Enantioselectivity of pinene against Leishmania amazonensis

Pinene is a major monoterpene described in various plant essential oils. It has two isomers, α- and β-pinene, that differ in the position of their double bond. Each isomer has two enantiomers, which have diverse pharmacological properties. Human tegumentary leishmaniasis (HTL) is a parasitic disease that significantly affects the morbidity, disability, and social stigmatization of poor and marginalized populations. This study aimed to investigate the activity of pinene enantiomers against the HTL causative agent, Leishmania amazonensis . The parasite viability was determined using the resazurin assay and transmission electron microscopy. The most active enantiomers against promastigotes and amastigotes were (+)-α-pinene and (+)-β-pinene with IC 50 ranging from 14 to 110 µg/mL. Furthermore, the positive enantiomers exhibited moderate cytotoxicity against the RAW 264.7 cell line and sheep erythrocytes and desirable selectivity indexes exceeding 10. In addition, in silico toxicological analysis revealed a pinene enantiomer profile similar to miltefosine. Parasites treated with (+)-α-pinene exhibited significant ultrastructural alterations suggestive of apoptosis, including mitochondrial swelling, myelin-like figures, and autophagosome-like structures. Additionally, (+)-α-pinene enhanced autophagic activity and increased ROS production in treated parasites, as assessed through monodansylcadaverine and H 2 DCFDA labeling, respectively. Our results demonstrated that (+)-α-pinene and (+)-β-pinene are the primary enantiomers responsible for the antileishmanial activity. These results provide an important point of view for the therapeutic use of enantiomers.