New Advances in Dye Analyses: In Situ Gel-Supported Liquid Extraction from Paint Layers and Textiles for SERS and HPLC-MS/MS Identification.

To date, it is still not possible to obtain exhaustive information about organic materials in cultural heritage without sampling. Nonetheless, when studying unique objects with invaluable artistic or historical significance, preserving their integrity is a priority. In particular, organic dye identification is of significant interest for history and conservation research, but it is still hindered by analytes' low concentration and poor fastness. In this work, a minimally invasive approach for dye identification is presented. The procedure is designed to accompany noninvasive analyses of inorganic substances for comprehensive studies of complex cultural heritage matrices, in compliance with their soundness. Liquid extraction of madder, turmeric, and indigo dyes was performed directly from paint layers and textiles. The extraction was supported by hydrogels, which themselves can undergo multitechnique analyses in the place of samples. After extraction, Ag colloid pastes were applied on the gels for SERS analyses, allowing for the identification of the three dyes. For the HPLC-MS/MS analyses, re-extraction of the dyes was followed by a clean-up step that was successfully applied on madder and turmeric. The colour change perceptivity after extraction was measured with colorimetry. The results showed ΔE values mostly below the upper limit of rigorous colour change, confirming the gentleness of the procedure.

Inside the History of Italian Coloring Industries: An Investigation of ACNA Dyes through a Novel Analytical Protocol for Synthetic Dye Extraction and Characterization.

The introduction of synthetic dyes completely changed the industrial production and use of colorants for art materials. From the synthesis of the first synthetic dye, mauveine, in 1856 until today, artists have enjoyed a wider range of colors and selection of chemical properties than was ever available before. However, the introduction of synthetic dyes introduced a wider variety and increased the complexity of the chemical structures of marketed dyes. This work looks towards the analysis of synthetically dyed objects in heritage collections, applying an extraction protocol based on the use of ammonia, which is considered favorable for natural anthraquinone dyes but has never before been applied to acid synthetic dyes. This work also presents an innovative cleanup step based on the use of an ion pair dispersive liquid-liquid microextraction for the purification and preconcentration of historical synthetic dyes before analysis. This approach was adapted from food science analysis and is applied to synthetic dyes in heritage science for the first time in this paper. The results showed adequate recovery of analytes and allowed for the ammonia-based extraction method to be applied successfully to 15 samples of suspected azo dyes from the Azienda Coloranti Nazionali e Affini (ACNA) synthetic dye collection, identified through untargeted HPLC-HRMS analyses.

Photons for Photography: A First Diagnostic Approach to Polaroid Emulsion Transfer on Paper in Paolo Gioli's Artworks.

The aim of this research is to study and diagnose for the first time the Polaroid emulsion transfer in the contemporary artist Paolo Gioli's artworks to provide preliminary knowledge about the materials of his artworks and the appropriate protocols which can be applied for future studies. The spectral analysis performed followed a multi-technical approach first on the mock-up samples created following Gioli's technique and on one original artwork of Gioli, composed by: FORS (Fiber Optics Reflectance), Raman, and FTIR (Fourier-Transform InfraRed) spectroscopies. These techniques were chosen according to their completely non-invasiveness and no requirement for sample collection. The obtained spectra from FTIR were not sufficient to assign the dyes found in the transferred Polaroid emulsion. However, they provided significant information about the cellulose-based materials. The most diagnostic results were obtained from FORS for the determination of the dye developers present in the mock-up sample which was obtained from Polacolor Type 88 and from Paolo Gioli's original artwork created with Polacolor type 89.

Dyes from the Ashes: Discovering and Characterizing Natural Dyes from Mineralized Textiles.

Vesuvius eruption that destroyed Pompeii in AD 79 represents one of the most important events in history. The cataclysm left behind an abundance of archeological evidence representing a fundamental source of the knowledge we have about ancient Roman material culture and technology. A great number of textiles have been preserved, rarely maintaining traces of their original color, since they are mainly in the mineralized and carbonized state. However, one outstanding textile sample displays a brilliant purple color and traces of gold strips. Since the purple was one of the most exclusive dyes in antiquity, its presence in an important commercial site like Pompeii induces us to deepen the knowledge of such artifacts and provide further information on their history. For this reason, the characterization of the purple color was the main scope of this research, and to deepen the knowledge of such artifacts, the SERS (Surface Enhanced Raman Scattering) in solution approach was applied. Then, these data were enriched by HPLC-HRMS analyses, which confirmed SERS-based hypotheses and also allowed to hypothesize the species of the origin mollusk. In this context, a step-by-step integrated approach resulted fundamental to maximize the information content and to provide new data on textile manufacturing and trade in antiquity.

