PhosphoLIMBO: An Easy and Efficient Protocol to Separate and Analyze Phospholipids by HPTLC From Plant Material

Phospholipids are major structural and regulatory elements of biological membranes and are involved in many different cellular and physiological processes. In this protocol, we provide an easy, cost-effective, and efficient method to obtain an overview of the phospholipid composition using high-performance thin layer chromatography (HPTLC). While the currently known phospholipid separation methods based on HPTLC display co-migration of certain lipid classes, the method we describe here allows the separation of all phospholipid classes, including anionic phospholipids in plant samples. This protocol combines elements of the classical Vitiello and Touchstone solvent systems to optimize phospholipid separation in a scaled pattern. Here, we provide a full characterization of this method, including statistical analyses of the retention factor of each phospholipid to show the robustness of the method and its efficiency in separating all phospholipid classes of a biological sample. Key features • Analysis of phospholipid composition through an easy, fast, robust, and cost-effective HPTLC method. • Separation of anionic phospholipids from the other phospholipid species. • Full overview of all phospholipids categories, including anionic phospholipids. • Qualitative and quantitative approach.