Abstract. Amalini YR, Muzakhar K, Su’udi M, Setyati D, Dwinianti EF, Ulum FB. 2025. Morphological, molecular, and phytochemical characteristics of wild edible fern (Diplazium esculentum) from Jember, East Java, Indonesia. Biodiversitas 26: 2794-2805. Diplazium esculentum, an edible pteridophyte with medicinal potential, exhibits morphological variation that risks improper medicinal use. This study aimed to comprehensively characterize D. esculentum from Jember, Indonesia, through an integrative approach, including morphological analysis, DNA barcoding, phytochemical profiling, and antioxidant activity assessment. Fifteen specimens, locally referred to as pakis menir, were collected from a single site within a rubber-mixed coffee plantation in Panti, Jember District, East Java, Indonesia. The examination of morphological traits, molecular identification using the rbcL marker, analysis of phytochemical profiles through GC-MS, prediction of biological activities using PASS online, and evaluation of antioxidant activity using the DPPH assay all pointed to the potential of D. esculentum. Morphological analysis confirmed D. esculentum's diagnostic traits, while rbcL DNA barcoding validated a 100% sequence match with D. esculentum (OL536867.1). GC-MS analysis of young fronds identified 25 bioactive metabolites, including 11 phenolics, 9 terpenoids, and 5 alkaloids, with 96% exhibiting high biological activity (Pa>0.7) according to PASS predictions. The dominant compound, 3,6-dimethylquinoline (12.38%), exhibited significant anticancer potential, while the frond extract showed strong antioxidant activity (IC50: 91.25±7.60 ppm), reflecting its phenolic richness. These results highlight the potential of D. esculentum's phytochemical wealth and pharmacological promise, supporting further exploration in pharmacognosy, nutraceuticals, and bioactive compounds.