Deepening Inside the Pictorial Layers of Etruscan Sarcophagus of Hasti Afunei: An Innovative Micro-Sampling Technique for Raman/SERS Analyses.

The Hasti Afunei sarcophagus is a large Etruscan urn, made up of two chalky alabaster monoliths. Dated from the last quarter of the third century BC, it was found in 1826 in the small town of Chiusi (Tuscany- Il Colle place) by a landowner, Pietro Bonci Casuccini, who made it part of his private collection. The noble owner's collection was sold in 1865 to the Royal Museum of Palermo (today under the name of Antonino Salinas Regional Archaeological Museum), where it is still displayed. The sarcophagus is characterized by a complex iconography that is meticulously illustrated through an excellent sculptural technique, despite having subjected to anthropic degradation and numerous restorative actions during the last century. During the restoration campaign carried out between 2016 and 2017, a targeted diagnostic campaign was carried out to identify the constituent materials of the artefact, the pigments employed and the executive technique, in order to get an overall picture of conservation status and conservative criticalities. In particular, this last intervention has allowed the use of the innovative micro-sampling technique, patented by the Cultural Heritage research group of Sapienza, in order to identify the employee of lake pigments through SERS analyses. Together with this analysis, Raman and NMR technique have completed the information requested by restorers, for what concerns the wax employed as protective layers, and allowed to rebuild the conservation history of the sarcophagus. In fact, together with the identification of red ocher and yellow ocher, carbon black, Egyptian blue and madder lake, pigments compatible with the historical period of the work, modern pigments (probably green Paris, chrome orange, barium yellow, blue phtalocyanine) have been recognized, attributable with not documented intervention during the eighteenth and twentieth centuries.

It's Only a Part of the Story: Analytical Investigation of the Inks and Dyes Used in the Privilegium Maius.

The Privilegium maius is one of the most famous and spectacular forgeries in medieval Europe. It is a set of charters made in the 14th century upon commitment by Duke Rudolf IV, a member of the Habsburg family, to elevate the rank and the prestige of his family. These five charters, now kept at the Österreichisches Staatsarchiv in Vienna, have been subjected to a thorough interdisciplinary study in order to shed light on its controversial story. The charters are composed of pergamenaceous documents bound to wax seals with coloured textile threads. The present contribution concerns the characterisation of the inks used for writing and of the dyes used to colour to the threads: Are they compatible with the presumed age of the charters? Though showing only a part of the whole story of the charters, dyes analysis could contribute in assessing their complex history from manufacturing to nowadays. The dyes were characterised with non-invasive in situ measurements by means of fibre optic (FORS) and with micro-invasive measurements by means of Surface Enhanced Raman Spectroscopy (SERS) and High-Performance Liquid Chromatography with Mass Spectrometry (HPLC-MS) analysis. The results showed that the threads of four of the charters (three dyed with madder, one with orchil) were apparently coloured at different dyeing stages, then re-dyed in the 19-20th century.

Iridoids of Chemotaxonomy Relevance, a New Antirrhinoside Ester and Other Constituents from Kickxia spuria subsp. integrifolia (Brot.) R.Fern.

In this work, the secondary metabolite content of the EtOH extract of Kickxia spuria subsp. integrifolia (Brot.) R.Fern. is reported. Fourteen compounds were isolated and identified by means of column chromatography and NMR and MS instrumental techniques, respectively. Among the identified compounds, the chemotaxonomic markers of the species were evidenced, whereas others were reported for the first time in the genus. Among these, a new antirrhinoside derivative (12) was recognized. The iridoid content showed a molecular pattern very similar to those reported for other taxa comprised in the Antirrhinae tribe of Plantaginaceae, thus providing an additional evidence that supports the current botanical classification of the Kickxia genus. Anyway, most of the recognized components are able to exert important pharmacological properties which might suggest the possible employment of also this species in traditional medicine just like it happens for some other species of the Kickxia genus.

