A rapid and sensitive procedure for determination of 5-N-acetyl neuraminic acid in lipopolysaccharides of Haemophilus influenzae:: a survey of 24 non-typeable H-influenzae strains
A rapid and sensitive procedure for determination of 5-N-acetyl neuraminic acid in lipopolysaccharides of Haemophilus influenzae:: a survey of 24 non-typeable H-influenzae strains
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DOI：
10.1016/s0008-6215(01)00242-7
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发表时间：
2001-10-15
影响因子：
3.1
通讯作者：
Schweda, EKH
中科院分区：
文献类型：
--
作者：
Bauer, SHJ;Månsson, M;Schweda, EKH
鉴于5-N-乙酰神经氨酸在细菌致病过程中的重要性，本文建立了一种灵敏、重复性好、可靠的测定脂多糖（LPS）中5-N-乙酰神经氨酸水平的方法，并应用于24株不同的非分型流感嗜血杆菌（NTHi）菌株。该方法包括通过高效阴离子交换色谱法与脉冲安培检测（HPAEC-PAD）的末端5-N-乙酰神经氨酸残基释放的神经氨酸酶处理O-脱酰LPS的分析。该过程相对较快，工具的努力是适度的。该程序的结果进行了比较，通过H-1 NMR和电喷雾电离质谱（ESI-MS）获得的数据。对24株NTHi菌株的LPS分析表明，5-N-乙酰神经氨酸是NTHi中LPS的共同成分。只有一个菌株（NTHi 432）没有显示任何唾液酸化。摩尔比（LPS /5-N-乙酰神经氨酸）在5/1和500/1之间。用HPAEC-PAD法检测了几株用其他方法（包括1H NMR和ESI-MS）不能检测到5-N-乙酰神经氨酸的菌株，结果表明这些菌株含有5-N-乙酰神经氨酸。该方法被施加到确定的水平的终端5-N-乙酰神经氨酸在LPS从NTHi菌株生长在不同的条件下，突变株含有非活性LPS生物合成基因。(C)2001爱思唯尔科技有限公司版权所有。
In view of the importance of 5-N-acetyl neuraminic acid in bacterial pathogenesis, a sensitive, reproducible and reliable method for the determination of 5-N-acetyl neuraminic acid levels in lipopolysaccharide (LPS) is described and applied to 24 different non-typeable Haemophilus influenzae (NTHi) strains. The method involves analysis by high-performance anion-exchange chromatography with pulsed amperometric detection (HPAEC-PAD) of terminal 5-N-acetyl neuraminic acid residues released by neuraminidase treatment of O-deacylated LPS. The procedure is relatively fast and the instrumental effort is moderate. The results of the procedure were compared with data obtained by H-1 NMR and electrospray ionisation-mass spectrometry (ESI-MS). The analysis of LPS from 24 NTHi strains showed that 5-N-acetyl neuraminic acid was found to be a common constituent of LPS in NTHi. Only one strain (NTHi 432) did not show any sialylation. Molar ratios (LPS /5-N-acetyl neuraminic acid) ranged between 5/1 and 500/1. Several strains in which no 5-N-acetyl neuraminic acid could be determined by other methods including 1H NMR and ESI-MS were shown to contain 5-N-acetyl neuraminic acid by this HPAEC-PAD procedure. The method was applied to determine levels of terminal 5-N-acetyl neuraminic acid in LPS from NTHi strains grown under different conditions and mutant strains containing inactive LPS biosynthetic genes. (C) 2001 Elsevier Science Ltd. All rights reserved.