© K. Douka,  T. Higham, E. Mastora

SINGLE AMINO ACID DATING

Archaeological bones are usually dated by radiocarbon measurement of extracted collagen. In Oxford, we have used ultrafilters to improve the recovery and quality of collagen. Sometimes, however, ultrafiltration is not good enough to completely decontaminate bone prior to dating. In some cases a combination of low collagen content, contamination from the burial environment or museum conservation work, such as addition of glues, preservatives and fumigants to ‘protect’ archaeological materials, result in highly contaminated samples that contain high molecular weight contaminants which will not be removable by ultrafiltration methods.

 

The new ORAU Jasco HPLC and SFC system, currently being used in the lab for amino acid separation and testing. 

Over the last decade in Oxford we have worked on developing methods to further improve the routine dating of archaeological bone by dating single amino acids using HPLC methods. It is possible, however, that single amino acids found in bone may have multiple sources. Some may be derived from bacteria or other organisms in the bone’s depositional environment for example having found their way into bone by leaching under conditions of poor preservation. Ho and co-workers first suggested isolating and dating hydroxyproline (HYP) specifically to circumvent this potential problem (Ho et al., 1969, Science). Other groups have attempted to reliably date HYP from bone, but their efforts have been hampered by methodological problems in determining the extent of column bleed and correcting for background.

 

HYP is found in mammalian collagen and constitutes around 10% of collagen carbon. It acts essentially as a bone specific biomarker because of the rarity with which it is found in non-mammalian material. Extraction and dating of HYP essentially provides us with a non-contaminated biomarker, and therefore a ‘gold-standard’ AMS date. In PalaeoChron we are therefore keen to further develop and apply the methods we currently use in the lab; a preparative mixed-mode chromatographic protocol designed for separation and dating of all the amino acids from bone collagen. An example of the type of separation currently possible is shown below, you can see the excellent separation of amino acids on one of our columns. HYP is one of the early eluting amino acids on the left hand side of the diagram. 

 

In addition to our preparative HPLC system suitable for mixed-mode chromatography, the research laboratory recently acquired a Supercritical Fluid Chromatography system. We have built a new Chromotography lab specifically for these instruments. Dr Thibaut Deviese (bottom right) is leading the amino acid development work. Dan Comeskey also works a great deal on the HPLC and method development. Several MSc and DPhil students have worked on single amino acid dating, whether developmentally or in an applied way. Currently, Eileen Jacob, Lorena Becerra Valdivia and Rachel Hopkins are involved. 

We have a methods paper about to be published. It can be seen in pre-proof stage online in the link below:

  • Devièse, T., Comeskey, D., McCullagh, J., Bronk Ramsey, C. and Higham, T.F.G. 2017. New protocol for compound specific radiocarbon analysis of archaeological bones. Rapid Communications in Mass Spectrometry (online in press) Link.

Copyright: James King-Holmes