Chemistry at the service of art

Understanding how the great painters such as Modigliani and da Vinci worked, determining the impact of restoration treatment on a work of art, knowing how to conserve it or even where it comes from... these are the subjects of Caroline Tokarski's research. However, she is no historian, nor is she a museum curator. Caroline Tokarski is a professor at the University of Bordeaux and a chemist at the Institute of Chemistry and Biology of Membranes and Nano-objects (CBMN - Bordeaux INP, CNRS and University of Bordeaux). The tool she and her research team use daily is a high-resolution mass spectrometer. Their preferred objects of study: proteins, lipids and sugars (or polysaccharides).

As head of the Proteome platform within the university's Health sciences and technologies research department, and a pioneer in the field, the researcher has been developing original proteomic analysis methods for the past 20 years - making it possible to identify all the proteins in a sample - applied in particular to the specific field of cultural heritage objects (works of art or bones).

Rebuilding the blocks

How is this method a real step forward? Because until recently, analysis methods enabled scientists to identify the basic building blocks of proteins, amino acids, without knowing how they were arranged. It's a bit like having a pile of a thousand stone blocks in front of you and not knowing whether they were all used to build a castle... or a pyramid!

The instrument that can be used to answer these questions, measuring masses that can provide information on the chemical composition, physical state or arrangement of molecules, is the high-resolution mass spectrometer. Thanks to its analyses, Caroline Tokarski's team has been able to identify, for example, that egg yolks and/or whites, milk, oils or animal glues have previously been used by painters to bind colour pigments.

"Proteins are enclosed in highly complex matrices that are modified by time and can be subject to environmental stresses such as temperature or humidity. We first need to extract these proteins and then extract structural information from them. It's always a challenge," she explains. Working with works of art requires the smallest sampling possible. It would be unthinkable to cut out a piece of Leonardo da Vinci's Last Supper, which is the subject of a European research project, in order to understand its composition! The samples taken from the works in the museums with which Prof. Tokarski collaborates are therefore very limited in size and quantity.

The sample was even invisible to the naked eye in a recent study published in the international journal Science Advances and featured in Nature. "The sample was taken using an abrasive film," explains Caroline Tokarski.
In this example, ivory was the focus of the study, the same ivory used to create the objects conserved at the Metropolitan Museum of Art (MET) in New York, a museum with which the Bordeaux researcher has been coordinating the CNRS ARCHE international laboratory for several years. This material, prized by many cultures for its aesthetic qualities, is commonly found in museum collections.

"With the limited quantities of samples and therefore proteins available, we have been able to reach previously inaccessible levels of information by optimising our preparation methods and adapting the associated analytical and data processing methods".

Ivory is composed of enamel and dentine in which collagen is found, a protein regularly targeted by the research team.
Thanks to numerous samples supplied by the American Museum of National History in New York, the researchers were able to build a database of species that are usually under-represented. By comparing the experimental data acquired, this database was then used to differentiate elephant ivory from hippopotamus ivory, sperm whale and even mammoth ivory, as well as from the horns and bones of deer and cattle.

Differentiating elephant ivory from hippopotamus ivory... on the basis of a single amino acid

While ivory from different species can be recognised by morphological analysis by specialists in the field, highly worked, polished or even weathered artefacts are more difficult to identify, explains Caroline Tokarski. "With the limited quantities of samples and therefore proteins available, we have been able to reach previously inaccessible levels of information by optimising our preparation methods and adapting the associated analytical and data processing methods".

Collagens are composed of several thousand constituent amino acids and the differences in sequences between elephant and hippopotamus collagen found in ivory objects can be resolved with the single side chain of two amino acids with very similar structures (with the same mass)! In this case, mass spectrometric sequencing of all or part of the protein chain was not enough: the species difference had to be detected down to the very last amino acid. Certain chemical modifications (such as deamidation, for example) can also alter an ancient object and change it over time. These phenomena, which are veritable chemical clocks, use these techniques to differentiate between older and more recent objects.

Thanks to advances in these analytical techniques, and in particular proteomic analysis, the researchers were able to determine the biological origins of ivory and bone from objects dating back to 4,000 BC. "Some objects from the Egyptian period had never been analysed, as the analysis methods available at the time were deemed too invasive," explains Caroline Tokarski.

While this micro-invasive method of analysis is of direct interest to museums, it could also have another, more surprising application, in ivory trafficking controls. International law enforcement agencies currently use morphological analysis (visual identification) or DNA analysis (which requires a lot of samples) to differentiate between elephant ivory (banned) and mammoth ivory (unregulated). They could benefit from this new technique to step up their controls while limiting the number of samples taken.
The development of techniques, while limiting the quantities of samples, is now enabling an increase in the quantity and quality of the information generated, allowing Caroline Tokarski to learn even more about the objects of the past.