Type 2 diabetes mellitus is the result of a combination of impaired insulin secretion with reduced insulin sensitivity of target tissues. There are an estimated 150 million affected individuals worldwide of whom, due to a lack of specific symptoms early on in this disorder and inadequate screening diagnostics, a large proportion remain undiagnosed. In this study, NMR based metabolomic analysis in conjunction with multivariate statistics was applied to examine the urinary metabolic changes in two animal models of T2DM as well as unmedicated human sufferers. The db/db mouse and obese Zucker (fa/fa) rat have autosomal recessive defects in the leptin receptor gene, causing type 2 diabetes. ¹H NMR spectra of urine were used in conjunction with uni- and multi-variate statistics to identify disease related metabolic changes in these two animal models and human sufferers. This study demonstrates metabolic similarities between the three species examined, including metabolic responses associated with general systemic stress, changes in the TCA cycle, perturbations in nucleotide metabolism and in methylamine metabolism. All three species demonstrated profound changes in nucleotide metabolism, including that of N-methylnicotinamide and N-methyl-2-pyridone-5-carboxamide, which may provide unique biomarkers for following T2DM progression.