Isotopic values of sea otters (modern and archaeological) from Southeast Alaska and Northern Oregon and potential prey items from Southeast Alaska

Integrating pre-industrial datasets into management frameworks is critical for establishing ecologically relevant baselines for conservation. Here, we use isotopic analysis of archaeological and modern sea otter (Enhydra lutris) specimens to anticipate future impacts of recolonization. We focus on Southeast Alaska (SEAK) and northern Oregon, where sea otter populations are recolonizing and competing with macroinvertebrate fisheries or remain extirpated with translocations being considered, respectively. We measured bulk bone collagen δ13C and δ15N and essential amino acid δ13C values of archaeological sea otters, and bulk isotopic values of vibrissae from modern SEAK sea otters. We compare these results with data of potential prey and archaeological samples from California. Isotopic data from SEAK reveal pre-industrial populations consumed infaunal bivalves and utilized both soft-sediment (33%) and kelp forest habitats (67%), with variation among sub-regions. If contemporary populations expand into this historical niche, conflict with Indigenous subsistence fisheries (bivalves) is likely to persist. Isotopic data of archaeological sea otters from northern Oregon suggest past consumption of low trophic level invertebrates, and a reliance on kelp forests (88%) rather than soft-sediment habitats, underscoring the significance of kelp forests for future translocations. Our work provides important perspectives on the potential future ecology of a recovering keystone predator.

Methods
Bulk tissue δ13C and δ15N values, along with weight percent [C] and [N], were measured via EA-IRMS using a Costech 4010 elemental analyzer (Valencia, CA) coupled to a Thermo Scientific Delta V Plus isotope ratio mass spectrometer (Bremen, Germany) at the University of New Mexico Center for Stable Isotopes (UNM-CSI; Albuquerque, NM). All samples and reference materials were calibrated against the internationally accepted standards for stable carbon and nitrogen isotope ratios, respectively Vienna-Pee Dee Belemnite (V-PDB) and atmospheric N2 . We report all isotopic data as δ values in parts per thousand, or per mil (‰), where δ13C or δ15N = 1000*[(Rsamp/Rstd)  1]. Here, Rsamp and Rstd are the 13C:12C or 15N:14N ratios of the sample and standard, respectively. The standard deviations of organic in-house reference materials (milk casein and tuna muscle) were 0.2 ‰ for both δ13C and δ15N values. As a control for the quality of our collagen samples, we converted measured weight [C]:[N] to atomic ratios: [C]:[N]atomic = [C]:[N]weight ∗ (14/12). 

The carbon isotope compositions of individual amino acids within a subset of archaeological sea otter samples (n=35) were measured via a gas chromatography-combustion system (GC-C; Thermo Scientific Trace 1310 and Isolink II) coupled to an IRMS at UNM-CSI. Derivatized samples and in-house amino acid reference mixtures were dried down under a gentle stream of N2 gas and brought up to final dilution in DCM. Sample DCM solutions were injected (1 mL) into a 60 m BPx5 gas chromatograph column (0.32 ID, 1.0 mm film thickness, SGE Analytical Science, Victoria, Australia) for amino acid separation (Trace 1310), and then combusted into CO2 gas (Isolink II). Isotopic ratios were analyzed with a Thermo Scientific Delta V IRMS. Samples were run in duplicate and bracketed with the in-house reference material; we report sample means across injections, and we discarded data where standard deviations >1.2 ‰. We measured δ13C values of 12 amino acids, six of which are considered essential – isoleucine (Ile), leucine (Leu), lysine (Lys), phenylalanine (Phe), threonine (Thr) and valine (Val), and six nonessential – alanine (Ala), aspartic acid/asparagine (Asx), glutamic acid/glutamine (Glx), glycine (Gly), hydroxyproline/proline (Pro), and serine (Ser). We use only essential amino acid δ13C data for all statistical analyses in this text.

The reagents used during derivatization add carbon to the side chains of amino acids, and δ13C values measured via GC-C-IRMS are thus a combination of intrinsic amino acid carbon and reagent carbon. However, because amino acid reference materials of known δ13C composition were derivatized and run with each batch of samples, we were able to correct for this carbon addition for each amino acid (see equation 3 from O'Brien et al. (2002)). These corrections are done on a daily, or ‘per run’ basis, which means unknown samples are typically corrected using four to six bracketed injections of the in-house reference material.

Reference: O'Brien, D. M., Fogel, M. L., & Boggs, C. L. (2002). Renewable and nonrenewable resources: amino acid turnover and allocation to reproduction in Lepidoptera. Proceedings of the National Academy of Sciences, 99(7), 4413-4418.