Plastid sequences of trnL-F are used to estimate the age of biogeographical events in New World Asclepiadoideae. These data suggest that the subfamily arose somewhere in the Old World in the Late Eocene (40 million years ago (Ma)) and that there were at least four independent episodes of dispersal to the New World. The MOG clade, comprising Metastelmatinae, Oxypetalinae, and Gonolobinae, probably colonized South America through transoceanic dispersal from Africa at 32 Ma. The three subtribes emerged at 24 Ma, radiating 15–16 Ma. Cynanchum L. subg. Mellichampia (A. Gray) Woodson (Cynanchinae) only arrived in the New World at 24 Ma and has since diversified gradually. Asclepias L. (Asclepiadinae) probably arrived in the New World through the Bering Strait at 20 Ma, dispersing to South America before the emergence of the Isthmus of Panama. More recently, tropical Marsdenia R. Br. (Marsdenineae) probably arrived in the New World by long-distance dispersal at 16 Ma. Most diversification in New World Asclepiadoideae is estimated to have occurred during the Miocene. The synchrony of events suggests an environmental influence on the diversification of the lineages. Nevertheless, individual rates of diversification point out the importance of intrinsic factors.