The tribe Eupatorieae has a chromosome base number that ranges from 4 to 25. A molecular phylogenetic analysis using cpDNA restriction site mutations was performed. Fifteen species representing 13 subtribes of the tribe Eupatorieae were examined, together with three species from the tribe Heliintheae as outgroups. A total of 103 restriction site mutations were detected, and 31 of these were phylogenetically informative. Parsimony analysis produced a single most parsimonious tree with 117 steps. This tree suggested that two clades diverged early in the evolution of the tribe Eupatorieae. One clade includes Neomirandea (x=17 and 25), Agerafina (x=17) and Sclerolepis (x=15) with the higher chromosome base numbers, and the other includes Mikana (x=17) and the remaining genera with lower chromosome base numbers (x=lO-ll). However, the monophyly of the former clade is supported with a low bootstrap value. In the latter clade, Mikana (x= 17) diverged first, then Sfevia (x=ll), and finally eight genera with x=10 diverged in succession. This result supports the hypothesis that the genera in the tribe Eupatorieae with x=10 evolved from an ancestor with a higher base number, and the tribe is of polyploid origin.