This study presents the first molecular evidence of Macrovipera razii from central Zagros, more than 300 km north-west of its prior records in southern Iran. Molecular analyses based on mitochondrial cytochrome b sequences identified the individuals from central Zagros as a lineage of M. razii. Specimens from the new localities are separated by a genetic distance of 1.46% from the known populations of M. razii. The results extend the known distribution range of M. razii as an endemic species of Iran.