公开(公告)号：US08563777B2

公开(公告)日：2013-10-22

申请号：US13155682

申请日：2011-06-08

申请人： Lloyd M. Smith ,  Michael R. Shortreed ,  Brian L. Frey ,  Margaret F. Phillips ,  Joshua J. Coon ,  Shane M. Lamos ,  Casey J. Krusemark ,  Peter J. Belshaw ,  Madhusudan Patel ,  Neil L. Kelleher

发明人： Lloyd M. Smith ,  Michael R. Shortreed ,  Brian L. Frey ,  Margaret F. Phillips ,  Joshua J. Coon ,  Shane M. Lamos ,  Casey J. Krusemark ,  Peter J. Belshaw ,  Madhusudan Patel ,  Neil L. Kelleher

IPC分类号： C07C211/10 ,  C07C211/11 ,  G01N33/58

CPC分类号： C07B59/001 ,  C07B59/00 ,  C07B59/008 ,  G01N30/7233 ,  G01N2458/15 ,  Y10T436/163333 ,  Y10T436/17

摘要： Relative quantification of metabolites by Electrospray Ionization Mass Spectrometry (ESI-MS) requiring a mechanism for simultaneous analysis of multiple analytes in two or more samples. Labeling reagents that are reactive to particular compound classes and differ only in their isotopic compositions facilitate relative quantification. Heavy and light isotopic forms of methylacetimidate were synthesized and used as labeling reagents for quantification of amine-containing molecules. Heavy and light isotopic forms of formaldehyde and cholamine were also synthesized and used independently as labeling reagents for quantification of amine-containing and carboxylic acid-containing molecules, such as found in biological samples. The labeled end-products are positively charged under normal acidic conditions involving conventional Liquid Chromatography Mass Spectrometry (LC/MS) applications. Labeled primary and secondary amine and carboxylic acid end-products generated higher signals concerning mass-spectra than pre-cursor molecules and improved sensitivity. Improved accuracy concerning relative quantification was demonstrated by mixing heavy and light labeled Arabidopsis extracts in different ratios.

摘要翻译： 通过电喷雾离子化质谱（ESI-MS）对代谢物进行相对定量，其需要同时分析两个或多个样品中多种分析物的机制。 对特定化合物类别具有反应性且仅在其同位素组成上不同的标记试剂便于相对定量。 合成了甲基乙酰亚胺酸的重和轻同位素形式，并用作定量含胺分子的标记试剂。 还合成了重和轻的同位素形式的甲醛和胆胺，并且独立地用作用于定量含胺和含羧酸的分子的标记试剂，例如在生物样品中发现的。 标记的最终产物在常规的液相色谱质谱（LC / MS）应用的正常酸性条件下带正电。 标记的伯胺和仲胺和羧酸终产物产生比前体分子更高的质谱信号，提高了灵敏度。 通过以不同的比例混合重和轻标记的拟南芥提取物来证明相对定量的改进的准确性。

公开(公告)号：US07982070B2

公开(公告)日：2011-07-19

申请号：US11726307

申请日：2007-03-21

申请人： Lloyd M. Smith ,  Michael R. Shortreed ,  Brian L. Frey ,  Margaret F. Phillips ,  Joshua J. Coon ,  Shane M. Lamos ,  Casey J. Krusemark ,  Peter J. Belshaw ,  Madhusudan Patel ,  Neil L. Kelleher

发明人： Lloyd M. Smith ,  Michael R. Shortreed ,  Brian L. Frey ,  Margaret F. Phillips ,  Joshua J. Coon ,  Shane M. Lamos ,  Casey J. Krusemark ,  Peter J. Belshaw ,  Madhusudan Patel ,  Neil L. Kelleher

IPC分类号： C07C211/10 ,  C07C211/11 ,  G01N33/58

CPC分类号： C07B59/001 ,  C07B59/00 ,  C07B59/008 ,  G01N30/7233 ,  G01N2458/15 ,  Y10T436/163333 ,  Y10T436/17

摘要： Relative quantification of metabolites by Electrospray Ionization Mass Spectrometry (ESI-MS) requiring a mechanism for simultaneous analysis of multiple analytes in two or more samples. Labeling reagents that are reactive to particular compound classes and differ only in their isotopic kit facilitating relative quantification and providing tangible evidence for the existence of specific functional groups. Heavy and light isotopic forms of methylacetimidate were synthesized and used as labeling reagents for quantification of amine-containing molecules, such as biological samples. Heavy and light isotopic forms of formaldehyde and cholamine were also synthesized and used independently as labeling reagents for quantification of amine-containing and carboxylic acid-containing molecules, such as found in biological samples. Advantageously, the labeled end-products are positively charged under normal acidic conditions involving conventional Liquid Chromatography Mass Spectrometry (LC/MS) applications. Labeled primary and secondary amine and carboxylic acid end-products also generated higher signals concerning mass-spectra than pre-cursor molecules and improved sensitivity. Improved accuracy concerning relative quantification was achieved by mixing heavy and light labeled Arabidopsis extracts in different ratios. Labeling strategy was further employed to ascertain differences in the amounts of amine-containing metabolites for two strains of Arabidopsis seeds.

