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PreprintCC BYChemRxiv

Off-line LCxLC-Mass Spectrometry Workflow using CID and EAD for the Quantification and Comprehensive Structural Double Bond Characterization of Glycerophospholipids in Plasma Samples

  • Laura Gisela González Iglesias1,2
  • Renzo Picenoni3
  • Guenter Boehm3
  • Gérard Hopfgartner1,2
  1. 1Department of Inorganic and Analytical Chemistry
  2. 2University of Geneva
  3. 3CTC Analytics AG
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Abstract

Here we describe an automated workflow using a PAL Robotic Tool Changer (RTC) system for plasma lipid sample preparation and off-line two-dimensional liquid chromatography (LCxLC) separation. The orthogonal chromatographic platform couples hydrophilic interaction liquid chromatography (HILIC) in the first dimension with rapid reversed-phase liquid chromatography (RPLC) in the second dimension, facilitated by automated fraction collection at 10-second intervals into 96-well plates. Lipid profiling and structural identification strategies were evaluated using collision-induced dissociation (CID) and electron-activated dissociation (EAD) across data-dependent acquisition (DDA), dataindependent acquisition (DIA / SWATH), and high-resolution multiple reaction monitoring (HRMRM) modes. Standard Reference Material NIST SRM 1950 (Human Plasma) was employed for lipid quantification and double-bond characterization of phosphatidylcholines (PCs). By combining automated modified Matyas liquid-liquid extraction, and off-line LCxLC, a total of 115 PC’s lipids could be detected in SRM 1950 plasma sample. For 74 PC’s including isomers, double assignments could be proposed based on the EAD spectra.

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Cite

BibTeXRIS
BibTeX
@misc{iglesias2026offline,
  title = {{Off-line LCxLC-Mass Spectrometry Workflow using CID and EAD for the Quantification and Comprehensive Structural Double Bond Characterization of Glycerophospholipids in Plasma Samples}},
  author = {Laura Gisela González Iglesias and Renzo Picenoni and Guenter Boehm and Gérard Hopfgartner},
  howpublished = {ChemRxiv},
  year = {2026},
  month = oct,
  doi = {10.26434/chemrxiv.15009982/v1},
  url = {https://chemrxiv.org/doi/full/10.26434/chemrxiv.15009982/v1},
  note = {CC BY},
}
RIS
TY  - UNPB
TI  - Off-line LCxLC-Mass Spectrometry Workflow using CID and EAD for the Quantification and Comprehensive Structural Double Bond Characterization of Glycerophospholipids in Plasma Samples
AU  - Laura Gisela González Iglesias
AU  - Renzo Picenoni
AU  - Guenter Boehm
AU  - Gérard Hopfgartner
PY  - 2026
DA  - 2026/10/06
PB  - ChemRxiv
DO  - 10.26434/chemrxiv.15009982/v1
UR  - https://chemrxiv.org/doi/full/10.26434/chemrxiv.15009982/v1
AB  - Here we describe an automated workflow using a PAL Robotic Tool Changer (RTC) system for plasma lipid sample preparation and off-line two-dimensional liquid chromatography (LCxLC) separation. The orthogonal chromatographic platform couples hydrophilic interaction liquid chromatography (HILIC) in the first dimension with rapid reversed-phase liquid chromatography (RPLC) in the second dimension, facilitated by automated fraction collection at 10-second intervals into 96-well plates. Lipid profiling and structural identification strategies were evaluated using collision-induced dissociation (CID) and electron-activated dissociation (EAD) across data-dependent acquisition (DDA), dataindependent acquisition (DIA / SWATH), and high-resolution multiple reaction monitoring (HRMRM) modes. Standard Reference Material NIST SRM 1950 (Human Plasma) was employed for lipid quantification and double-bond characterization of phosphatidylcholines (PCs). By combining automated modified Matyas liquid-liquid extraction, and off-line LCxLC, a total of 115 PC’s lipids could be detected in SRM 1950 plasma sample. For 74 PC’s including isomers, double assignments could be proposed based on the EAD spectra.
ER  -