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学科主题: 分析化学
题名: Qualitative and quantitative analysis of ceramide species in biological samples in an untargeted lipidomic profiling platform by high resolution LC-MS
作者: Li J(李佳) ;  Zhao XJ(赵欣捷) ;  Hu CX(胡春秀) ;  Chen SL(陈世礼) ;  Zhou LN(周丽娜) ;  Xu GW(许国旺)
会议文集: Proceeding of HPLC 2011
会议名称: 37th International Symposium on High Performance Liquid Phase Separations and Related Techniques
会议日期: 2011-10-8
出版日期: 2011
会议地点: 大连
通讯作者: 许国旺
出版者: 待补充
出版地: 待补充
合作性质: 墙报
部门归属: 1808
主办者: 中国化学会色谱专业委员会
摘要: Ceramide plays important roles in several aspects such as cell apoptosis, growth and differentiation, as well as signal transduction. Recently Ceramide is unveiled to be responsible for insulin resistance. Analysis of ceramides is challenging due to its low abundance. SPE (solid phase extraction) and mild alkaline hydrolysis are reported to be used to remove other lipids thus to enrich ceramides. Such methods are complicated and will lose information of other lipids. Here UHPLC-LTQ-Orbitrap was adopted for untargeted lipid profiling of cellular lipid extract. The UHPLC separation was performed on a Waters C8 column (1.7 μm, 2.1*100 mm) by gradient elution, and data was acquired in ESI(+) in resolution 30,000 after ESI source parameters were optimized by tuning with ceramide standard. Triple-quadrupole MS was employed for fragmentation of ceramide species. The quantification of ceramides was based on EIC in a low mass error tolerance window (<5 ppm), by comparing EIC area of ceramides with that of internal standard C17-ceramide spiked before lipid extraction. The dynamic range was more than 2.5 orders of magnitude for ceramide and its on-column LOQ was 0.23 pmol, such low LOQ benefited from significantly high S/N of high resolution EIC. Besides ceramides, other lipids, including LPC, LPE, PC, PE, SM, DG, TG, were also analyzed at the same time, thus a lipid global profile could be obtained. This method was applied in lipidomic analysis of human myotube cells incubated with 13C-labelled palmitate. In total 11 ceramides were separated and unambiguously identified. Both the normal form and the 13C-labelled form eluted at the same retention time, and the MS2 characteristic verified the de novo pathway of synthesis of ceramide, which makes the structural elucidation more confident. This developed platform shows great sensitivity and robustness for specific lipid analysis in an untargeted profiling way, and high mass accuracy combined with fragmentation pattern is advantageous for identification of unknown lipids.
英文摘要: Ceramide plays important roles in several aspects such as cell apoptosis, growth and differentiation, as well as signal transduction. Recently Ceramide is unveiled to be responsible for insulin resistance. Analysis of ceramides is challenging due to its low abundance. SPE (solid phase extraction) and mild alkaline hydrolysis are reported to be used to remove other lipids thus to enrich ceramides. Such methods are complicated and will lose information of other lipids. Here UHPLC-LTQ-Orbitrap was adopted for untargeted lipid profiling of cellular lipid extract. The UHPLC separation was performed on a Waters C8 column (1.7 μm, 2.1*100 mm) by gradient elution, and data was acquired in ESI(+) in resolution 30,000 after ESI source parameters were optimized by tuning with ceramide standard. Triple-quadrupole MS was employed for fragmentation of ceramide species. The quantification of ceramides was based on EIC in a low mass error tolerance window (<5 ppm), by comparing EIC area of ceramides with that of internal standard C17-ceramide spiked before lipid extraction. The dynamic range was more than 2.5 orders of magnitude for ceramide and its on-column LOQ was 0.23 pmol, such low LOQ benefited from significantly high S/N of high resolution EIC. Besides ceramides, other lipids, including LPC, LPE, PC, PE, SM, DG, TG, were also analyzed at the same time, thus a lipid global profile could be obtained. This method was applied in lipidomic analysis of human myotube cells incubated with 13C-labelled palmitate. In total 11 ceramides were separated and unambiguously identified. Both the normal form and the 13C-labelled form eluted at the same retention time, and the MS2 characteristic verified the de novo pathway of synthesis of ceramide, which makes the structural elucidation more confident. This developed platform shows great sensitivity and robustness for specific lipid analysis in an untargeted profiling way, and high mass accuracy combined with fragmentation pattern is advantageous for identification of unknown lipids.
内容类型: 会议论文
URI标识: http://cas-ir.dicp.ac.cn/handle/321008/116078
Appears in Collections:中国科学院大连化学物理研究所_会议论文

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Recommended Citation:
Li J,Zhao XJ,Hu CX,et al. Qualitative and quantitative analysis of ceramide species in biological samples in an untargeted lipidomic profiling platform by high resolution LC-MS[C]. 见:37th International Symposium on High Performance Liquid Phase Separations and Related Techniques. 大连. 2011-10-8.
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