Periodic Reporting for period 2 - PARCA (Advance in Proteomics and Analysis of dyes and Recovery of Charred and Aged textiles)
Reporting period: 2024-02-01 to 2025-01-31
Background. Archaeological textiles hold significant importance in comprehending past cultures and histories. They play a crucial role in confirming or validating historical sources, shedding light on the utilization of specific species, providing evidence of trade routes, and indicating technological levels. However, textiles often face complex preservation challenges, such as susceptibility to various breakdown processes like insect damage, fire, different degrees of carbonization, soil contamination, and partial or total mineralization. These textiles are typically composed of proteinaceous fibers, including those derived from hair, such as furs and wool, composed mainly of keratins (IFs) and keratins associated proteins (KAPs). In addition to the protein component, textiles are often dyed, and the identification of dyes provides valuable information for reconstructing the socio-cultural background of the textile.
Aim of the project. The PARCA project addresses both components of wool textiles—dyes, specifically anthraquinone ones, and keratins and keratin-associated proteins. The aim is to individuate an innovative protocol to demonstrate the possibility of achieving extraction and characterization of both component in a single workflow, getting yields comparable to individual analytical processes. This means the sample processing time and the number of samples required is strongly reduced. The protocol is first developed on dyed and undyed wool specimens and then applied to textile mock-ups subjected to varying degrees of thermal aging, with the aim of evaluate the modifications in protein pattern step by step at different temperatures. The ultimate goal is to apply such protocols to semi- or fully charred archaeological specimens, thus providing a new tool for understanding the degradative processes linked to the events that triggered them, as in the case of the eruption of Mount Vesuvius in 79 AD, thus allowing us to look at both sides of the same coin.
Aim of the project. The PARCA project addresses both components of wool textiles—dyes, specifically anthraquinone ones, and keratins and keratin-associated proteins. The aim is to individuate an innovative protocol to demonstrate the possibility of achieving extraction and characterization of both component in a single workflow, getting yields comparable to individual analytical processes. This means the sample processing time and the number of samples required is strongly reduced. The protocol is first developed on dyed and undyed wool specimens and then applied to textile mock-ups subjected to varying degrees of thermal aging, with the aim of evaluate the modifications in protein pattern step by step at different temperatures. The ultimate goal is to apply such protocols to semi- or fully charred archaeological specimens, thus providing a new tool for understanding the degradative processes linked to the events that triggered them, as in the case of the eruption of Mount Vesuvius in 79 AD, thus allowing us to look at both sides of the same coin.
In the outgoing period of the project, specifically carried out at the Museum Conservation Institute - MCI (Smithsonian Institution), the identification of suitable procedure to get the simultaneous extraction of dyes and protein from a single sample was achieved. This step, developed on madder, cochineal, indigo and weld dyed and undyed wool samples, actually brought to two different protocols, which can be used based on the state of conservation of the textile (i.e. for a strong soil contamination, one is more effective than the other): a slightly modified urea protocol and a recently proposed new TCEP/CAA procedure.
In addition, different methods of cleanup and workflow for proteins and dyes were investigated to develop protocols that did not result in a loss of aliquots of the analytes of interest and to maximize the recovery of both components from the extracting solution. These protocols investigated the application of two types of paramagnetic beads, unmodified and carboxylate-coated hydrophilic magnetic beads, and dialysis and stage-tip protocols.
This aimed at ensuring the same high efficiency in terms of analytical sensitivity for both dye and protein, as in their single workflow.
Once the best procedures were identified, during the second year they were applied to same kind of specimens but subjected to a thermal aging protocol. The laboratory dyed and undyed mock-ups were subjected to a progressive thermal ramp, from 200°C to 300°C at different times, thus trying to simulate the conditions of charred archaeological samples to evaluate its effectiveness and determine the survival threshold of proteins and dyes at the various thermal aging steps. These data were compared with FTIR analysis and morphological information by SEM, to also have a correlation between destructive analysis and morphological appearance and nondestructive investigations. A threshold of protein survival was defined, thus allowing us to trace a path for the first time in evaluating the degradation process and the temperatures reached based on protein modifications.
In returning phase at Sapienza University of Rome, Dept of Environmental Biology, the project focused on evaluating the latest aging to reconstruct the trend of protein modifications at different thermal conditions and photo-oxidative stress conditions and then move on to analyzing the archaeological samples from the Vesuvian Area and Greek area. This fundamental step, currently being finalized, aims to validate and enrich the information obtained from the mock-ups by comparing it with unique findings, revealing new information which may be a new key to open a door on the history and events of these materials.
In addition, different methods of cleanup and workflow for proteins and dyes were investigated to develop protocols that did not result in a loss of aliquots of the analytes of interest and to maximize the recovery of both components from the extracting solution. These protocols investigated the application of two types of paramagnetic beads, unmodified and carboxylate-coated hydrophilic magnetic beads, and dialysis and stage-tip protocols.
This aimed at ensuring the same high efficiency in terms of analytical sensitivity for both dye and protein, as in their single workflow.
Once the best procedures were identified, during the second year they were applied to same kind of specimens but subjected to a thermal aging protocol. The laboratory dyed and undyed mock-ups were subjected to a progressive thermal ramp, from 200°C to 300°C at different times, thus trying to simulate the conditions of charred archaeological samples to evaluate its effectiveness and determine the survival threshold of proteins and dyes at the various thermal aging steps. These data were compared with FTIR analysis and morphological information by SEM, to also have a correlation between destructive analysis and morphological appearance and nondestructive investigations. A threshold of protein survival was defined, thus allowing us to trace a path for the first time in evaluating the degradation process and the temperatures reached based on protein modifications.
In returning phase at Sapienza University of Rome, Dept of Environmental Biology, the project focused on evaluating the latest aging to reconstruct the trend of protein modifications at different thermal conditions and photo-oxidative stress conditions and then move on to analyzing the archaeological samples from the Vesuvian Area and Greek area. This fundamental step, currently being finalized, aims to validate and enrich the information obtained from the mock-ups by comparing it with unique findings, revealing new information which may be a new key to open a door on the history and events of these materials.
Results reached so far have shown that it was possible to extract dyes and proteins simultaneously from a single sample, devising a workflow that does not lose the analytical sensitivity of the two individual processes. This has several effects in the field of archaeometric analysis of archaeological fabric: it is possible to strongly reduce the number of samples required from archaeological artifacts, which are valuable and unique, and to maximize the information content obtained from the single extraction. In addition, the use of clean-up methods for isolation and purification of individual components, in some cases applied for the first time to these matrices, have in some cases improved extraction yields. Such protocols have also been applied to thermal-aged samples, simulating the conditions of charred archaeological samples, to evaluate the threshold of protein survival, related to the reached temperature. The new methodology offers new important insight on such precious relics, not only identifying the material employed, but also its degradation state and the processes it has been subjected to (i.e. temperature reached, burying condition). Thus, when it is applied to real archeological samples, it would be able to give historian or archaeologists novel important clues about historical events these manufacts have been subjected to.