A new analytical method for determination of the nitrogen isotopic composition of methionine: Its application to aquatic ecosystems with mixed resources

Compound-specific nitrogen isotope (delta N-15) analysis of amino acids is a powerful tool for estimating the trophic positions (TPs) of animals. The TP of an animal can be represented as a linear function of the isotopic difference between glutamic acid (delta N-15(Glu)) and phenylalanine (delta N-15(phe)). However, the method using delta N-15(Glu) and delta N-15(phe) cannot be applied to animals in mixed food webs where basal resources are derived from both terrestrial and aquatic primary producers, because the mean value of delta N-15(phe) relative to delta N-15(Glu) differs greatly between terrestrial plants (+8.4 parts per thousand) and aquatic algae (-3.4 parts per thousand). To resolve this problem, the delta N-15 of methionine (delta N-15(Met)) is useful. Because the C-N bond of methionine is not cleaved in its initial metabolic step, theoretically there should be little diversity in delta N-15(Met) relative to delta N-15(Glu) among primary producers and a small trophic discrimination factor for methionine in animal metabolism. We developed a dual-column-coupled GC-C-IRMS method to determine delta N-15 met . Data collected from controlled feeding experiments and wild samples demonstrated that the isotopic difference between methionine and phenylalanine in terrestrial food webs (Delta(Met-Phe) = - 16.5 +/- 0.5 parts per thousand) is clearly distinguishable from that in aquatic food webs (Delta(Met-Phe) = - 5.0 +/- 0.5 parts per thousand). This approach allowed us to determine ecologically reasonable TP values for carnivores in a stream food web, which were substantially underestimated with the conventional method. This method has potential utility in assessing TP for animals that rely on varying proportions of both terrestrial- and aquatic-derived resources, with no requirement to characterize delta N-15 in their basal resources.