Add time:08/01/2019 Source:sciencedirect.com
Several conditions of acidic anhydrous methanolysis were examined to optimize the release and minimize the degradation of unphosphorylated 2-keto-3-deoxy-d-manno-octonic acid (KDO) from bacterial lipopolysaccharides and polysaccharides. The reaction was monitored by capillary gas chromatography after derivatization by trifluoroacetic anhydride. The best results were obtained by use of 2 M hydrochloric acid at 60°C for 2 h. Under these conditions a single KDO component appeared, and KDO was quantitatively released from all model compounds except when glycosidically linked to hexosamines. For quantitative cleavage of this linkage a reaction time of 6 h was required at 60°C, giving rise to 5/2-10% of secondary KDO products. The KDO detection limit was about 250 pmol (50 ng) and the molar response was the same as for glucose. The KDO derivative gave a mass spectrometric fragmentation pattern consistent with a pyranosidic methylketoside methyl ester structure. Differentation of KDO linkage types could be obtained by determination of the rates of KDO release by mild methanolysis.
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