Tracking the synthesis and metabolism of amino acids in mouse liver and muscle tissue using nitrogen isotopes (δ15N)
Stable isotope analysis has long been a staple method for estimating trophic position and tracing nutrients in individuals and ecosystems. More recently the use of compound specific stable isotope analysis of nitrogen in amino acids (AA-CSIA) has emerged as a promising technique that resolves some of the ambiguities in analyses of bulk tissue. Compound specific isotope analyses rely upon groupings of amino acids as either “source”, i.e., no sign of isotopic enrichment from diet to consumer, and “trophic”, i.e., considerable isotopic enrichment. However, these classifications overlook the metabolic and catabolic processes that drive amino acid δ15N values and fractionation patterns. We completed controlled feeding experiments on mice (Mus musculus), varying the proportion of protein and carbohydrates in the diet, in order to begin addressing the isotopic routing and synthesis of nitrogen from dietary components.
We measured δ15N values in mouse liver that were consistently lower than muscle in both trophic and source amino acids in all treatments; these differences were substantially larger in low protein diets (average trophic: 6.9‰) versus high protein diets (average trophic: 3.6‰). Additionally, in low protein treatments, amino acids in liver were depleted in 15N by 2-5‰ compared to the diet. In combination with muscle tissue 15N enrichment, these values cannot be explained solely through routing of amino acids or reduced nitrogen metabolism. Our results confirm that animals consuming higher protein diets preferentially metabolize non-essential amino acids, regardless of trophic or source designations. However, when a diet is low in protein, our data indicate an alternate biosynthesis of both trophic and source essential amino acids from a pool of nitrogen depleted in 15N. Our study represents a major step toward understanding nitrogen isotope dynamics in amino acids, without which interpretation of AA-CSIA data is incomplete. Furthermore, data from this and future studies may be applied to modeling efforts to examine de novo amino acid synthesis contributions by gut microbiota.