A Validated GC‐FID Method for Determination of Very Long‐Chain Polyunsaturated Fatty Acids (C24–C30 <i>n</i>  − 3) in Triglyceride Fish Oils

ABSTRACT A quantitative and in‐house validated gas chromatography–flame ionization detector (GC‐FID) method for the analysis of methyl esters of very long chain (C24–C30, 4–8 double bonds) polyunsaturated fatty acids (VLC‐PUFAs) in triglyceride (TG) fish oils has been developed. Quantification of VLC‐PUFA was obtained by use of empirical response factors relative to the long‐chain PUFA DHA. The identification of VLC‐PUFA was achieved using GC‐positive chemical ionization‐mass spectroscopy (PCI‐MS) and electron ionization‐mass spectroscopy (EI‐MS) fragmentation patterns. Method performance parameters have been investigated: Results showed a high linearity ( r = 0.999, n = 15, p &lt; 0.001), an overall accuracy/recovery of 98% ± 0.3%, repeatability of 0.8%, and precision of 1.7%. Limit of detection of individual VLC‐PUFA was in the range of 0.2 mg/g ( S / N = 3). The results have shown that the GC‐FID method is applicable for quantifying the levels of individual VLC‐PUFA and the total amount of VLC‐PUFA levels in TG fish oils in the range of 20–70 mg/g fish oil. Practical Applications : Our results have shown that it is possible to obtain a routine quantitative determination of the methyl esters of the individual VLC‐PUFA in triglyceride fish oils after an initial GC‐EI‐MS and GC‐PCI‐MS confirmation of the fatty acid structures. Investigation of method performance parameters has shown that our requirements for linearity, accuracy, repeatability, precision, and limit of quantification and detection have been met. The results show that the GC‐FID method is able to quantify the total amount of VLC‐PUFA levels in triglyceride fish oils in the range of 20–70 mg/g fish oil. This method may represent a useful tool for determining VLC‐PUFA content in commercial fish oil products. The method could also have the potential to be used with other types of lipids, such as ethyl esters, free fatty acids, phospholipids, wax esters, and even extracts of biological samples.