Lipid extraction effects on stable isotope values (δ¹³C and δ¹⁵N) of elasmobranch muscle tissue

Given the known effect of lipid content on δ¹³C values and the potential effect of urea on δ¹⁵N values, examining the effects of lipid extraction, which can potentially extract both, is of particular importance for elasmobranch isotope ecology. Through analysing paired δ¹³C, total %C, δ¹⁵N, total %N and C:N values of non-lipid extracted (BULK) and lipid extracted (LE) muscle samples from twenty-one elasmobranch species, we assessed whether lipid extraction was required: (i) to remove lipids given reported low lipid content and, (ii) to determine if δ¹⁵N values were affected and whether this relates to the retention of isotopically light urea by elasmobranchs. The mean (±SD) δ¹³C values of eight out of twenty-one species significantly increased following lipid extraction with two species, the Greenland (Somniosus microcephalus) and whale (Rhincodon typus) shark, showing a marked increase (5.0±0.4‰ and 3.3‰, respectively). The mean (±SD) and maximum increase in δ¹³C values were 0.6±1.2‰ and 5.9‰, respectively. For δ¹⁵N data, thirteen species showed a significant increase following lipid extraction and a concomitant reduction in total percent nitrogen (%N). The C:N ratio for these species also increased from unexpectedly low values of <3.0 to ~3.0, the value expected for pure protein. The mean and maximum observed increase in δ¹⁵N values were 0.6±0.6‰ and 2.3‰, respectively. There was no effect of increasing animal size on δ¹³C and δ¹⁵N difference (LE–BULK) for the two species examined. Field sampled animals (sampled immediately upon capture in the marine environment) showed a greater δ¹⁵N difference than animals sampled in the laboratory (sampled several hours after capture in the marine environment) (1.0±0.5‰ and 0.4±0.4‰ respectively), while estuarine sampled animals (sampled immediately) showed the smallest difference (0.1±0.6‰). The δ¹³C data demonstrate that lipid extraction is required to remove lipids from elasmobranch muscle tissue given both intra- and inter- species variability. In addition, the increase in δ¹⁵N values, decrease in %N and increase in C:N ratio indicate that lipid extraction is removing soluble urea. Given lower δ¹⁵N diet-tissue discrimination factors for large marine predators, removal of urea is required to elucidate accurate trophic position estimates and relative food web position of elasmobranchs and for diet reconstruction. It is recommended that investigators undertake lipid extraction trials on elasmobranch muscle tissue to determine effects on δ¹³C and δ¹⁵N values on a species-by-species basis.