Stop-flow Comprehensive Two-dimensional Liquid Chromatography Combined with Mass Spectrometric Detection for Phospholipid Analysis

A novel comprehensive two-dimensional liquid chromatographic (LCxLC) system for characterization of phospholipid (PL) molecular species belonging to six phospholipid classes was developed. To tackle such a task, a silica hydrophilic interaction liquid chromatography (HILIC) column was used as the first dimension (D1), and reversed-phase liquid chromatography (RP-LC) with a C18 column was used as the second dimension (D2) in combination with mass spectrometric detection. Fraction transfer from the D1 to the D2 was performed by means of a two-position ten-port switching valve, operated under stop-flow conditions. The capability of the investigated LCxLC approach was demonstrated in the separation of phospholipid molecular species contained in two Folch-extracted cow's milk and plasma samples.