Characterisation of cell membrane lipids of the thermophilic archaeon Sulfolobus acidocaldarius using high-performance thin layer chromatography and mass spectrometry

Abstract

Cell membrane lipids of archaea dier signicantly in their chemical structure from typical eubacterial cell membrane lipids, instead of glycerol ester-bond lipids archaea exclusively form glycerol ether-bond lipids. The lipid part itself is formed by connecting isoprene building blocks up to a C20-back bone (i.e. phytanol) exhibiting multiple methyl branchings. The membranes mainly consist of diphythanyl glycerol- and dimeric bis-diphythanyl glycerol-based lipids diering in their polar head groups (e. g. phosphoinositol, dihexoside etc.). Structural variation in the bis-diphythanyl glycerol-based lipid portion strongly depends on the growth conditions of Sulfolobus acidocaldarius as e.g. cyclopentane-ring formation. Archaeal lipids were analyzed by high-performance thin layer chromatography (normal phase as well as reverse-phase). For practical reasons, normal phase HPTLC was used. As lipid staining agent primuline was used being fully compatible with subsequent mass spectrometric analysis. This analytical strategy was applied to archaeal lipid extracts dissolved in dierent extraction solvents (methanol/chloroform, isopropanol/dimethylsulfoxide). After TLC separation the separated lipids were scraped from the TLC plate, extracted from the stationary phase and measured by mass spectrometry. Final mass spectrometric detection was performed by matrix-assisted laser desorption/ionization in combination with either a reectron time-of-ight or a hybrid quadrupole/ reflectron time-of-ight. After optimizing measurement conditions for the samples like adapting the matrix system, laser power and rastering over the sample during the desorption/ionization process the dierent lipids were analyzed. During this thesis an algorithm was developed that allows to deduce the number of cyclic structures from isotopic structures detected in the dierent samples.Data on the detailed composition of archaeal lipids will serve as valuable information for subsequent liposome experiments for drug incorporation.