Supercritical fluid chromatography-mass spectrometry enables simultaneous measurement of all phosphoinositide regioisomers.

Phosphoinositide species, differing in phosphorylation at hydroxyls of the inositol head group, play roles in various cellular events. Despite the importance of phosphoinositides, simultaneous quantification of individual phosphoinositide species is difficult using conventional methods. Here we developed a supercritical fluid chromatography-mass spectrometry method that can quantify the molecular species of all seven phosphoinositide regioisomers. We used this method to analyze (1) profiles of phosphoinositide species in mouse tissues, (2) the effect of lysophosphatidylinositol acyltransferase 1-depletion on phosphoinositide acyl-chain composition in cultured cells, and (3) the molecular species of phosphatidylinositol-3-phosphate produced during the induction of autophagy. Although further improvement is needed for the absolute quantification of minor phosphoinositide regioisomers in biological samples, our method should clarify the physiological and pathological roles of phosphoinositide regioisomers at the molecular species level.