Rhododendron anthopogon D. Don., a small compact Himalayan shrub growing in Nepal, is a known medicinal plant used to treat sore throat, colds, blood disorders, bone disease, potato allergies, and vomiting, and to relieve liver disorders, headaches and back pain. The present study investigated the chemical composition and bioactivities of the leaf essential oil from R. anthopogon from Dhankuta, Nepal. The essential oil from leaves was obtained by hydrodistillation and a detailed chemical analysis was conducted by gas chromatography mass spectrometry (GC-MS). The enantiomeric distribution of monoterpenoid components was determined using chiral gas chromatography and represents the first chiral examination of R. anthopogon essential oil. The essential oil was screened for antimicrobial activity using the microbroth dilution test, and for cytotoxic activity against MCF-7, MDA-MB-231, and 5637 using the MTT assay. A total of 70 volatile components were identified from the essential oil. The major components were alpha-pinene (21.5%), delta-cadinene (13.8%), beta-pinene (9.5%), limonene (5.9%), delta-amorphene (4.6%), alpha-muurolene (4.5%), and (E)-caryophyllene (3.2%) with other minor constituents (< 3%). The essential oil showed marginal antibacterial and cytotoxic activities, but no antifungal effects.