Abstract
A high-performance liquid chromatography analysis method with an evaporative light-scattering detector has been developed for the separation and quantitative analysis of free fatty acids in biological matrices. Core-shell reversed-phase high-performance liquid chromatography separation of 10 free fatty acids is achieved within 10.5 min using a methanol/water (0.05% trifluoroacetic acid) eluent gradient. After optimization, the drift tube and nebulization temperature of the evaporative light-scattering detector was set at 35°C, nitrogen flow-rate at 1.1 standard liter per minute and column temperature at 25°C. All calibration curves showed good regression (r2 > 0.9975). A validation procedure following the International Conference on Harmonisation guidelines was implemented to certify the method. Relative standard deviations did not exceed 1.5% and 4.25% for repeatability and reproducibility respectively.
Highlights
The development of fast high-performance liquid chromatography (HPLC) methods for routine analysis is of interest for a rapid identification and quantification of free fatty acids (FFAs).Gas chromatography (GC) is obviously the most commonly used method for FFA analysis
A high-performance liquid chromatography analysis method with an evaporative light-scattering detector has been developed for the separation and quantitative analysis of free fatty acids in biological matrices
Core-shell reversed-phase high-performance liquid chromatography separation of 10 free fatty acids is achieved within 10.5 min using a methanol/water (0.05% trifluoroacetic acid) eluent gradient
Summary
The development of fast high-performance liquid chromatography (HPLC) methods for routine analysis is of interest for a rapid identification and quantification of free fatty acids (FFAs).Gas chromatography (GC) is obviously the most commonly used method for FFA analysis. There is a risk of structural degradation during this methylation process since it requires high temperature under alkaline conditions. For this reason, accuracy of GC analysis can be greatly affected when unstable molecules like long chain polyunsaturated FFAs need to be quantified [1]. The use of a non-absorbing solvent is required, limiting the choice of the mobile phase To overcome this drawback, different alternative detection systems can be implemented such as an evaporative light-scattering detector (ELSD). In ELSD, the solvent is evaporated and the only requirement is a low volatility of the compounds as compared to the mobile phase. For an optimal operation, several ELSD parameters need to be tuned even though ELSD optimization is seldom reported in literature
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