Abstract
Cetacean adipose tissues contain an extremely very wide variety of acyl-chains present in triacylglycerols and / or wax esters. In addition, changes in the lipid composition across organs suggest fine stratification. It therefore remains technically challenging to describe precisely the lipid organization of these tissues. In the present study, we used in parallel HR-MAS NMR (High Resolution Magic Angle Spinning Nuclear Magnetic Resonance) and GC (gas-chromatography) to characterize and quantify the lipids and fatty acyl-chains from the blubber and melon of two odontocete species. Both methods generated very similar compositions, but each presented clear advantages. While GC underestimated the amount of short branched fatty acyl-chains, which are specific to cetacean adipose tissues and most probably of primary importance for their functioning, HR-MAS NMR allowed for their exact quantification. Conversely, when HR-MAS NMR could only discriminate a few types of fatty acyl-chain families, GC unambiguously identified and quantified most of them. In addition, this technique allowed for the determination of the wax esters molecular species. Our results further suggest that the stratification of these adipose tissues relies on changes in the triacylglycerol to wax ester ratio and in the fatty acyl composition of triacylglycerols, but not on changes in the wax esters composition. Altogether, our data show that the complementarities of these two approaches result in lipid analyses of unprecedented precision, paving the way for the detailed description of the fatty acyl composition of cetacean adipose tissues and the understanding of their functioning.
Highlights
Adipose tissues having highly specialized functions are one of the major adaptations of cetaceans to the marine environment
Dephinidae, monodontidae and phocoenidae accumulate short isobranched fatty acids (FA) (i-FA) like IVA, while other odontocetes contain longer i-FA, the ziphiidae accumulating for example C10-C12 i-FA [10,20]
Concerning the melon, we found in both species that the contents of WE, IVA and i-FA were higher in the center than in the external layer, whereas this later was enriched in C16, C18 and very-long-chain FA, as previously described [9,12,21]
Summary
ChemicalsAll chemicals, solvents, and lipid standards/references were purchased from Merck (Darmstadt, Germany) or Sigma (St Louis, MO, USA).Origin of samplesSamples were taken from fresh carcasses of recently stranded animals provided by the French Stranding Network (described in [27]). Subsamples of 20 mm long, 5 mm wide and 5 mm thick were taken from frozen blubbers or melons, bisected, and each half was sent frozen to one of the analysis platforms (HR-MAS NMR and GC). A piece of tissue (about 10 mg) was placed in a 4 mm zirconium oxide (ZrO2) MAS rotor and D2O was added into the rotor for 2H field locking. Parameters were optimized: pulse calibration at PLW1 = 20W was determined and manual shimming was performed to obtain linewidth under 7 Hz. 1H HR-MAS NMR spectra were performed, at room temperature, with a 30 ̊ pulse, a 2 s delay, 64 scans (spectral width = 10 000 Hz, TD = 32 K and SI = 64 K) using a Watergate sequence. Three sub-samples were independently analyzed three times each, and manual integrations gave an error below 5%
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