Evaluation of the Antifungal and Antiproliferative Properties of the Lichen Roccella tinctoria DC. Extracts and Main Components.

In this work, phytochemical analysis on different extracts of Roccella tinctoria DC. was reported using different techniques with respect to the past. Twenty volatile and three non-volatile compounds were identified, some of which were found in this species for the first time. The methanolic extracts and their non-volatile components were then evaluated for their antitumor effects in cancerous A549 and Mz-ChA-1 cells and for their tolerability in non-cancerous BEAS-2B and H69 cells, showing IC50 values from 94.6 µg/mL to 416.4 µg/mL, in general. The same extracts and compounds were also tested for their antifungal effects in Candida albicans, with only compound 2 being active, with an MIC50 value of 87 µg/mL. In addition, they were tested for their anti-Candida adhesion activity, anti-Candida biofilm formation, and anti-Candida mature biofilm inhibition, with efficacy percentages generally above 50% but not for all of them. Lastly, the DF3 extract and compounds 1-2 were tested in vivo according to the Galleria mellonella survival assay, showing positive mortality rates above 50% at different concentrations. All these biological assays were conducted on this species for the first time. Comparisons with other lichens and compounds were also presented and discussed.

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.

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.

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.

Phytochemical analysis on the aerial parts of Teucrium capitatum L. with aspects of chemosystematics and ethnobotany.

The phytochemical analysis on the aerial parts of Teucrium capitatum L. collected from a new population in Central Italy, led to the identification of eight compounds, i.e. pheophytin a (1), poliumoside (2), apigenin (3), luteolin (4), cirsimaritin (5), cirsiliol (6), 8-O-acetyl-harpagide (7) and teucardoside (8) belonging to four different classes of secondary metabolites. Pheophytin a (1) represents a newly identified compound in the genus whereas compounds (7-8) are newly identified compound in the species. The chemotaxonomic and ethnobotanical aspects relative to the presence of these compounds were widely discussed suggesting important conclusions for both.

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.

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.

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.

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.

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.

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.

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.

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.

A new multi analytical approach for the identification of synthetic and natural dyes mixtures. The case of orcein-mauveine mixture in a historical dress of a Sicilian noblewoman of nineteenth century.

In this paper, the application of a multi-analytical approach for the characterisation of synthetic and natural dyes in a historical textile is presented. The work is focused on a historical dress of a Sicilian noblewoman, dating from about 1865-1870. Firstly, SERS on fibre was performed, in order to individuate the classes of dyes employed. The SERS spectra suggested the presence of two main dyes: mauveine and orcein. In order to confirm these preliminary results, two different extraction protocols were applied. The extracts obtained were analysed by ESI-MS, MALDI-ToF and UHPCL-MS analyses, confirming the SERS results. In particular, the application of the ammonia mild extraction technique allowed to selectively extract the phenoxazonic dyes, separating them already in the extraction step from the synthetic ones. Thanks to this multi-analytical approach, this dress could be considered as one of the first examples of employment of synthetic dyes in association with natural ones.

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.

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.

Iridoid glycosides and polyphenolic compounds from Teucrium chamaedrys L.

In this work, the phytochemical analysis of Teucrium chamaedrys L. collected in Italy was reported. Eight compounds were isolated and identified by means of classical column chromatography and spectroscopic techniques, such as NMR and MS. In detail, these compounds were: verbascoside (1), forsythoside b (2), samioside (3), alyssonoside (4), harpagide (5), 8-O-acetyl-harpagide (6), cirsiliol (7) and ß-arbutin (8). The presence of these compounds, in particular iridoids and phenyl-ethanoid glycosides, has a chemotaxonomic relevance and results to be in perfect accordance with the current botanical classification of the species. In addition, it provides a phytochemical rationale for the use of this particular plant in the ethno-pharmacological field. Conversely, it is worth of mention the absence of potentially toxic components, unlike to what observed in other species of the genus which can no longer be used for ethno-medicinal purposes.

Lignans and secoiridoid glycosides from the stem barks of Jasminum tortuosum.

This paper reports on the first phytochemical analysis ever performed on Jasminum tortuosum Willd. This analysis, mainly carried out by means of column chromatography separation, NMR spectroscopy and mass spectrometry, led to the isolation and the identification of four compounds, namely the lignans ginkgool (1) and olivil-4'-O-ß-glucopyranoside (2) and the secoiridoids oleoside dimethyl ester (3) and oleoside 11-methyl ester (4). The presence of these compounds is significant from a chemotaxonomic point of view, confirming the correct botanical classification of the species and, from a phytochemical standpoint, may suggest its possible use in the ethno-medicinal field.