A comparison of carbon and nitrogen stable isotope ratios of fish tissues following lipid extractions with non-polar and traditional chloroform/methanol solvent systems

Stable isotope ratios act as chemical tracers of animal diet, and are used to study food web dynamics. Because carbon stable isotope values are influenced by tissue lipid content, a number of extraction methods have been used to remove lipid bias, but, in some species and tissues, extractions also alter nitrogen isotope values. We have analyzed delta(13)C and delta(15)N in Atlantic bluefin tuna liver and white muscle, and whole Atlantic herring, fish tissues covering a wide range of lipid content (bulk C:N 3.1-12.5). In order to compare delta(13)C and delta(15)N values from traditional chloroform/methanol extractions with non-polar solvent alternatives, we analyzed samples following (1) no treatment, (2) lipid removal using chloroform/methanol (2:1), and (3) Soxhlet extractions using chloroform, diethyl ether or hexane. Chloroform/methanol and chloroform extractions produced the lowest C:N values and highest delta(13)C values. In bluefin tuna, chloroform and hexane extractions significantly altered liver delta(15)N, and all methods significantly altered delta(15)N values in white muscle. Whole Atlantic herring delta(15)N was not altered by any extraction method, while the 2:1 chloroform/methanol extraction most completely removed fish tissue lipid components. Our results indicate that delta(15)N effects are not limited to common chloroform/methanol extractions and suggest that chloroform/methanol is the most effective extraction for delta(13)C correction. Given evidence for delta(15)N alteration among all tested methods, mathematical correction approaches should be further explored as an alternative to lipid correction.