An efficient separation and quantification of the individual neutral and polar lipid classes and their constituent fatty acids was achieved by the combination of two different detection techniques: Iatroscan TLC-FID and GC-FID. The solvent composition and ratio of development system, the sample size, the fidelity, and precision were tested in order to estimate the effectiveness of separation of individual neutral and polar lipid classes and the quantitative reproducibility of the Iatroscan TLC-FID technique. GC-FID method, with a high-quality capillary column, allowed sensitive and reproducible fatty acid qualitative and quantitative analyses, separation of fatty acid structural isomers (e.g., n-C16:0, iso-C16:0 and anteiso-C16:0), positional isomers (e.g., C18:1ω-9 and C18:1ω-7), geometrical isomers (cis-trans), and homologues (e.g., C16:0, C17:0, C18:0, etc.) in standards and complex lipid samples. Seventeen (17) lipid classes and fifty-two (52) saturated (SFA), monounsaturated (MUFA), and polyunsaturated (PUFA) fatty acids were identified and quantified, respectively, in samples of standard lipid and fatty acid mixtures, simulating the composition of natural lipids and their fatty acid methyl esters in common foods. The wide number of applications establishes this combination of Iatroscan TLC-FID and GC-FID methods as a powerful tool for lipid class and fatty acid analysis of any fat origin. 1. Introduction Fat and fatty acids especially polyunsaturated ones contribute to important aspects of fish, meat, and plant products’ quality and are critical for their nutritional and sensory value. Therefore, it is required to improve methods for the separation of total lipids into their neutral and polar fractions and for the analysis of their fatty acids (FA). Neutral lipid FA composition of animal and marine fat is strongly associated with their diet, whereas polar lipids regulate the function of membrane cells. Iatroscan is an instrument that combines thin-layer chromatography (TLC) resolution efficacy with the capacity of quantification by flame ionization detection (FID) [1, 2]. Neutral and polar lipids (e.g., triglycerides, sterols, phospholipids) cannot be separated by gas chromatography (GC) or high-performance liquid chromatography (HPLC) without fractionation, saponification, derivatization, or other pretreatment methods [3] and HPLC also requires availability of suitable columns for analysis of different polarity lipid. On the other hand, TLC/FID is adaptable and flexible as the separation can be achieved by adjusting polarity of solvent system
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