., 431,1-15. 

Sonesson A., Larsson L., Schiitz A., Hagmar L. and Hallberg T. (1990) Comparison of the Limulus amebocyte lysate test and gas chromatography-mass spectrometry for measuring lipopolysaccharides (endotoxins) in airborne dust from poultry processing industries. Appl. Environ. Microbiol., 56, 1271-78. 

Unfortunately, some of the same physiochemical characteristics make quantitation of NO difficult. Most studies in vivo and in vitro have used indirect indices of NO production such as breakdown products of NO metabolism, second messengers, or biological events in effector systems. Recent developments in probe technology and mass spectrometry have allowed the direct detection of NO in some situations. Constitutive NO synthases (cNOS) in vascular endothelium and other tissues produce small quantities of NO for continuous maintenance of vascular tone and cellular communication. Inducible NO synthases (iNOS) are induced by immunological stimuli such as interleukins and lipopolysaccharides, and synthesize relatively large amounts of NO for long periods. In general, therefore, it is much more difficult to study constitutive than inducible NO production. 

Analysis of a hydrolyzate of the lipopolysaccharide from Pasteurella pseudotuberculosis Group IIA, by g.l.c. of the derived alditol acetates, revealed an unknown component. Mass spectrometry of the alditol acetate obtained after reduction with sodium borodeuteride showed a fragmentation mode compatible with the alditol acetate from a 6-deoxy-heptose. The sugar was subsequently identified as 6-deoxy-D-manno-heptose. ... 

The chemical structure of lipid A of lipopolysaccharide isolated from Comamonas testosteroni was recently determined by lida et al. (1996) by means of methylation analysis, mass spectrometry and NMR. The lipid A backbone was found to consist of 6-0-(2-deoxy-2-amino-P-D-glucopyrano-syl)-2-deoxy-2-amino-alpha-D-glucose which was phosphorylated in positions 1 and 4. Hydroxyl groups at positions 4 and 6 were unsubstituted, and position 6 of the reducing terminal residue was identified as the attachment site of the polysaccharide group. Fatty acid distribution analysis and ES/MS of lipid A showed that positions 2,2, 3 and 3 of the sugar backbone were N-acylated or O-acylated by R-3-hydroxydecanoic acid and that the hydroxyl groups of the amide-linked residues attached to positions 2 and 2 were further O-acylated by tetradecanoic and dodecanoic acids, respectively. 

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