Cyclotides are small disulfide-rich plant peptides. Their unique structural features, intrinsic bioactivities, and enormous sequence diversity make them interesting molecules for pharmaceutical applications. So far the amino acid sequences of nearly 400 individual cyclotides have been reported and they have been discovered in plant species of the Violaceae, Cucurbitaceae, Fabaceae, Solanaceae, Poaceae, and Rubiaceae families. Rubiaceae is the fourth largest angiosperm family and comprises about 650 genera and 13,000 species. The family includes coffee, which is a major export commodity and it was the top agricultural export for 12 countries in 2004, as well as the world's seventh largest legal agricultural export by value in 2005. Additionally, the family comprises timber species such as bilinga, many ornamental plants, and important plants of medicinal purpose such as quinine and ipecac. In part due to the wide geographical distribution, plants of the coffee family are estimated to constitute about 8% of the world's flora. In this book chapter, we will (1) summarize our efforts to identify and characterize novel cyclotide-expressing plants within the Rubiaceae family, (2) analyze the sequence diversity of cyclotides from Rubiaceae, (3) discuss their evolution in flowering plants based on the distribution of cyclotide-bearing Rubiaceae species, and (4) provide a brief overview of bioactive properties of cyclotides that have been isolated from Rubiaceae plant species. Based on studies of cyclotides in Rubiaceae their number has been predicted to be greater than 50,000 and we believe cyclotides are one of the largest protein families within the plant kingdom.