The Deposition by Raffaello Sanzio: New analytical insights on old cross sections for the characterisation of pictorial palette.

In 2020, 500th anniversary of Raffaello Sanzio death, his Deposition (1507), -the altarpiece known also as the Pala Baglioni, today located at the Borghese Gallery in Rome-has been subjected to conservative revision and preventive conservation project. This included in-depth diagnostic campaigns through most modern non-invasive techniques, together with the analysis of old cross sections from the same Pala. These latters, prepared between 1966 and 1972, preserved in ICR laboratory of chemistry and testing materials archive, have been used to deepen the knowledge of Raffaello painting techniques. The use of such cross sections was fundamental to verify the original pictorial film and restoration re-paintings before the conservation intervention in the same years. In this paper, the results of analytical insights on Raffaello pictorial palette are presented. The information is obtained by the analysis of the old ICR stratigraphic sections, through the use of Scanning Electron Microscope with Energy Dispersive X-Ray analysis (SEM-EDX) and micro-Raman (632.8 nm), while Surface Enhanced Raman Scattering (SERS) analysis through colloidal paste has been tested for the identification of organic lake-pigments present in low concentration and for the successful recognition of copper resinate, whose SERS spectrum is here reported for the first time, according to our knowledge. This combined diagnostic approach has made it possible to recognize the pigments employed in the different pictorial layers, such species in traces and those from organic materials, responding to open questions arising previous non-invasive analyses and highlighting further aspects of the illustrious master refined painting technique.

Multi-Technique Assessment of Chelators-Loaded PVA-Borax Gel-like Systems Performance in Cleaning of Stone Contaminated with Copper Corrosion Products.

Currently, one of the most important challenges for the conservation of stone artworks is the removal of metal corrosion products on their surfaces. Traditional cleaning methods, which typically involve the application of aqueous solutions containing chelating agents capable of complexing these metal ions, have shown some weaknesses. These weaknesses become apparent when such methods are applied to statues and other vertical surfaces or when aiming to limit the cleaning process to a specific area with controlled application times. Furthermore, the porosity of the stone surface plays a role concerning the cleaning efficiency. To address these issues, chelating agents can be incorporated into gel-like materials. This study is a proof of concept to evaluate the cleaning efficacy of various gel formulations composed of polyvinyl alcohol (PVA), borax (B), and agarose (AG), loaded with two chelators: ethylenediaminetetraacetic acid (EDTA) and potassium sodium tartrate (PST or Rochelle salt). Three types of carbonate stones (travertine, Lecce stone, and Carrara marble) characterized by different porosities were artificially stained with copper sulphates and treated with the different PVA-B-AG formulations. The effectiveness of the treatment was directly monitored on the stones using a multi-technique approach that included scanning electron microscopy with energy dispersive spectroscopy (SEM-EDS) and non-invasive portable nuclear magnetic resonance (NMR). Additionally, the rheological properties of the gels were investigated, and the Fourier transform infrared attenuated total reflection spectroscopy (FTIR ATR) was used to analyse the chemical structure of the gel before and after treatment, aiming to understand the changes induced by the cleaning process.

Early Stages of Metal Corrosion in Coastal Archaeological Sites: Effects of Chemical Composition in Silver and Copper Alloys.

In this study, metal disks with different chemical composition (two Ag-based alloys and three Cu-based alloys) were buried in the soil of coastal archaeological sites for a period of 15 years. The aim was to naturally induce the growth of corrosion patinas to obtain a deeper insight into the role of alloying elements in the formation of the patinas and into the degradation mechanisms occurring in the very early stages of burial. To reach the aim, the morphological, compositional and structural features of the patinas grown over 15 years were extensively characterized by optical microscopy, field emission scanning electron microscopy coupled with energy dispersive spectrometry, X-ray diffraction and micro-Raman spectroscopy. Results showed that the Cu amount in Ag-based alloys strongly affected the final appearance, as well as the composition and structure of the patinas. Corrosion mechanisms typical of archaeological finds, such as the selective dissolution of Cu, Pb and Zn and internal oxidation of Sn, occurred in the Cu-based alloys, even if areas enriched in Zn and Pb compounds were also detected and attributed to an early stage of degradation. In addition, some unusual and rare compounds were detected in the patinas developed on the Cu-based disks.

Applying Gel-Supported Liquid Extraction to Tutankhamun's Textiles for the Identification of Ancient Colorants: A Case Study.

