Exploring molecular biomarkers in lava tube speleothems offers insights into environmental dynamics, biogeochemical processes, and subsurface life, with implications for astrobiology, particularly in Martian analog environments. Despite extensive biomarker research in marine and terrestrial settings, lava tube studies are limited. This study employed molecular, isotope and mineralogical characterization to analyze sulfate speleothems from six lava tubes of Lanzarote (Canary Islands), considered analog for the Moon and Mars. The combination of mass spectrometry, thermogravimetry, and mineralogical techniques, revealed geological processes, biomarkers and their biological sources. The identified minerals were primarily calcium and sodium sulfates. Sulfur isotope analyses indicated volcanic and oceanic origins, while carbon isotope composition and pyrolysis analyses suggested influences from vegetation and microbial activity. Lipidic profiles highlighted branched and n-alkanes, as well as palmitic and stearic acid methyl esters, suggesting microbial origins. These findings contribute to understanding geological and environmental dynamics in lava tubes and recognizing potential biosignatures on Earth and other planets.