Two distinct isozymes (UMPH-1 and UMPH-2) which hydrolyse pyrimidine 5'-nucleotides have been identified in mouse, rat and Chinese hamster tissues. One of these, UMPH-1 appears to be specific for pyrimidine 5'-nucleotides, whereas the other, UMPH-2 has a broader substrate specificity. UMPH-1 and UMPH-2 show differences in their expression in different tissues. The evidence suggests that UMPH-1 and UMPH-2 are coded by distinct structural gene loci, Umph-1 and Umph-2. These isozymes have not been identified in man, and it is suggested that they may have overlapping electrophoretic mobility.