The identification of the dyes present on a linen fragment from the tomb of Pharaoh Tutankhamun is the objective of the present study. Fiber optic reflectance spectroscopy (FORS) was applied to the archaeological sample for preliminary identification of the dyes and to better choose the extraction methodology for different areas of the sample. The innovative gel-supported micro-extraction with agar gel and the Nanorestore Gel® High Water Retention (HWR) gel were applied to the archaeological sample after testing of the best concentration for the extraction of the agar gels substrates, performed on laboratory mock-ups by means of UV-Vis transmittance spectroscopy. Immediately after extraction, Ag colloidal pastes were applied on the gel surface and Surface Enhanced Raman Scattering (SERS) analysis was performed directly on them. The combination of information deriving from FORS and SERS spectra resulted in the successful identification of both indigo and madder and, in hypothesis, of their degradation products.

Unearthed opium: development of a UHPLC-MS/MS method for the determination of Papaver somniferum alkaloids in Daunian vessels.

Introduction: The analysis of organic residue in ancient vessels to investigate early-age civilization habits is an important archeological application that needs advanced analytical methods. However, these procedures should meet inherent requisites such as low sampling invasiveness and high sensitivity for trace analysis. This study deals with the development of advanced analytical methods for the detection of opium alkaloids in ceramic vessels and its first application to the study of Daunian pots dating back to the VIII-IV sec BC. Methods: All the stages of the analytical procedure, from sampling to analysis, were carefully optimized. Concerning sampling, the traditional scraping approach was compared with a swabbing strategy which permitted minimizing sample encroachment. Extraction was based on pressurized liquid extraction or ultrasound-assisted liquid extraction, followed by dispersive liquid-liquid microextraction, which allowed concentration enrichment. On the other hand, a UHPLC-MS/MS method was specifically developed and validated to obtain reliable data. Some Daunian pots, belonging to the Ceci-Macrini private archeological collection, were selected for sample withdrawal as their iconography could suggest opium usage. Results: Several of the analyzed samples resulted positive to thebaine and less frequently to morphine and codeine; furthermore, 70% of the analyzed items tested positive for at least one opium alkaloid. Positive findings were common to all the samples collected in the pots, suggesting that scraping and swabbing provided comparable results and validating this unusual sampling strategy. All samples were additionally analyzed by UHPLC-HRMS to further improve the confidence level of the identified compounds. The obtained results shed new light on the hypothesis of opium usage by the ancient Daunian civilization. Furthermore, this study provided suitable analytical tools for further investigations on the same topic, with a good level of confidence in the quality of the results.

Non-Invasive Assessment of PVA-Borax Hydrogel Effectiveness in Removing Metal Corrosion Products on Stones by Portable NMR.

The cleaning of buildings, statues, and artworks composed of stone materials from metal corrosion is an important topic in the cultural heritage field. In this work the cleaning effectiveness of a PVA-PEO-borax hydrogel in removing metal corrosion products from different porosity stones has been assessed by using a multidisciplinary and non-destructive approach based on relaxation times measurement by single-sided portable Nuclear Magnetic Resonance (NMR), Scanning Electron Microscopy-Energy Dispersive Spectroscopy (SEM-EDS), and Raman Spectroscopy. To this end, samples of two lithotypes, Travertine and Carrara marble, have been soiled by triggering acidic corrosion of some copper coins in contact with the stone surface. Then, a PVA-PEO-borax hydrogel was used to clean the stone surface. NMR data were collected in untreated, soiled with corrosion products, and hydrogel-cleaned samples. Raman spectroscopy was performed on PVA-PEO-borax hydrogel before and after cleaning of metal corrosion. Furthermore, the characterization of the dirty gel was obtained by SEM-EDS. The combination of NMR, SEM-EDS and Raman results suggests that the mechanism behind the hydrogel cleaning action is to trap heavy metal corrosion products, such as Cu2+ between adjacent boron ions cross-linked with PVA. Moreover, the PVA-PEO-borax hydrogel cleaning effectiveness depends on the stone porosity, being better in Carrara marble compared to Travertine.

Street art graffiti: Discovering their composition and alteration by FTIR and micro-Raman spectroscopy.

