Ecological complexity may improve ecosystem function, stability and adaptability to natural and anthropogenic disturbances. Intraspecific trophic variation can represent a significant component of total community variation and can influence food web structure and function. Thus, understanding how trophic niches are partitioned between intraspecific and interspecific processes could improve our understanding of food web dynamics. We examined gut contents, fatty acids and stable isotope ratios in round goby (Neogobius melanostomus) and yellow perch (Perca flavescens) across six sites in Lake Michigan, USA, to determine patterns in intra- and interspecific trophic composition (i.e., mean gut or fatty acid composition) and diversity (i.e., the diversity of gut items or fatty acids). We also examined relationships between fatty acid diversity and gut content characteristics to understand potential mechanisms shaping individual trophic phenotypes. There was significant variation in both trophic composition and diversity among sites, and individual and spatial variation was as important to total trophic variation as species identity. Round goby that consumed dreissenid mussels had more diverse fatty acid profiles than those that consumed other benthic invertebrates, whereas yellow perch fatty acid diversity was not related to gut content composition. Our results confirm that intraspecific variation in resource use can be as important to trophic dynamics as interspecific variation, and that spatial variation in lower level food web processes or habitat may strongly structure local food web dynamics. Individual-level examination of trophic diversity, in concert with trophic composition, could provide additional information about the resilience, function and adaptability of local food webs.