摘要翻译： 通过电喷雾离子化质谱（ESI-MS）对代谢物进行相对定量，其需要同时分析两个或多个样品中多种分析物的机制。 标记对特定化合物类别具有反应性的试剂，仅在其同位素试剂盒中有所不同，便于相对定量，并为特定官能团的存在提供有形证据。 合成了甲基亚酰亚胺酸的重和轻同位素形式，并用作定量含胺分子（如生物样品）的标记试剂。 还合成了重和轻的同位素形式的甲醛和胆胺，并且独立地用作用于定量含胺和含羧酸的分子的标记试剂，例如在生物样品中发现的。 有利地，标记的最终产物在常规的液相色谱质谱（LC / MS）应用的正常酸性条件下带正电荷。 标记的伯胺和仲胺和羧酸终产物也产生了比前体分子更高的质谱信号，提高了灵敏度。 通过以不同比例混合重和轻标记的拟南芥提取物，提高了相对定量的准确性。 进一步使用标记策略来确定两种拟南芥种子的含胺代谢物的量的差异。

公开(公告)号：US08592216B2

公开(公告)日：2013-11-26

申请号：US12759950

申请日：2010-04-14

申请人： Brian L. Frey ,  April L. Jue ,  Casey J. Krusemark ,  Lloyd M. Smith ,  Joshua J. Coon

发明人： Brian L. Frey ,  April L. Jue ,  Casey J. Krusemark ,  Lloyd M. Smith ,  Joshua J. Coon

IPC分类号： G01N37/00

CPC分类号： C07K1/13

摘要： The present invention provides methods for enhancing the fragmentation of peptides for mass spectrometry by modifying the peptides with a tagging reagent containing a functional group, such as a tertiary amine, having a greater gas-phase basicity than the amide backbone of the peptide. These high gas-phase basicity functional groups are attached to a peptide by reacting the tagging reagent to one or more available carboxylic acid groups of the peptide. Linking these high gas-phase functional groups to the peptides leads to higher charge state ions from electrospray ionization mass spectrometry (ESI-MS), which fragment more extensively during fragmentation techniques, particularly non-ergodic fragmentation techniques such as electron capture dissociation (ECD) and electron transfer dissociation (ETD).

摘要翻译： 本发明提供了通过用含有与肽的酰胺主链相比具有更大的气相碱度的官能团的标记试剂（例如叔胺）修饰肽来增强肽质谱分离的方法。 这些高气相碱性官能团通过使标记试剂与肽的一个或多个可用的羧酸基团反应而连接到肽上。 将这些高气相官能团连接到肽导致来自电喷雾离子化质谱（ESI-MS）的更高电荷状态离子，其在片段化技术期间更广泛地进行片段，特别是非遍历断裂技术，例如电子俘获解离（ECD） 和电子转移离解（ETD）。

公开(公告)号：US20100330680A1

公开(公告)日：2010-12-30

申请号：US12759950

申请日：2010-04-14

申请人： Brian L. Frey ,  April L. Jue ,  Casey J. Krusemark ,  Lloyd M. Smith ,  Joshua J. Coon

发明人： Brian L. Frey ,  April L. Jue ,  Casey J. Krusemark ,  Lloyd M. Smith ,  Joshua J. Coon

IPC分类号： G01N37/00 ,  C07K1/13 ,  C07K1/14 ,  C07K2/00

CPC分类号： C07K1/13

摘要： The present invention provides methods for enhancing the fragmentation of peptides for mass spectrometry by modifying the peptides with a tagging reagent containing a functional group, such as a tertiary amine, having a greater gas-phase basicity than the amide backbone of the peptide. These high gas-phase basicity functional groups are attached to a peptide by reacting the tagging reagent to one or more available carboxylic acid groups of the peptide. Linking these high gas-phase functional groups to the peptides leads to higher charge state ions from electrospray ionization mass spectrometry (ESI-MS), which fragment more extensively during fragmentation techniques, particularly non-ergodic fragmentation techniques such as electron capture dissociation (ECD) and electron transfer dissociation (ETD).

摘要翻译： 本发明提供了通过用含有与肽的酰胺主链相比具有更大的气相碱度的官能团的标记试剂（例如叔胺）修饰肽来增强肽质谱分离的方法。 这些高气相碱性官能团通过使标记试剂与肽的一个或多个可用的羧酸基团反应而连接到肽上。 将这些高气相官能团连接到肽导致来自电喷雾离子化质谱（ESI-MS）的更高电荷状态离子，其在片段化技术期间更广泛地进行片段，特别是非遍历断裂技术，例如电子俘获解离（ECD） 和电子转移离解（ETD）。