SEPARATION OF NEUTRAL LIPIDS BY HIGH-PERFORMANCE LIQUID-CHROMATOGRAPHY - QUANTIFICATION BY ULTRAVIOLET, LIGHT-SCATTERING AND FLUORESCENCE DETECTION

The recent increased use of cell cultures to model physiological event s, in particular signal transduction and traumatic injury, has produce d a need for a quantitative, high-performance liquid chromatographic s eparation of neutral lipid classes with a high degree of resolution an d reproducibility. We report an isocratic separation using a Phenomene x Selectosil silica column (5 mu m). Two solvents were used, 1.2% 2-pr opanol in n-hexane containing 0.1% acetic acid (90%) and n-hexane (10% ) at a dow-rate of 0.6 ml/min. Column temperature was maintained at 55 degrees C and this temperature was critical for baseline resolution o f 1,3-diacylglycerol and cholesterol. The use of 10% n-hexane permitte d the resolution of low polarity compounds such as butylated hydroxyto luene, triacylglycerols and cholesteryl esters. All of the detectors u sed produced standard curves with linearity over a wide concentration range.