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Updated: May 15, 2026
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Radiolabeling and Quantification of Cellular Levels of Phosphoinositides by High Performance Liquid Chromatography-coupled Flow Scintillation
Published on: January 6, 2016
Stop-flow comprehensive two-dimensional liquid chromatography combined with mass spectrometric detection for
Paola Dugo1, Nermeen Fawzy, Filomena Cichello
1Dipartimento di Scienze del Farmaco e dei Prodotti della Salute, University of Messina, viale Annunziata, 98168 Messina, Italy. pdugo@unime.it
Journal of Chromatography. A
|January 22, 2013
Summary
A new two-dimensional liquid chromatography method effectively characterizes phospholipid molecular species in milk and plasma. This advanced technique enhances separation and identification of complex lipid profiles for detailed analysis.
Area of Science:
- Analytical Chemistry
- Biochemistry
- Chromatography
Background:
- Phospholipids (PLs) are crucial biological molecules with diverse functions.
- Accurate characterization of individual phospholipid molecular species is essential for understanding biological processes and disease states.
- Existing chromatographic methods often face limitations in resolving complex phospholipid mixtures.
Purpose of the Study:
- To develop a novel comprehensive two-dimensional liquid chromatography (LC × LC) system for the detailed characterization of phospholipid molecular species.
- To establish an efficient method for separating and identifying phospholipids across six different classes.
- To demonstrate the system's capability using real-world biological samples.
Main Methods:
- Utilized a hydrophilic interaction liquid chromatography (HILIC) column as the first dimension ((1)D).
- Employed reversed-phase liquid chromatography (RP-LC) with a C18 column as the second dimension ((2)D).
- Integrated mass spectrometric detection (MS) and a two-position ten-port switching valve for fraction transfer under stop-flow conditions.
Main Results:
- Successfully developed and implemented a HILIC×RP-LC-MS system for phospholipid analysis.
- Demonstrated the system's effectiveness in separating phospholipid molecular species from Folch-extracted cow's milk and plasma samples.
- Identified up to 16 and 14 distinct phosphatidylcholine species in milk and plasma, respectively, using both positive and negative ionization modes.
Conclusions:
- The novel comprehensive two-dimensional LC system provides enhanced separation power for complex phospholipid mixtures.
- This method enables detailed characterization of phospholipid molecular species in biological matrices like milk and plasma.
- The HILIC×RP-LC-MS approach offers a robust platform for lipidomic research and biomarker discovery.
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