The membrane glycerolipids of four phototrophs that were isolated from an edaphic assemblage were determined by UPLC-MS after cultivation in a laboratory growth chamber. Identification was carried out by 18S and 16S rDNA sequencing. The algal species were (Charophyta), sp. (Chlorophyta), and (Bacillariophyta), and the cyanobacterium was (Cyanobacteria). The glycerolipid profile of sp. was dominated by monogalactosyldiacylglycerol (MGDG) species, with MGDG(18:3/16:4) accounting for 68.6%, whereas MGDG(18:3/16:3) was the most abundant glycerolipid in (50.1%). A ratio of digalactosyldiacylglycerol (DGDG) species to MGDG species (DGDG/MGDG) was shown to be higher in (0.26) than in sp. (0.14). This ratio increased under high light (HL) as compared to low light (LL) in all the organisms, with its highest value being shown in cyanobacterium (0.38-0.58, LL-HL). High contents of eicosapentaenoic acid (EPA, C20:5) and hexadecenoic acid were observed in the glycerolipids of . Similar Fourier transform infrared (FTIR) and Raman spectra were found for and sp. Specific bands at 1629.06 and 1582.78 cm were shown by in the Raman spectra. Conversely, specific bands in the FTIR spectrum were observed for at 1143 and 1744 cm . The results of this study point out differences in the membrane lipid composition between species, which likely reflects their different morphology and evolutionary patterns.