Rapid separation of neutral lipids, free fatty acids and polar lipids using prepacked silica sep‐Pak columns

TLDR

The study presents a rapid method for separating neutral lipids, free fatty acids, and polar lipids using small prepacked silica Sep‑Pak columns. The method employs sequential elution with hexane/methyl‑tert‑butyl ether to isolate cholesteryl esters and triglycerides, acidification to release fatty acids and cholesterol, and a final step with methyl‑tert‑butyl ether, methanol, and ammonium acetate to elute polar lipids. The technique achieves recoveries of ≥96 % for neutral lipids and ≥98 % for phospholipids, though phosphatidylethanolamine and phosphatidylinositol co‑elute, and phosphatidylcholine, sphingomyelin, and lysophosphatidylcholine form a second fraction.

Abstract

Abstract A method is described for the separation of neutral lipid, free fatty acid and polar lipid classes using small (600 mg), prepacked silica Sep‐Pak columns. Combinations of hexane and methyltertiarybutylether were used to progressively elute cholesteryl ester first then triglyceride from the column. After column acidification, fatty acids were eluted followed by cholesterol. Recoveries of these lipids were 96% or greater. Polar lipids were eluted from the column using combinations of methyltertiarybutylether, methanol and ammonium acetate. Phospholipid classes could not be separated completely from each other. Phosphatidylethanolamine and phosphatidylinositol eluted together, whereas the more polar phosphatidylcholine, sphingomyelin and lysophosphatidylcholine were eluted as a second fraction. Recoveries of each phospholipid was greater than 98%.