Carlo Crescenzi

This study explores the feasibility of using headspace solid-phase microextraction (HS-SPME) combined with gas chromatography-mass spectrometry (GC-MS) to analyse the volatilome of human melanoma A375 cells. The goal was to identify and characterise the volatile organic compounds (VOCs) and monitor alterations induced by treatment with thapsigargin (TG), a drug known to disrupt calcium homeostasis, thereby inducing endoplasmic reticulum stress and ultimately triggering cell death. Reproducibility of experimental conditions is a major issue in biological experiments, which presents intrinsic variability of the samples to be analysed. In our case, initial analysis revealed a significant batch effect, accounting for 56.88% of the total variance. To address this, external parameter orthogonalisation (EPO) was applied, which successfully reduced the batch variance to just 0.16%. After this correction, the treatment factor became the dominant source of variation, explaining 47.12% of the total variance with strong statistical significance ( p -value = 0.001). A supervised classification model using partial least squares discriminant analysis (PLS-DA) was developed and validated to characterise the differences between treated and untreated cells. The model achieved a mean overall accuracy of 92.21% and an area under the curve (AUC) of 0.974, indicating excellent discrimination between the two classes. The robustness of these findings was confirmed by repeated double cross-validation and permutation testing, which showed that the model’s predictive ability was not due to random chance. The results demonstrate that TG treatment induces a reproducible and highly discriminant volatilome signature in A375. This suggests that VOCs could potentially serve as biomarkers for monitoring cellular responses to drug treatments. Graphical abstract Graphical abstract created with BioRender.com

A first general and practical method for the synthesis of valuable 2-(hetero)aryl glycidic ethyl esters has been developed using commercially available reagents and a catalyst. A telescoped three-step seven-reaction process, based on a Knoevenagel/nitro-Michael/hydroxylation/double elimination/epoxidation/esterification sequence, provides the epoxides in up to 73% overall yield. This one-pot protocol features (i) aldehydes as versatile feedstocks, among the reagents used in the process; (ii) all steps proceeding at room temperature in benign solvents; and (iii) scalability of the reaction up to 5 mmol. The epoxides are elaborated to obtain new attractive β-amino α-hydroxy esters, bearing a quaternary stereocenter, including tryptamine- and morpholine-based esters.

The huge amount of citrus peel produced worldwide represents an economic burden for society. However, this agricultural by-product is a rich source of natural molecules, potentially endowed with interesting pharmacological activities. In this regard, we decided to investigate if the polymethoxyflavones contained in citrus peel waste could be exploited as novel vasorelaxant agents. A hydroalcoholic blond orange (Citrus sinensis) peel extract, obtained by ultrasonication, was partitioned in dichloromethane. Column chromatography allowed for the isolation of four polymethoxyflavones, namely, scutellarein tetramethyl ether, nobiletin, tangeretin, and sinensetin, identified by nuclear magnetic resonance (NMR) spectroscopy and UPLC-HRMS/MS and confirmed by multivariate curve resolution of NMR fractional spectra. The four molecules showed interesting in vitro vasorelaxant activity, at least, in part, due to the blockade of smooth muscle Ca_V1.2 channels. Molecular modeling and docking analysis elucidated the binding mode of the polymethoxyflavones at the homology model of the rat Ca_V1.2c subunit and provided the structural basis to rationalise the highest activity of scutellarein tetramethyl ether in the set and the dramatic effect of the additional methoxy group occurring in nobiletin and sinensetin. In conclusion, citrus peel can be considered a freely available, valuable source of vasoactive compounds worthy of pharmaceutical and/or nutraceutical exploitation.

Sphingolipids play crucial roles in cellular membranes, myelin stability, and signalling responses to physiological cues and stress. Among them, sphingosine 1-phosphate (S1P) has been recognized as a relevant biomarker for neurodegenerative diseases, and its analogue FTY-720 has been approved by the FDA for the treatment of relapsing–remitting multiple sclerosis. Focusing on these targets, we here report three novel polymeric capture phases for the selective extraction of the natural biomarker and its analogue drug. To enhance analytical performance, we employed different synthetic approaches using a cationic monomer and a hydrophobic copolymer of styrene-DVB. Results have demonstrated high affinity of the sorbents towards S1P and fingolimod phosphate (FTY-720-P, FP). This evidence proved that lipids containing phosphate diester moiety in their structures did not constitute obstacles for the interaction of phosphate monoester lipids when loaded into an SPE cartridge. Our suggested approach offers a valuable tool for developing efficient analytical procedures. Graphical Abstract

