Influence of Bonded-Phase Column Type, Mobile Phase Composition, Temperature and Flow-Rate in the Analysis of Triglycerides by Reverse-Phase High Performance Liquid Chromatography

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Abstract
Two chromatographic systems for RP-HPLC analysis of triglycerides, operating under isocratic conditions using octadecylsilane and octylisilane bonded phases, are described. The influence of such chromatographic factors as bonded phase column type, mobile phase composition, temperature and flow rate on retention, analysis selectivity and efficiency, and separation of mixtures of homogeneous triglycerides was assessed. Linear relationships were established for the logarithm of the capacity factor and selectivity for each triglyceride in relation to temperature, the proportion of certain mobile phase components and flow rate. The octadecylsilane bonded phase was more selective when analyzing triglycerides with a partition number below 48, while octylsilane was appropiate for separating mixtures of long chain saturated triglycerides to the detriment of the resolution of triglycerides with low partition numbers. ACN/ACE/THF (58/38/4) was a suitable mobile phase for use with the octadecylsilane bonded phase, and ACN/THF /H2O (60/40/1) for the octylsilane bonded phase. A column temperature of 30°C and a flow rate of 1.5 mL/min resulted in acceptable resolution and analysis time in both systems.