Fatty acid profiling provides information on dietary intakes and an understanding of lipid metabolism. High throughput techniques such as fingertip prick (FTP) sampling has gained popularity in recent years as a simplified method for basic research, and could be further used to assess disease risk in the population, and other similar high-throughput techniques have the potential to assist in the monitoring and labeling of fatty acids in the food supply. With the advancement of high-throughput sample analysis techniques, a more complete understanding of storage stability is required as a larger volume of samples are produced with equal amounts of time to analyze them. Energy-assisted analysis techniques have the potential to help ameliorate some of these issues. Presently, FTP blood, whole blood and salmon storage stability is assessed under various storage conditions, and both microwave-assisted direct transesterification and indirect ultrasound-assisted extraction techniques are assessed. It is determined that storage of FTP blood and whole blood samples at -20°C results in significant and nearly complete highly unsaturated fatty acid (HUFA) degradation compared to all other temperatures examined. This degradation is determined to be the result of hemolysis and subsequent iron release from erythrocytes initiating fatty acid peroxidation reactions. Direct transesterification of FTP blood is reduced from as long as three hours to one minute with microwave-assisted energy and fatty acid extraction from ground flaxseed is reduced to 40 minutes from as long as 24 hours without compromising fatty acid profiles. Results of the current study provides insight into the storage stability of food sample and blood samples collected via high-throughput techniques, and provides support for the utilization of further high-throughput energy-assisted analytical methods that can help to minimize the potentially detrimental effects that long-term storage can have on fatty acid profiles.

The determination and accurate identification of fatty acids (FAs) in biological samples remain challenging due to the complexity of these compounds and confounding sample matrix effects. This thesis highlights a range of method developments and applications using advanced multidimensional gas chromatography (MDGC) and comprehensive two dimensional gas chromatography (GCxGC) techniques, coupled with mass spectrometry (MS) for FA profiling and structural identification.An investigation of appropriate combinations of stationary phases was studied prior to the subsequent multidimensional analytical method development. Recently introduced ionic liquid (IL) capillary GC stationary phases covering a wide polarity range have been evaluated for fatty acid methyl esters (FAMEs) analysis in terms of their elution patterns and retention indices (e,g. ECL) on the IL columns.The performance of a newly proposed integrated GCxGC / heart-cut (H/C) MDGC-FID method for isomeric PUFA separation was then evaluated. As a practical application, abundant long chain UFA isomers with carbon numbers ranging from 18 to 22 in marine oil, as well as a dairy food product, can almost be fully separated by using GCxGC. Additionally, more than 7 other FA compounds were found in the same region by switching the system to H/C MDGC-FID under a modified GC condition. Both these applications employed fast analysis conditions.Next, GCxGC was coupled to the quadrupole-accurate mass time of flight mass spectrometry (QTOFMS). This was done to achieve accurate detection and structural confirmation of individual known or unknown branched FA obtained through predicted empirical formula and accurate mass information. The approach was applied to phospholipid fatty acids (PLFAs) from complex forest soil samples in order to investigate the microbial community. Several high abundance branched hydroxyl- (OH-), iso-/anteiso- and cyclopropyl- (cy-) FAME were clearly determined. Tentative identities of trace level OH-FAME and unusual epoxidised FAME were found according to their predicted empirical formulae and elemental compositions. Epoxidised FA were previously found in the metabolic pathway of a Gram-positive soil bacterium Bacillus megaterium. The findings indicate that this approach has great potential towards fingerprinting and FA biomarker discovery in environmental research. Lastly, comprehensive fast GCxMS with a positive chemical ionisation mode (PCI) was proposed for FA analysis in medical studies. Method performance was evaluated by application to different types of FAs including PUFAMEs and bacterial FAMEs (e.g. branched FAMEs) as well as to real biological samples (e.g. wild type and transgenic mouse brains associated with Alzheimer's disease). Methanol was selected as the most appropriate PCI chemical reagent for total FA profiling. A comparison of both rapid comprehensive GCxPCI-QMS and GCxGC approaches, for fast screening capabilities and the rapid quantitative determination of long chain PUFA isomers in mice brain associated with Alzheimer's disease, was conducted in this study. An evaluation of both methods was based on the comparison of their 2D presentations, calibration linearity and minimum levels of detection. Fast GCxMS data demonstrated the good quantification potential of this approach towards biological FA analysis, especially in medical studies.

Gas chromatography was evaluated to determine its usefulness to analyze fatty acids in natural food products. Several different types of foods were analyzed including liquid and solid materials. Because of its simplicity, speed, efficiency and minimal costs, the direct transesterification method was found to be superior to previously reported methods and thus is a very useful analytical procedure.