M. Fresnais1,2, P. Richardin2, A.,Gimatb2, M.,Sepúlveda3, E. Leize-Wagner1, and A. Charrié1
- Laboratoire de spectrométrie de masse des interactions et des systèmes (LSMIS), 1 rue Blaise Pascal, 67008 Strasbourg, France
- Centre de recherche et de restauration des musées de France (C2RMF), Palais du Louvre, Porte des Lions, 14 quai François Mitterrand, 75001 Paris, France
- Laboratorio de Análisis e Investigación arqueométrico, Departemento de Antropología, Universidad de Tarapacá, 18 de sept. 2222, Casilla 6D, Arica, Chile
Two pre-Hispanic mummies from the Andean coast, belonging to a corpus of 16 mummies from the San Miguel de Azapa (Arica, Chile), were radiocarbon dated and analyzed in order to replace them in their historical context and to study the conservation state of the hair fibers and the heavy metal presence. The radiocarbon dating placed both mummies in the Formative period (1700 years BC to 500 years AD). Global and elemental analyses were performed using scanning electron microscopy coupled with energy dispersive X-ray spectroscopy and using X-ray fluorescence spectroscopy. These combined techniques enabled to prove the good global conservation state of the mummies’ hair and to detect iron, lead, bromide and also arsenic in some cases, in significant amounts inside the hair fibers. Fourier transformed infra-red spectroscopy seemed to prove the good conservation state of the hair surface at a structural level that is why the conservation of hair proteins at a molecular level will be investigated by a proteomics approach in future work.
CovalX Technology Used (Click each option to learn more)
Samples of modern hair proteins and ancient hair proteins were washed and degreased before being submerged in 5 mL of extraction solution (7 M urea, 2 M thiourea, 50 mM DTT, 50 mM Tris-HCl (pH 7.5), 0.1% Triton X-100). Next, the hair samples were incubated for 18 hours at 37 °C without agitation before being decanted. Soluble and insoluble materials were separated from one another and the protein extract (soluble sample) was alkylated with 1 M iodoacetamide (IAA) and 3 M Tris-HCl in the dark for 10 minutes at room temperature. From this alkylation, the extract was purified using dialysis cartridges (3.5 kDa, 3-12 mL) against water. The water was changed twice, once after 2 hours and then again over 2 days. The extract was freeze dried overnight before being stored at -20 °C.
The samples were resuspended in extraction buffer (25 or 50 mM Tris-HCl) and then dry droplets were placed on a stainless steel target with a saturated solution of sinapic acid in ACN/H2O 0.1% TFA 1:1 (v/v). To prevent saturation of the detector by matrix ions, gated suppression was applied. The samples were analyzed using a mass spectrometer that had been modified with a CovalX high mass detection system. From the preliminary data analysis, it was found that the two archaeological hair samples had a similar signature that differed significantly from modern hair. The ancient hair had no defined protein profile and no intact protein was identified while there was a clear protein profile in the modern hair. Thus, it is thought that ancient keratin was less strong than modern keratin due to the molecular interactions ensuring protein cohesion inside the fiber.