Mass spectrometry based profiling of the salmon lipidome

Introduction: Lipid metabolism is important in salmon farming, both for energy and for omega-3 fatty acids in human diet. To complement mathematical modelling of metabolism in the Digital Salmon project (DigiSal), we have developed and applied analytical tools to characterize the salmon lipidome (SL). UPLC and UPC2 separation in combination with mass spectrometric (MS) based detection provide the most selective and sensitive instrumental platform for high resolution qualitative characterization of lipid profiles in biological samples. Importantly, this methodological platform can also be tuned for quantitative analysis. We illustrate its application to 100 samples from a feed switch trial with contrasting levels of long-chain polyunsaturated fatty acids, conducted both in fresh and salt water. Methods: Lipid samples were extracted using a two phase system (water-methanol-chloroform) and a Precellys bead homogenizer. The extracts were analysed using a Waters Acquity UPLC Synapt HDMS QTof mass spectrometer operated in MSE. A RP-UPLC method was established and samples were analysed twice, in both positive and negative ESI mode, for the most comprehensive coverage of the SL. Raw data were processed with a Progenesis software with embedded LipidBlast database for identification. Further, data were exported for multivariate analysis using Unscrambler and MetaboAnalyst. Results and Discussion: The poster will show the MS based lipid profiling workflow we have established for the salmon lipidome. This includes discussion of the challenges in robust chromatographic separation and identification of lipid species based on MS and MS/MS information. The lipid samples studied originate from a feeding trial using both vegetable and marine oils and incubation of the salmon in fresh water and sea water1. Utilizing the bead homegenizer the lipid extraction has proven to be very effective so we are able to extract lipidome of as tiny samples as even 5 mg. The extraction method has been applied to fish liver and muscle; nevertheless, it can easily be adopted to other similar tissue types. Our approach includes also a fast semi-quantitative determination of lipid classes in addition to the high resolution UPLC-MSE profiling. Usually around 2-3000 species (lipids and compounds of nonpolar and mid-polar character) are detected with the high resolution method while the semi-quant lipid method covers the ten most abundant lipid classes. A statistical data analysis of the results from this abovementioned salmon study unambiguously showed the differentiation among the individual groups and identified lipids with the highest impact on the group clustering. The data obtained represent the starting point for a comprehensive study on how the diet affects polyunsaturated fatty acids profile in salmons.