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S -phenylmercapturic acid (PMA) is a specific urinary biomarker of benzene at exposure levels lower than 1  ppm. However, measuring PMA in urine is an expensive task by either GC or HPLC due to the necessity of extensive sample pretreatment. In the present study, a commercial chemiluminescence enzyme-linked immunosorbent assay (ELISA) test for PMA and GC-MS were used for screening urine samples of 60 workers employed in petrochemical settings. The ELISA results were evaluated by comparison with the GC-MS. Overall, the ELISA test proved sensitive (limit of detection = 0.1  µg  l -1 ), rapid, robust and reliable, affording results in good agreement with the GC-MS (54% of measurements) and no false-negatives. On the other hand, 46% of the ELISA assays were assigned as false-positives (arbitrarily established when ELISA >5  µg  l -1 , GC-MS [ABSTRACT FROM AUTHOR]

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The Heck reaction is widely employed to build a variety of biologically relevant scaffolds and has been successfully implemented in the production of active pharmaceutical ingredients (APIs). Typically, the reaction with terminal alkenes gives high yields and stereoselectivity toward the trans- substituted alkenes product, and many green variants of the original protocol have been developed for such substrates. However, these methodologies may not be applied with the same efficiency to reactions with challenging substrates, such as internal olefins, providing trisubstituted alkenes. In the present work, we have implemented a Heck reaction protocol under green conditions to access trisubstituted alkenes as final products or key intermediates of pharmaceutical interest. A set of preliminary experiments performed on a model reaction led to selecting a simple and green setup based on a design of experiments (DoE) study. In such a way, the best experimental conditions (catalyst loading, equivalents of alkene, base and tetraalkylammonium salt, composition, and amount of solvent) have been identified. Then, a second set of experiments were performed, bringing the reaction to completion and considering additional factors. The protocol thus defined involves using EtOH as the solvent, microwave (mw) irradiation to achieve short reaction times, and the supported catalyst Pd EnCat ® 40, which affords an easier recovery and reuse. These conditions were tested on different aryl bromides and internal olefines to evaluate the substrate scope. Furthermore, with the aim to limit as much as possible the production of waste, a simple isomerization procedure was developed to convert the isomeric byproducts into the desired conjugated E alkene, which is also the thermodynamically favoured product. The approach herein disclosed represents a green, efficient, and easy-to-use handle towards different trisubstituted alkenes via the Heck reaction., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2024 Rossino, Marrubini, Brindisi, Granje, Linciano, Rossi and Collina.)

Despite the considerable steps taken in the last decade in the context of antineoplastic drug (AD) handling procedures, their mutagenic effect still poses a threat to healthcare personnel actively involved in compounding and administration units. Biological monitoring procedures usually require large volumes of sample and extraction solvents, or do not provide adequate sensitivity. It is here proposed a fast and automated method to evaluate the urinary levels of cyclophosphamide and iphosphamide, composed of a miniaturized solid phase extraction (µSPE) followed by ultrahigh-performance liquid chromatography-tandem mass spectrometry (UHPLC-MS/MS) analysis. The extraction procedure, developed through design of experiments (DoE) on the ePrep One Workstation, required a total time of 9.5 min per sample, with recoveries of 77-79% and a solvent consumption lower than 1.5 mL per 1 mL of urine sample. Thanks to the UHPLC-MS/MS method, the limits of quantification (LOQ) obtained were lower than 10 pg/mL. The analytical procedure was successfully applied to 23 urine samples from compounding wards of four Italian hospitals, which resulted in contaminations between 27 and 182 pg/mL.

Formaldehyde (FA) is a toxic compound and a human carcinogen. Regulating FA-releasing substances in commercial goods is a growing and interesting topic: worldwide production sectors, like food industries, textiles, wood manufacture, and cosmetics, are involved. Thus, there is a need for sensitive, economical, and specific FA monitoring tools. Solid-phase microextraction (SPME), with O -(2,3,4,5,6-pentafluorobenzyl)-hydroxylamine (PFBHA) on-sample derivatization and gas chromatography, is proposed for FA monitoring of real-life samples. This study reports the use of polydimethylsiloxane (PDMS) as a sorbent phase combined with innovative commercial methods, such as multiple SPME (MSPME) and cooling-assisted SPME, for FA determination. Critical steps, such as extraction and sampling, were evaluated in method development. The derivatization was performed at 60 °C for 30 min, followed by 15 min sampling at 10 °C, in three cycles (SPME Arrow) or six cycles (SPME). The sensitivity was satisfactory for the method's purposes (LOD-LOQ at 11-36 ng L -1 , and 8-26 ng L -1 , for SPME and SPME Arrow, respectively). The method's linearity ranges from the lower LOQ at trace level (ng L -1 ) to the upper LOQ at 40 mg L -1 . The precision range was 5.7-10.2% and 4.8-9.6% and the accuracy was 97.4% and 96.3% for SPME and SPME Arrow, respectively. The cooling MSPME set-up applied to real commercial goods provided results of quality comparable to previously published data.

