We have determined the time course of [U-¹³C]-glucose utilization and transformations in SCID mice via bolus injection of the tracer in the tail vein. Incorporation of ¹³C into metabolites extracted from mouse blood plasma and several tissues (lung, heart, brain, liver, kidney, and skeletal muscle) were profiled by NMR and GC–MS, which helped ascertain optimal sampling times for different target tissues. We found that the time for overall optimal ¹³C incorporation into tissue was 15–20 min but with substantial differences in ¹³C labeling patterns of various organs that reflected their specific metabolism. Using this stable isotope resolved metabolomics (SIRM) approach, we have compared the ¹³C metabolite profile of the lungs in the same mouse with or without an orthotopic lung tumor xenograft established from human PC14PE6 lung adenocarcinoma cells. The ¹³C metabolite profile shows considerable differences in [U-¹³C]-glucose transformations between the two lung tissues, demonstrating the feasibility of applying SIRM to investigate metabolic networks of human cancer xenograft in the mouse model.