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.