Amino acid‐specific δ¹⁵N trophic enrichment factors in fish fed with formulated diets varying in protein quantity and quality

Abstract

Compound‐specific isotope analysis (CSIA) of amino acids (AAs) in consumer tissues is a developing technique with wide‐ranging applications for identifying nitrogen (N) sources and estimating animal trophic level. Controlled experiments are essential for determining which dietary conditions influence variability in N stable isotopes (δ15N) trophic enrichment factors in bulk tissue (TEFbulk) and AAs (TEFAA). To date, however, studies have not independently evaluated the effect of protein quantity and quality (digestibility) on TEFs, complicating the application of AA‐δ15N values for estimating trophic levels. We conducted a 98‐d feeding experiment using five formulated isoenergetic feeds prepared with a high‐quality protein source to evaluate the effect of protein quantity and quality on TEFs of liver and muscle tissues of juvenile Pacific yellowtail (Seriola lalandi), a carnivorous fish species. We decreased protein digestibility using well‐established protocols that do not change AA profiles. Growth rates were higher in diets with higher protein content, and isotopic equilibrium was reached for both fish tissues and all treatments. Protein quantity and quality influenced isotope discrimination depending on tissue type and AA. In liver tissue, bulk TEFs showed a limited but significant relationship with protein quality, but did not differ with protein quantity or quality in muscle. None of the pre‐established source AAs (Lys, Met, Phe, and Gly) TEFs varied significantly with protein quantity or quality in liver tissue. However, in muscle tissue, TEFPhe increased significantly with protein content and decreased in response to reduced digestibility, indicating it may not serve as proxy for baseline isotopic values used to calculate trophic level. Among trophic AAs, TEFLeu decreased significantly with increasing protein quantity in liver tissue, while both Leu and Ile TEFs decreased with lower protein digestibility in muscle tissue. Our results indicate that CSIA‐AA in liver tissue provides more robust source and trophic AA‐δ15N values than in muscle.

A Guide to Using Compound-Specific Stable Isotope Analysis to Study the Fates of Molecules in Organisms and Ecosystems

“…In addition, consumers that excrete nitrogen as ammonia rather than as urea or uric acid have higher Δ 15 NC-D values [74], possibly because the relatively greater toxicity of ammonia requires this molecule to be excreted rapidly, which can cause substantial fractionation [37,75]. Values of Δ 15 NC-D can also vary among tissues [76,77] with the highest values occurring in tissues with very active metabolic processing of AA, such as liver [73]. Unique physiological processes can also affect Δ 15 NC-D. A recent study of captive leopard sharks (Triakis semifasciata) found surprisingly low Δ 15 NC-D values for trophic AA [72], likely related to the ability of elasmobranchs to retain and recycle urea nitrogen into newly-synthesized AA [31,76].…”

Compound‐specific isotope analysis of amino acids as a new tool to uncover trophic chains in soil food webs

“…Although assumed to change minimally between diet and consumer, source AAs such as Phe may also vary in trophic enrichment (Nuche‐Pascual et al. ) and may even be depleted in 15 N compared to the diet (Steffan et al. ).…”

“…A recent review by McMahon and McCarthy (2016) showed that the trophic fractionation of Glu and Phe is not as consistent as initially assumed. Fractionation of Glu may vary depending on diet quality (Hoen et al 2014, Blanke et al 2017, Nuche-Pascual et al 2018 and mode of N excretion (Germain et al 2013). Although assumed to change minimally between diet and consumer, source AAs such as Phe may also vary in trophic enrichment (Nuche-Pascual et al 2018) and may even be depleted in 15 N compared to the diet (Steffan et al 2013).…”

“…Fractionation of Glu may vary depending on diet quality (Hoen et al 2014, Blanke et al 2017, Nuche-Pascual et al 2018 and mode of N excretion (Germain et al 2013). Although assumed to change minimally between diet and consumer, source AAs such as Phe may also vary in trophic enrichment (Nuche-Pascual et al 2018) and may even be depleted in 15 N compared to the diet (Steffan et al 2013). To account for variable TDF Glu-Phe among different consumers, the use of multi-TDF equations has been shown to yield more exact estimates of trophic position in some studies (Germain et al 2013, McMahon et al 2015b, Wu et al 2018.…”

Errors associated with compound specific δ15N analysis of amino acids in preserved fish samples purified by high pressure liquid chromatography

“…This approach assumes that analytical and methodological errors are similar for CSIA-AA and bulk SIA. However, Elemental Analyzer -Isotope Ratio Mass Spectrometry (EA-IRMS) used in bulk SIA typically operates with an analytical precision (0.1 -0.2 ‰) that is substantially higher than what is reported in CSIA-AA studies (0.4 -1.0 ‰; Bradley et al 2016;Broek and McCarthy 2014;Broek et al 2013;Chikaraishi et al 2015;Hetherington et al 2017;Nuche-Pascual et al 2018;Ogawa et al 2013;Ruiz-Cooley et al 2017;Vane et al 2018;Vokhshoori et al 2019). This lower precision is the result of a lengthy and time-consuming analytical process inherent to the Gas Chromatography-Combustion-IRMS (GC-C-IRMS) used for almost all CSIA-AA work.…”