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Ezine

  • Published: Oct 5, 2009
  • Author: Steve Down
  • Channels: Gas Chromatography
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Life on Earth was influenced by organic molecules from outer space. There is a growing body of thought to support this theory, based on the discovery of amino acids in extraterrestrial material. They have been found in meteorites which landed on Earth and the essential amino acid glycine was discovered in particles shed by the comet Wild 2 in its recent flypast.

Their role in the origins of life is supported by the fact that there tends to be more of the L-amino acids in meteorites than the D-enantiomers, up to 18% more in the case of isovaline in the much-studied Murchison meteorite. It is the L-enantiomers that dominate living systems, building up peptides and proteins.

The presence of amino acids in other parts of space would support their arrival on Earth. One region of interest is Mars because it once held water and organic compounds and may even have supported life at one stage. Space exploration probes have been kitted out with on-board instrumentation for a range of experiments and gas chromatographs have proven sufficiently sturdy and reliable for space flight. This fits in nicely with the amino acid hunt since they can be analysed by GC.

The main obstacle is that the amino acids need to be derivatised to allow them to pass through the GC column but that can be accomplished in space as evidenced by several missions, including the Rosetta and Sample Analysis at Mars (SAM) missions. These two programs managed the automated, remote control sampling, derivatisation and analysis of samples but there was no attempt to separate the L- and D-amino acids from each other.

This goal has been brought closer by two Italian researchers who have compared two derivatisation procedures suitable for the enantiomeric separation of amino acids in space exploration studies. Maria Chiara Pietrogrande and Giulia Basaglia from the University of Ferrara performed the resolution of the derivatised amino acids under energy saving conditions (low column temperature and short analysis time) to mirror those in space.

The amino acid derivatives of 20 proteinogenic amino acids from both reactions were analysed by GC/MS using a chiral column that has been proven to function in space, as part of the Rosetta mission. A temperature program ranging from 60-180°C was chosen to effect separation, providing an acceptable compromise between short retention times, amino acid resolution, and column temperature. A single quadrupole mass spectrometer with an electron ionisation source was used for detection.

In the first instance, the amino acids were derivatised with methyl chloroformate in heptafluoro-1-butanol (HFB) for 1 minute, before extraction with chloroform containing sodium chloride. A total of 14 enantiomeric pairs were resolved under constant elution order with the D-isomer eluting before the L-isomer.

Six pairs (Ala, Val, Ile, Leu, Met, Glu) were well resolved and 8 were less well-resolved because they had higher molecular masses (Thr, Phe, Lys, Tyr, Trp) or had the bis-ester (Asp) or bis-acyl derivatives (Gln, Ser) as the more stable derivative.

For the alternative reaction, the amino acids were derivatised with a perfluoroalcohol-perfluoro anhydride combination for one hour before removing the reagents with nitrogen and extracting with ethyl acetate. The three reagent pairs were trifluoroacetic anhydride (TFAA)-2,2,2-trifluoroethanol, TFAA-HFB and heptafluorobutyric anhydride-HFB. They gave similar retention time patterns and chiral separation behaviour.

Once again, 14 enantiomeric pairs were separated, with 8 pairs (Ala, Val, Ile, Leu, Met, Glu, Phe, Tyr) well resolved and 6 (Pro, Thr, Asp, Lys, Gln, Trp) poorly separated.

For amino acid quantitation, both methods displayed good linearity and produced detection limits of about 0.5 nmol. These are at the right level for in situ analysis of extraterrestrial environments, since concentrations in Earth-landed meteorites are at the sub-nmol level.

The complex GC/MS data were processed using an autocovariance function previously developed by the research group to extract information on the enantiomeric pairs. This automated procedure is suited to the high-throughput analysis of data derived from space studies, reducing the time and labour needed to allow rapid delivery of the results.

Both derivatisation methods involve one-step reactions and produce similar results, making them both suitable candidates for future missions. The next challenge, according to the researchers, is to fully automate the derivatisation for remote control conditions, so that extraction from the local material and derivatisation can be accomplished in one reactor coupled to the GC/MS system.


The views represented in this article are solely those of the author and do not necessarily represent those of John Wiley and Sons, Ltd.

Mars

 Mars: courtesy NASA

 

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