It has been recently demonstrated, utilizing interspecific introgression lines of tomato, generated from the cross between Solanum lycopersicum and the wild species Solanum pennellii, that the efficiency of photosynthate partitioning exerts a considerable influence on the metabolic composition of tomato fruit pericarp. In order to further evaluate the influence of source-sink interaction, metabolite composition was determined by gas chromatography-mass spectrometry in a different population. For this purpose, we used 23 introgression lines resulting from an interspecific cross between S. lycopersicum and the wild species Solanum chmielewskii under high (unpruned trusses) and low (trusses pruned to one fruit) fruit load conditions. Following this strategy, we were able to contrast the metabolite composition of fruits from plants cultivated at both fruit loads as well as to compare the network behavior of primary metabolism in the introgression line population. The study revealed that while a greater number of metabolic quantitative trait loci were observed under high fruit load (240) than under low fruit load (128) cultivations, the levels of metabolites were more highly correlated under low fruit load cultivation. Finally, an analysis of genotype x fruit load interactions indicated a greater influence of development and cultivation than genotype on fruit composition. Comparison with previously documented transcript profiles from a subset of these lines revealed that changes in metabolite levels did not correlate with changes in the levels of genes associated with their metabolism. These findings are discussed in the context of our current understanding of the genetic and environmental influence on metabolic source-sink interactions in tomato, with particular emphasis given to fruit amino acid content.