Although to date the phylogenetic position of the provisional order Eulipotyphla has been assessed by various molecular markers, it has not been conclusively clarified due to low statistical supporting values and inconsistent results. To clarify the phylogenetic position of Eulipotyphla, we cloned cDNAs for pepsinogens A and C from five mammalian species belonging to four different orders and determined their nucleotide sequences. Molecular phylogenetic analysis based on the 1st and 2nd codon positions of the protein-coding region of cDNA sequences strongly supported the close relationship between Eulipotyphla and Chiroptera. Carnivora was found to be a sister group to these two orders. The monophyly of the order Rodentia and that of the cohort Glires (Rodentia and Lagomorpha) was also shown by the present phylogenetic trees of pepsinogens.