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Itchy leaves Laportea decumana (Roxb). Wedd is an indigenous plant in Maluku, Indonesia, and is used traditionally to treat complaints such as fatigue and joint and muscle pains. To provide scientific proof of the analgesic effect of L. decumana ethanolic extract tested in in vivo assays while investigating its bioactive phytochemicals using liquid chromatography tandem high-resolution mass spectrometry (LC-HRMS) profiling and Cox-2 inhibition assay. To investigate the analgesic activity of the ethanolic extract, assays were conducted on male mice Balb/c strain by chemical induction using acetic acid (i.p.) and heat induction (hotplate). Mice were divided into six groups consisting of six mice, i.e., the baseline group; positive control group (paracetamol 80 mg/kg BW); groups treated with extracts in dosages of 100, 200, and 400 mg/kg bodyweight (BW); and negative control group (acetic acid 0.6%, i. p.). The crude extract was partitioned with liquid–liquid fractionation to yield hexane, ethyl acetate, and water fractions. The extract and fraction were assayed for Cox-2 enzyme inhibition, and the chemical profiles were analyzed using untargeted LC-HRMS. The analgesic assays revealed the dose-dependent effect of the extracts, of the effect of treatment with 400 mg/kg BW was not significantly different with that of paracetamol (p < 0.05). The ethyl acetate fraction showed IC 50 of 19.25 μg/mL on Cox-2 inhibition (IC 50 celexocib 18.48 μg/mL). LC-HRMS showed a distinctive profile of the ethyl acetate fraction compared with those in the extract and other fractions. This study presents scientific evidence of the analgesic activity of the L. decumana ethanol extract given orally to experimental animals. Inflammatory inhibition plays a role in the overall analgesic mechanism by Cox-2 inhibition of the extract and all fractions. This finding is also supported by the phytochemical profiles of the extract and fractions, showing the presence of compounds reported elsewhere as anti-inflammatory activity. [Display omitted] • Itchy leaves Laportea decumana (Roxb.) Wedd is a indigenous plant that grows in eastern Indonesia, specifically in Maluku. • The local community usually uses its leaves to treat complaints such as pain, fatigue, and muscle pains. • Studies of the analgesic activity and phytochemical profile of this plant's extract may support its development as herbal medicine for analgesia. [ABSTRACT FROM AUTHOR]