Leaves of Melaleuca leucadendra contain the novel flavonol glycoside, myricetin 3-O-β-4C1-galactopyranuronoid. In addition, known fifteen phenolics were identified. All isolates are characterized for the first time from this plant. Structures were established by conventional methods and confirmed by spectral methods of analysis, including one and two-dimensional nuclear magnetic resonance spectroscopy (1D and 2D-NMR) and high resolution electro-spray ionization mass spectrometry (HRESIMS), as well. Assessment of some immunological and biological efficacy, of the extract in combination with a parallel cytotoxicity evaluation, using the method of cellular reduction of 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) technique was carried out. Besides, evaluation of the antioxidant effectiveness, using the free radical 2,2-diphenyl-1-picrylhydrazyl (DPPH) and the oxygen radical absorption capacity (ORAC) methods was performed. In addition, the cytotoxicity against liver (Huh-7), breast (MCF-7) and prostate (PC-3) cancers using the neutral red assay (NRU) technique for the extract and the new flavonol glycoside also, was assessed.