Abstract: The phospholipids in pasteurized milk samples were analyzed and identified by ultra-high performance liquid chromatography-triple quadrupole-time of flight tandem mass spectrometry (UPLC-Triple-TOF MS/MS). The chromatographic separation was performed on an ACQUITY UPLC? BEH C18 (2.1 mm × 100 mm, 1.7 μm) column using gradient elution with water/methanol/acetonitrile (1:1:1, V/V) as mobile phase A and isopropanol/acetonitrile (1:1, V/V) as mobile phase B. Data acquisition was done in the positive ion mode using electrospray ionization (ESI). According to the high-resolution mass spectral data of retention time, quasi-molecular ions and secondary fragments combined with information on the mass spectral fragmentation pathways of reference standards, a total of 68 phospholipid molecules were identified including 22 phosphatidylcholine (PC, 7.98%), 17 phosphatidylethanolamine (PE, 56.60%), 10 phosphatidylserine (PS, 1.70%), 7 phosphatidylinositol (PI, 1.33%), and 12 sphingomyelin (SM, 32.39%). This study achieved comprehensive qualitative and quantitative determination of phospholipids in milk by using the UPLC-Triple-TOF MS/MS method with high sensitivity, precision and stability. This method could detect more phospholipids in milk than other detection techniques and thus provide more basic data for research on milk phospholipids.

Key words: pasteurized milk; phospholipid; phospholipidomics; ultra-high perfomance liquid chromatography-triple quadrupole-time of flight tandem mass spectrometry