Dietary protein content and digestibility influences discrimination of amino acid nitrogen isotope values in a terrestrial omnivorous mammal

Abstract

Rationale Ecologists increasingly determine the δ15N values of amino acids (AA) in animal tissue; “source” AA typically exhibit minor variation between diet and consumer, while “trophic” AA have increased δ15N values in consumers. Thus, trophic‐source δ15N offsets (i.e., Δ15NT‐S) reflect trophic position in a food web. However, even minor variations in δ15Nsource AA values may influence the magnitude of offset that represents a trophic step, known as the trophic discrimination factor (i.e., TDFT‐S). Diet digestibility and protein content can influence the δ15N values of bulk animal tissue, but the effects of these factors on AA Δ15NT‐S and TDFT‐S in mammals are unknown. Methods We fed captive mice (Mus musculus) either (A) a low‐fat, high‐fiber diet with low, intermediate, or high protein; or (B) a high‐fat, low‐fiber diet with low or intermediate protein. Mouse muscle and dietary protein were analyzed for bulk tissue δ15N using elemental analyzer‐isotope ratio mass spectrometry (EA‐IRMS), and were also hydrolyzed into free AA that were analyzed for δ15N using gas chromatography‐combustion‐IRMS. Results As dietary protein increased, Δ15NConsumer‐Diet slightly declined for bulk muscle tissue in both experiments; increased for AA in the low‐fat, high‐fiber diet (A); and remained the same or decreased for AA in the high‐fat, low‐fiber diet (B). The effects of dietary protein on Δ15NT‐S and on TDFT‐S varied by AA but were consistent between variables. Conclusions Diets were less digestible and included more protein in Experiment A than in Experiment B. As a result, the mice in Experiment A probably oxidized more AA, resulting in greater Δ15NConsumer‐Diet values. However, the similar responses of Δ15NT‐S and of TDFT‐S to diet variation suggest that if diet samples are available, Δ15NT‐S accurately tracks trophic position. If diet samples are not available, the patterns presented here provide a basis to interpret Δ15NT‐S values. The trophic‐source offset of Pro‐Lys did not vary across diets, and therefore may be more reliable for omnivores than other offsets (e.g., Glu‐Phe).

Reevaluating trophic discrimination factors (Δδ13CandΔδ15N) for diet reconstruction

“…We define a single-source system where all food items have the same primary production origin (e.g., omnivore consuming C 3based protein and carbohydrates) and a mixed system where two or more food items differ in their primary production (e.g., omnivore consuming C 3 -based protein and C 4 -based carbohydrates). We hypothesized that, like TDF-δ 15 N (Robbins et al, 2005;Whiteman et al, 2021), TDF-δ 13 C would be primarily related to dietary protein (Farquhar et al, 1989). In marine systems, the δ 13 C values of phytoplankton (marine algae) are largely controlled by growth rates due to differences in temperature, nutrient availability, and levels of dissolved CO 2 (Graham et al, 2010), with δ 13 C values intermediate between C 3 and C 4 plants (À23.0‰ [À31.9 to À12.9], n = 1434) (Graham et al, 2010;McMahon et al, 2013).…”

“…We define a single‐source system where all food items have the same primary production origin (e.g., omnivore consuming C 3 ‐based protein and carbohydrates) and a mixed system where two or more food items differ in their primary production (e.g., omnivore consuming C 3 ‐based protein and C 4 ‐based carbohydrates). We hypothesized that, like TDF‐δ 15 N (Robbins et al, 2005; Whiteman et al, 2021), TDF‐δ 13 C would be primarily related to dietary protein content (Figure 2a–c) (McMahon et al, 2010) but that patterns would differ between a single‐source and mixed system because metabolic routing would lead TDF‐δ 13 C to strongly depend on the δ 13 C value of the dietary protein source relative to that of the energy source (Figure 2d,e) (Hobbie, 2017). We predicted that in a single‐source system, or a mixed system where protein has a higher δ 13 C value than the energy source, preferential routing would cause TDFs to be positive (Figure 2b–d).…”

Reconstructing Hominin Diets with Stable Isotope Analysis of Amino Acids: New Perspectives and Future Directions

“…A feeding trial with mice showed that higher protein led to smaller TDF Glx – Phe , probably because proteins were used as an energy source instead of lipids (Whiteman et al. 2021 ). However, the TDF for proline to leucine spacing (TDF Pro – Lys ) did not change significantly, indicating that these two amino acids may provide a more precise estimate of trophic position when the diet quality is unknown ( figure 3 ).…”

“…Threonine, which becomes 15 N depleted during trophic transfer, has been proposed as a new biomarker for protein consumption (Fuller and Petzke 2017 ), but more studies are needed to understand how δ 15 N threonine is affected by diet quality (Whiteman et al. 2021 ).…”

Extreme bill dimorphism leads to different but overlapping isotopic niches and similar trophic positions in sexes of the charismatic extinct huia

“…Values of δ 15 N bulk have traditionally been used to estimate trophic position (DeNiro and Epstein 1981 ; Minagawa and Wada 1984 ; Peterson and Fry 1987 ; Post 2002 ), whilst δ 13 C bulk data are more useful for determining sources of carbon for an organism (Rounick and Winterbourn 1986 ; Peterson and Fry 1987 ; O’Leary et al 1992 ; Post 2002 ). With the introduction of compound-specific stable isotope analysis (CSIA) of nitrogen in amino acids, it is now possible to more accurately assess an animal’s trophic position (TP) using δ 15 N values of certain amino acids ( δ 15 N AA ) (Chikaraishi et al 2011 , 2014 ; Whiteman et al 2019 , 2021 ). This approach is based on the fractionation between the so-called ‘trophic’ and ‘source’ amino acids in metabolic processes.…”