ASE extraction method for simultaneous carbon and nitrogen stable isotope analysis in soft tissues of aquatic organisms

Since lipids are depleted in 13C relative to proteins and carbohydrates, variations in lipid composition among species and within individuals significantly influence δ13C and may result in misleading ecological interpretations. Whereas lipid extraction before IRMS analysis constitutes a way of stable isotope result lipid-normalisation, such a procedure was given up because of the un-controlled effects of the methods used (i.e., “Bligh & Dyer”, Soxhlet, etc.) on δ15N. The aim of this work was to develop a simple, rapid and efficient lipid extraction method allowing for simultaneous C and N stable isotope analysis in the biological soft tissues of aquatic organisms. The goal was to be free from the lipid influence on δ13C values without interfering with δ15N values. For that purpose, the modern automated pressurized liquid extraction technique ASE (accelerated solvent extraction) was selected. Eel muscles representative of a broad range of fat contents were extracted via ASE by using different semi-polar solvents (100% dichloromethane and 80% n-hexane/20% acetone) and by operating at different temperature (ambient temperature and 100 °C) and pressure (750 and 1900 psi) conditions. The results were discussed in terms of lipid extraction efficiency as well as δ13C and δ15N variability.