Enantioselective separation of amino acids as biomarkers indicating life in extraterrestrial environments

Traces of prebiotic amino acids, i.e., the building blocks of proteins, are excellent biomarkers that could pro-vide evidence of extinct or extant life in extra-terrestrial environments. In particular, characterization of the enantio-meric excess of amino acids gives relevant information about the biotic or abiotic origin of molecules, because it is generally assumed that life elsewhere could be based on eitherLorDamino acids, but not both. The analytical pro-cedures used in in-situ space missions for chiral discrimina-tion of amino acids must meet severe requirements imposed by flight conditions: short analysis time, low energy con-sumption, robustness, storage for long periods under extreme conditions, high efficiency and sensitivity, automation, and remote-control operation. Such methods are based on gas chromatography, high-pressure liquid chromatography, and capillary electrophoresis, usually coupled with mass spec-trometry; of these, gas chromatography–mass spectrometry (GC–MS) is the only such combination yet used in space missions. Preliminary in-situ sample derivatization is required before GC–MS analysis to convert amino acids into volatile and thermally stable compounds. The silylation reagent most commonly used,N-(tert-butyldimethylsilyl)-N-methyltrifluoroacetamide, isunsuitable for detection of homochirality, and alternative derivatization techniques have been developed that preserve the stereochemical configuration of the original compounds and are com-patible with spaceflight conditions. These include the reagent N,N-dimethylformamide dimethylacetal, which has already been used in the Rosetta mission, a mixture of alkyl chloroformate, ethanol, and pyridine, a mixture of perfluorinated anhydrides and perfluoro alcohols, and hexafluoroacetone, the first gaseous derivatizing agent. In all the space instruments, solvent extraction of or-ganic matter and chemical derivatization have been combined in a single automatic and remote-controlled procedure in a chemical reactor. Liquid-based separation systems have been used in space missions. In particular, microchip capillary electrophoresis, based on microfluidic lab-on-a-chip systems, enables high-performance chemical analysis of amino acids with low mass and volume equipment and lowpower and reagent consumption. Couplingwith laser-induced fluorescence detectors results in ultra-low limits of detection. This critical reviewdescribes applications of the on-board instruments used in the Rosetta mission to comets and in the more recent Mars exploration program, i.e., the Mars Science Laboratory and ExoMars missions.