To identify novel genes associated with iron metabolism, we performed gene chip studies in 2 models of iron-deficiency: iron-deprived rats and rats deficient in the principal intestinal iron transporter, divalent metal transporter 1 (i.e. Belgrade rats). Affymetrix rat genome gene chips were utilized (RAE230) with cRNA samples derived from duodenum and jejunum of experimental and control animals. Computational analysis and statistical data reduction identified 29 candidate genes, which were induced in both models of iron-deficiency. Gene ontology analysis showed enrichment for genes related to lipid homeostasis, and one gene related to this physiological process, a leukocyte-type, arachidonate 12-lipoxygenase (Alox15), was selected for further examination. Real-time PCR studies demonstrated strong induction of Alox15 throughout the small and large intestine, and in the liver of iron-deficient rats. Polyclonal antibodies were developed and utilized to demonstrate that proteins levels are significantly increased in the intestinal epithelium of iron-deprived rats. HPLC analysis revealed altered intestinal lipid metabolism indicative of Alox15 activity, which resulted in the production of biologically active lipids (12-HETE, 13-HODE and 13-HOTE). The overall effect is a perturbation of intestinal lipid homeostasis, which results in the production of lipids essentially absent in the intestine of control rats. We have thus provided mechanistic insight into the alteration in lipid metabolism that occurs during iron-deficiency, in that induction of Alox15 mRNA expression may be the primary event. The resulting lipid mediators may be related to documented alterations in villus structure and cell proliferation rates in iron-deficiency, or to structural alterations in membrane lipid composition.