Paints used in street art are modern materials subjected to degradation processes, which are very complex and difficult to predict without taking into account of several factors. This study investigates three outdoor murals in Lazio, - namely "graffiti", a word now used to indicate a spontaneous street art tendency consisting in images and writings realized by spray paints in public spaces to provoke passersby -with the aim to discover materials application techniques and chemical composition and figure out whether alteration phenomena occurred. Twenty-two samples were collected, and their stratigraphy was studied by optical microscopy. Fourier Transformed Infrared spectroscopy was used to identify binders and their degradation products in paints and preparatory layers, while for characterization of organic pigments used in all different stratigraphy layers of samples micro-Raman spectroscopy analyses was carried out. Furthermore, micro-Raman spectroscopy allowed to study an unusual patina formed on the surface of a pink paint. This information is useful for artists as well as for conservators, who must face numerous issues related to the preservation of this modern and labile kind of artistic expression, very fashionable nowadays but often created without care for materials duration. Conservation issues were also deepened by interviews with several contemporary mural authors. Artists underlined how contemporary murals are a very heterogeneous means of expression. Different cultural tendencies coexisting result in different attitude towards conservation.

Lucio Fontana and the light: Spectroscopic analysis of the artist's collection at the National Gallery of Modern and Contemporary Art.

In this work the results deriving from the characterization of materials used by Lucio Fontana to realize some of his artworks are presented. Specifically, object of analyses are three artworks from the collection of National Gallery of Modern and Contemporary Art in Rome, whose complex composition required the combination of different diagnostic techniques for a complete characterization. Microscopic fragments from the artworks were analyzed through X-Ray Fluorescence (XRF), Fourier Transformed InfraRed (FTIR) spectroscopy, in transmission and in Attenuated Total Reflectance (ATR) mode, and Raman spectroscopy to obtain information on the atomic and molecular species and to individuate pigments and binders. In one case, the richness in different materials and the not homogenous distribution on the artwork surface required the additional use of portable techniques, as Visible Light Reflectance and Raman spectroscopies with optical fiber probes, to map the surface. The combination of these techniques resulted to be a powerful tool in order to obtain a complete characterization of artist's choices and provided new information to understand the evolution of his technique.

A new byciclic monoterpene glucoside and a new biflavone from the male reproduction organs of Wollemia nobilis.

In this work, the fifth part of an ongoing phytochemical study on Wollemia nobilis was reported. The attention was now focused on the male reproduction organs of which the content in both primary and secondary metabolites was analyzed. Twenty compounds, belonging to seven different classes of natural compounds, were identified from the ethanolic extract by means of Column Chromatography and NMR and MS Spectroscopy. They all represent new compounds for the studied organ whereas some of them are also new constituents of the genus or even previously undescribed phytochemicals. Their presence was able to display a general overview of these organs from the phytochemical standpoint and to provide more elements in confirmation with the current botanical classification of the species. Moreover, they add a further experimental evidence of the tendency of this species to accumulate different metabolites in different organs. This characteristic as well as the occurrence of several compounds with added value, make this plant a possible candidate for large scale cultivation with extractive purposes to obtain useful phytochemicals for botanicals and pharmaceutical fields. Moreover, they offer the opportunity to develop an additional method of conservation and protection for this endangered and very rare species.

Pedicularis L. Genus: Systematics, Botany, Phytochemistry, Chemotaxonomy, Ethnopharmacology, and Other.

In this review, the relevance of the plant species belonging to the Pedicularis L. genus has been considered from different points of view. Particular emphasis was given to phytochemistry and ethnopharmacology, since several classes of natural compounds have been reported within this genus and many of its species are well known to be employed in the traditional medicines of many Asian countries. Some important conclusions on the chemotaxonomic and chemosystematic aspects of the genus have also been provided for the first time. Actually, this work represents the first total comprehensive review on this genus.

New Developments in the Synthesis of EMICORON.

This paper reports on the modification of two synthetic steps in the usual protocol used for obtaining EMICORON. EMICORON is a benzo[ghi]perylen-diimide, which was synthesized for the first time in our laboratory in 2012, and has shown to have in vivo antitumor activities that interferes with the tumor growth and development using a multi-target mechanism of action. The provided modifications, which involved the reaction times, the reaction conditions, and the work-up procedures, allowed the global yield of the process to be increased from 28% to about 40%. Thus, this new procedure may be more suitable for recovering higher amounts of EMICORON to be used in further preclinical studies.