This paper studies the phytochemical analysis of the acetone extracts of the Ramalina fraxinea and Ramalina fastigiata lichens and the antioxidant, antimicrobial and antitumour activities of these species and their constituents. The phytochemical analysis of two Ramalina species was evaluated using HPLC-UV test. The depsides (evernic acid, obtusatic acid, sekikaic acid and atranorin), depsidones (protocetraric acid) and dibenzofurane (usnic acid) were identified from these lichens. Antioxidant activity was evaluated by DPPH assay, reducing power assay and by measuring the amounts of total phenolics in extracts. Antimicrobial activity was tested towards five bacterial and 10 fungal species, using broth microdilution method to determine the minimum inhibitory concentration. Cytotoxic activity was tested using MTT method on the human epithelial carcinoma (Hela), human lung carcinoma (A549) and human colon carcinoma (LS174) cells. As a result of the study, tested samples showed strong free radical scavenging activity with I50 values within the range of 285.45-423.51 μg/mL. Absorbance for reducing power was found to be from 0.0043 to 0.1747. The total amount of phenol concentrations in extracts of Ramalina fraxinea and Ramalina fastigiata was 32.63 and 33.49 μg PE/mg, respectively. Methyl evernate showed the strongest antimicrobial properties with the least measured MIC value being 0.125 mg/mL. In addition, all samples exhibited strong anticancer activities against tested cells (I50 values were between 24.63 and 161.37 μg/mL). These results indicate that lichen appears to be a possible natural biopharmaceutical.