2.6 Stable isotope analysis
Ethanol-preserved liver tissues were rinsed in distilled water and freeze-dried for 48 h. Once freeze-dried, samples were ground to a fine powder using a laboratory bead-beater. Ground samples were weighed in 8 x 5 mm pressed standard weight tin capsules using a high precision microbalance (repeatability = 0.0008 mg). Elemental percentages of carbon, nitrogen, sulfur, and stable isotope ratios (δ13C, δ15N and δ34S) were measured using a Pyrocube elemental analyser (Elementar, Langenselbold, Germany) linked to a visION continuous-flow isotope ratio mass spectrometer (Elementar, Langenselbold, Germany) at the Universidad de Antofagasta Stable Isotope Facility (UASIF), Chile. Stable isotope ratios are expressed using δ notation and are reported in units of per mil (‰) relative to the following standards: Vienna Pee Dee Belemnite for carbon, air for nitrogen, and Vienna Canyon Diablo Troilite for sulfur. International standards were used in each batch to provide a multi-point calibration using the ionOS software package v4.1.005 (Elementar, Langenselbold, Germany). Certified reference material USGS40 and USGS41a were used for carbon and nitrogen and IAEA-SO-5, IAEA-SO-6 and IAEA-S2 for sulfur. Repeated analysis of standards showed analytical errors (± 1 SD) of ± 0.04 ‰ for δ13C, ± 0.06 ‰ for δ15N, and ± 0.6 ‰ for δ34S. We used two calibration standards, a) sulfonamide (Elementar, Germany) and b) an in-house standard (rainbow trout dorsal muscle) to correct for instrument drift.