Anemia is a widespread hematological disorder in vertebrates. In channel catfish (Ictalurus punctatus), anemia is a persistent problem in culture environments, however, its causes and physiological impacts are not well understood. To better understand the symptoms and characterize associated biomarkers of anemia, 1-D 1H and 2-D 1H J-resolved NMR were used to analyze metabolite changes in healthy and anemic channel catfish kidney and liver tissue. Additionally, standard physiological analytical techniques were used to analyze blood plasma. NMR analyses revealed energy sources such as glucose were depleted and many metabolites associated with anaerobic metabolism or alternative energy pathways such as lactate, creatine, alanine, acetate and myo-inositol had changed. Energy demanding processes were reduced, such as muscle function, as indicated by reductions in taurine and inosine, and protein synthesis. Stress and oxidative stress related metabolites changed, with increases in valine, leucine and isoleucine and decreases in glutathione. Inhibitory neurotransmitters such as 4-aminobutyric acid (GABA) increased, and excitatory neurotransmitters such as glutamine and glutamate generally decreased, although there were tissue-specific differences. Immune function also decreased in anemic fish. Blood analyses revealed decreased respiratory gas transport capabilities through reductions in erythrocytes and hemoglobin-markers. Taken together, these tissues and analytical techniques produced complementary results, and metabolite expression in anemic channel catfish was similar to that observed in other vertebrates exposed to anoxia, severe hypoxia or experiencing oxidative stress from deficiencies in iron homeostasis.