Cyclotides are small disulfide-rich peptides that are characterized by a head-to-tail cyclized peptide backbone and a knotted arrangement of three conserved disulfide bonds. Their primary function in plants is thought to be as defense agents, based on their potent insecticidal activity, but they also have a range of other biological activities, including insecticidal, nematocidal, molluscicidal, antimicrobial, anti-HIV, and antitumor activities. Until now, these molecules have been identified in five major families of angiosperms, namely the Rubiaceae, Violaceae, Poaceae, Cucurbitaceae, Solanaceae and Fabaceae, despite this, the work involving Brazilian plants are scarce. In order to address these deficiencies, our group has been dedicated to study this class of interesting natural products in Brazilian plants. Hence, this work describes the isolation and characterization of two cyclotides from medicinal plant Anchietea pyrifolia (Mart.) G. Don collected on Atlantic Forest.