We recently cloned the murine homologue of Cyp26B1, a novel retinoic acid (RA)-metabolizing enzyme and showed that its gene expression pattern is unique from that of Cyp26A1 during early embryogenesis. Here, we complete this comparative expression analysis from embryonic day (E) 12 to postnatal stages. Cyp26B1 expression was found in developing tendons and precartilaginous elements and in perichondrium by E14.5, while Cyp26A1 expression was restricted to extremities of rib and vertebral cartilage. Cyp26A1 and Cyp26B1 were expressed, in the distal epithelium and mesenchyme of the limbs and genital tubercle, respectively. High Cyp26B1 expression was found in craniofacial areas undergoing morphogenetic growth, whereas Cyp26A1 message was restricted to the mouth and dental epithelium. Cyp26A1 alone was expressed in the developing neural retina, while both genes were co-expressed in the retinal pigment epithelium. Cyp26B1 was specifically expressed in the developing hindbrain (pons, cerebellum) and forebrain (striatum, hippocampus), with forebrain expression persisting postnatally. In addition, Cyp26B1 was expressed at specific levels of the differentiating upper and lower thoracic spinal cord, adjacent to the cervical and lumbar regions that express the RA-synthesizing enzyme RALDH-2. In viscera, Cyp26B1 transcripts were detected in the developing lung, kidney, spleen, thymus and testis, whereas Cyp26A1 transcripts were found in the diaphragm and outer stomach mesenchyme. Cyp26B1 was also specifically expressed in dermis surrounding the developing hair follicles. Regulated RA metabolism may therefore be required in many developing systems.