Four types of neutral glycosphingolipids (LacCer, Gb3Cer, Gb4Cer, and IV3αGalNAc-Gb4Cer; 10 pmol each) were analyzed using high-performance liquid chromatography (HPLC)-electrospray ionization quadrupole ion trap time-of-flight (ESI-QIT-TOF) mass spectrometry (MS) with a repeated high-speed polarity and MSn switching system. This system can provide six types of mass spectra, including positive and negative ion MS, MS2, and MS3 spectra, within 1 s per cycle. Using HPLC with a normal-phase column, information on the molecular weights of major molecular species of four neutral glycosphingolipids was obtained by detecting [M+Na]+ in the positive ion mode mass spectra and [M−H]− in the negative ion mode mass spectra. Sequences of glycosphingolipid oligosaccharide were obtained in the negative ion MS2 spectra. In addition, information on the ceramide structures was clearly obtained in the negative ion MS3 mass spectra. GlcCer molecular species were analyzed by HPLC-ESI-QIT-TOF MS with a reversed-phase column using 1 pmole of GlcCer. The structures of the seven molecular species of GlcCer, namely, d18:1-C16:0, d18:1-C18:0, d18:1-C20:0, d18:1-C22:0, d18:1-C23:0, d18:1-C24:1, and d18:1-C24:0, were characterized using positive ion MS and negative ion MS, MS2, and MS3. The established HPLC-ESI-QIT-TOF MS with MSn switching and a normal phase column has been successfully applied to the structural characterization of LacCer and Gb4Cer in a crude mixture prepared from human erythrocytes.

译文

四种类型的中性鞘糖脂 (LacCer,Gb3Cer,Gb4Cer和IV3αGalNAc-Gb4Cer; 使用高效液相色谱 (HPLC)-电喷雾电离四极杆离子阱飞行时间 (ESI-qit-tof) 质谱 (MS) 分析,并具有重复的高速极性和MSn切换系统。该系统可以在每个周期1 s内提供六种类型的质谱,包括正负离子MS,MS2和MS3光谱。使用带有正相色谱柱的HPLC,通过在正离子模式质谱中检测 [M Na] 和在负离子模式质谱中检测 [M-h]-,获得了有关四种中性鞘糖脂的主要分子种类的分子量的信息。在负离子MS2光谱中获得了鞘糖脂寡糖序列。此外,在负离子MS3质谱中清楚地获得了有关神经酰胺结构的信息。GlcCer分子种类通过hplc-esi-QIT-TOF MS用反相色谱柱使用1 pmole的GlcCer进行分析。GlcCer的七个分子物种的结构,即d18:1-C16:0,d18:1-C18:0,d18:1-C20:0,d18:1-C22:0,d18:1-C23:0,d18:1-C24:1和d18:1-C24:0,使用正离子MS和负离子MS,MS2和ms3进行表征。建立的具有MSn切换和正相柱的hplc-esi-QIT-TOF MS已成功应用于由人红细胞制备的粗混合物中LacCer和Gb4Cer的结构表征。

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