Structure and stability of biological materials – characterisation at the nanoscale

Mummies are witnesses of the past harbouring information about the
lives and fates of our ancestors. By examining them, the conditions
of living, dietary, lifestyle and cultural habits as well as
maladies in ancient times can be revealed. Knowledge of these
maladies can be used to ascertain the evolution of diseases and may
be helpful in characterising and treating them today. Uncovering
information from mummies, however, depends on the preservation of
the mummy tissue. Once degradation sets in, the molecular structure
of the tissue is changed, and much information is lost. Favourable
environmental conditions can slow down the process of decay and,
hence, preserve organic material for long periods of time. As
discussed in this work, biological tissue, which has substructural
arrangements that are advantageous for withstanding mechanical
load, might also be particularly favourable for preservation after
the organism’s death. To address the question concerning the degree
of preservation and to retrieve additional information from ancient
tissue, two quasi-non-invasive analysis techniques, atomic force
microscopy and Raman spectroscopy, were used. With these methods,
the submicron structure, chemical composition, and nanomechanical
properties of small mummified tissue samples were determined. In
preliminary tests on recent collagen, the main connective tissue
protein of vertebrates, results showed that in addition to imaging
by atomic force microscopy, Raman spectroscopy is able to verify
the alignment of this protein. Based on this knowledge, the
arrangement and degree of collagen preservation in mummified human
skin was investigated. Samples extracted from a 5300-year-old
glacier mummy, the Iceman, were analysed. Extremely well-preserved
collagen fibrils, in which the micro, ultra, and molecular
structure were largely unaltered, were found. These results were in
contrast, to the collagen fibrils found in the dermis of the
Zweeloo mummy, a bog body of a female dating to the Roman period
(78–233 AD). The Zweeloo mummy collagen fibrils showed moderate
decomposition likely due to the acidic environment in the bog.
Therefore, mummification due to freeze-drying, as in the Iceman,
seems to be particularly beneficial for tissue preservation. The
Iceman collagen, moreover, was found to be slightly stiffer than
recent collagen, indicating that dehydration due to freeze-drying
changed the mechanical properties of the tissue. This change likely
improves the resilience of the freeze-dried collagen, stiffens the
skin, and in turn maintains the skin’s protective function that
prevents the underlying tissue from decomposing. Finally, also the
preservation of red blood cells in wound tissue samples from the
Iceman was observed. Single and clustered red blood cells were
found whose morphological and molecular characteristics were
similar to those of recent red blood cells. The ancient corpuscles
moreover featured the typical red blood cell structure that
indicates the preservation of healthy cells in Iceman tissue.
Because fibrin, a protein formed during blood coagulation, was also
detected, it appears that the clustered cells resembled remnants of
a blood clot. The structure of the blood clot, stabilised by
fibrin, may have been a protective envelope, which prevented the
red blood cells from decomposing. Nonetheless, Raman spectra of the
cells provided first indications of slight red blood cell
degradation. These investigations emphasise the fundamental
importance of the substructure and molecular arrangement of
tissues, indicating that a tissue’s overall function and stability
correlate with its molecular properties, in particular, the degree
of cross-linking and the arrangement of the tissue molecular
constituents. Last but not least the results show that ancient
tissue can be preserved and its molecular properties probed and
addressed even after millennia.