Phylogenetic relationships within Periplocoideae (Apocynaceae s.l.) were investigated using sequence data from nuclear ribosomal DNA and two plastid regions: trnD-trnT and trnT-trnF. We included 43 ingroup taxa, representing all but three of 32 currently recognized genera. In the ITS and total evidence analyses, Phyllanthera grayi (P. I. Forst.) Venter is supported as sister to the rest of the subfamily. We identified several well-supported clades, and three of six genera represented here by more than one species (Cryptolepis R. Br., Pentopetia Decne., and Periploca L.) are supported as monophyletic. Pollinial species appear in four distinct, well-defined clades, suggesting either three or four independent origins of pollinia within the subfamily. It is most parsimonious, therefore, to view pollinia in Periplocoideae as non-homologous with those of Secamonoideae and Asclepiadoideae, a hypothesis that is supported by their structural differences.