Many studies have explored the extraction of bioactive compounds from different onion solid wastes, such as bulb, skin, and peel. However, onion leaves have received limited attention despite their potential as a valuable source of nutraceutical compounds. This study aimed to valorise, for the first time, the agricultural waste in the form of spring onion leaves (CN, Cipollotto Nocerino) to obtain antioxidant-rich polyphenolic extracts. A Box–Behnken design (BBD) was used to assess the impact of microwave-assisted extraction (MAE) variables (temperature, time, extraction volume, and ethanol concentration) on total polyphenol content (TPC) measured by Folin–Ciocalteu method and the antioxidant power determined by FRAP assay. Response surface methodology (RSM) was applied, and regression equations, analysis of variance, and 3D response curves were developed. Our results highlighted that the TPC values range from 0.76 to 1.43 mg GAE g⁻¹ dw, while the FRAP values range from 8.25 to 14.80 mmol Fe(II)E g⁻¹ dw. The optimal extraction conditions predicted by the model were 60 °C, 22 min, ethanol concentration 51% (v/v), and solvent volume 11 mL. These conditions resulted in TPC and FRAP values of 1.35 mg GAE g⁻¹ dw and 14.02 mmol Fe(II)E g⁻¹ dw, respectively. Furthermore, the extract obtained under optimized conditions was characterized by UHPLC-ESI-Orbitrap-MS analysis. LC/MS–MS platform allowed us to tentatively identify various compounds belonging to the class of flavonoids, saponins, fatty acids, and lipids. Finally, the ability of CN optimal extract to inhibit the intracellular reactive oxygen species (ROS) release in a hepatocarcinoma cell line using an H₂O₂-induced oxidative stress model, was evaluated. The results highlighted the potential of CN extract as a valuable source of polyphenols with significant antioxidant properties, suitable for various applications in the food and pharmaceutical industries.

In this study we explore the potential of combining traditional zooarchaeological determination and proteomic identification of morphologically non-diagnostic bone fragments (ZooMS) collected from the Uluzzian levels of three Italian sites: Uluzzo C Rock Shelter, Roccia San Sebastiano cave, and Riparo del Broion. Moreover, we obtained glutamine deamidation ratios for all the contexts analysed during routine ZooMS screening of faunal samples, giving information on collagen preservation. We designed a selection protocol that maximizes the efficiency of the proteomics analyses by excluding particularly compromised fragments (e.g. from taphonomic processes), and that aims to identify new human fragments by favouring bones showing morphological traits more similar to Homo. ZooMS consistently provided taxonomic information in agreement with the faunal spectra outlined by traditional zooarchaeology. Our approach allows us to delineate and appreciate differences between the analysed contexts, particularly between the northern and southern sites, related to faunal, environmental, and climate composition, although no human remains were identified. We reconstructed the faunal assemblage of the different sites, giving voice to morphologically undiagnostic bone fragments. Thus, the combination of these analyses provides a more complete picture of the faunal assemblage and of the paleoenvironment during the Middle-Upper Palaeolithic transition in Italy.

An enantioselective one‐pot catalytic strategy to dihydroquinoxalinones, featuring novel 1‐phenylsulfonyl‐1‐cyano enantioenriched epoxides as masked α‐halo acyl halide synthons, followed by a domino ring‐opening cyclization (DROC), is documented. A popular quinine‐derived urea served as the catalyst in two out of the three steps performed in the same solvent using commercially available aldehydes, (phenylsulfonyl)acetonitrile, cumyl hydroperoxide and 1,2‐phenylendiamines. Medicinally relevant 3‐aryl/alkyl‐substituted heterocycles are isolated in generally good to high overall yield and high enantioselectivity (up to 99 % ee). A rare example of excellent reusability of an organocatalyst at higher scale, subjected to oxidative conditions, is demonstrated. Mechanistically, labile α‐ketosulfone has been detected as the intermediate involved in the DROC process. Theoretical calculations on the key epoxidation step rationalize the observed stereocontrol, highlighting the important role played by the sulfone group.