Quantitative analysis of phosphatidylcholine in rat liver tissue by nanoflow liquid chromatography/tandem mass spectrometry.

A quantitative method was developed for the determination of phosphatidylcholine (PC) species concentration using nanoflow LC-ESI-MS/MS. In this study, a calibration method is developed to determine the effect of PC carbon chain length on MS peak intensity. Using the multiple standard addition method, a relationship between the peak intensities of different PC species from nanoflow LC-MS and carbon chain length is established first using different injection amounts of PC standards. From this relationship, a calibration curve for each carbon chain length can be obtained for the concentration calculation. It was found that the MS peak area of PC species analyzed by nanoflow LC-MS linearly decreased with increased acyl carbon numbers, and that the effect of the degree of acyl chain unsaturation on MS peak intensity was minimized when the injection amount was maintained at less than 1 pmol. The method was applied for the quantitative calculation of 34 PC species from rat liver, which were identified from data-dependent MS/MS analysis during nanoflow LC separation.