Peripheral nerve injury is one of the most debilitating pathologies that severely impair patients' life. Although many efforts have been made to advance in the treatment of such a complex disorder, successful strategies to ensure full recovery are still scarce. The aim of the present work was to develop flexible and mechanically resistant platforms intended to act as a support and guide for neural cells during the regeneration process of peripheral nerve injury. For this purpose, poly(lactic- co -glycolic acid) (PLGA)/poly(d,l-lactic acid) (PDLLA)/poly(ethylene glycol) 400 (PEG)-multichannel-based scaffolds (MCs) were prepared through a multistep process involving electrospun microfibers coated with a polymer blend solution and used as a sacrificial mold. In particular, scaffolds characterized by random (MCR) and aligned (MCA) multichannel were obtained. A design of experiments approach (DoE) was employed to identify a scaffold-optimized composition. MCs were characterized for morphological and mechanical properties, suturability, degradability, cell colonization, and in vivo safety. A new biodegradable, biocompatible, and safe microscale multichannel scaffold was developed as the result of an easy multistep procedure.

Clove oil (CO) is a powerful antioxidant essential oil (EO) with anti-inflammatory, anesthetic, and anti-infective properties. It can be therefore considered a good candidate for wound-healing applications, especially for chronic or diabetic wounds or burns, where the balance of reactive oxygen species (ROS) production and detoxification is altered. However, EOs require suitable formulations to be efficiently administered in moist wound environments. Chitosan hydrophobically modified by an ionic interaction with oleic acid (chitosan oleate, CSO) was used in the present work to stabilize CO nanoemulsions (NEs). The dimensions of the NE were maintained at around 300 nm as the volume distribution for up to six months, and the CO content did not decrease to under 80% over 4 months, confirming the good stabilizing properties of CSO. The antioxidant properties of the CO NE were evaluated in vitro by a 2,2-diphenil-2-picrylhydrazyl hydrate (DPPH) assay, and in fibroblast cell lines by electron paramagnetic resonance (EPR) using α -phenyl- N -tert-butyl nitrone (PBN) as a spin trap; a protective effect was obtained comparable to that obtained with α -tocopherol treatment. In a murine burn model, the ability of CO formulations to favor macroscopic wound closure was evidenced, and a histological analysis revealed a positive effect of the CO NE on the reparation of the lesion after 18 days. Samples of wounds at 7 days were subjected to a histological analysis and parallel dosage of lipid peroxidation by means of a thiobarbituric acid-reactive substances (TBARS) assay, confirming the antioxidant and anti-inflammatory activity of the CO NE.

In April 2015, a fishing boat that departed from Libya with about 1,000 migrants on board sank in the Mediterranean Sea. Most of the migrants were packed in the hull of the boat and drowned in the shipwreck. After fifteen months, the ship was recovered from the seabed and brought to a Sicilian naval area for forensic investigations. Skeletal remains belonging to more than 700 people were retrieved. A selected sample composed of 80 victims was considered in order to evaluate the possibility of achieving genetic profiles useful for a positive identification from these challenging specimens. The molecular features of the DNA recovered from a significant number of real casework samples exposed to seawater for long periods of time were described for the first time. Three different DNA extraction protocols and three different commercial kits were employed in order to generate genetic profiles based on the characterization of 21 autosomal STR loci. The combination of multiple DNA extractions and the cross-checking of multiple PCR amplifications with different kits allowed to obtain reliable genetic profiles characterized by at least 16 STR markers in more than 70% of the samples. The factors that could have affected the different quality of the genetic profiles were investigated and the bone preservation was examined through microscopic and macroscopic analyses. The approach presented in this study could be useful in the management of the genetic analysis of bone samples collected in other similar DVI scenarios. The genetic profiles recovered from the bone samples will be compared in kinship analysis to putative relatives of the victims collected in Africa in order to obtain positive identifications., Competing Interests: Conflict of interest The authors have declared no conflict of interest and that the study was performed with the authorization of the Italian Government’s Office of the Commissioner for Missing Persons (UCPS-Ufficio Commissario Straordinario per le Persone Scomparse) and of the judicial authority., (Copyright © 2022 Elsevier B.V. All rights reserved.)

The development of a successful strategy to ensure a full recovery in patients affected by peripheral nerve injury (PNI), one of the most debilitating pathologies, is, still today, a major clinical challenge. Herein, spermidine (SP), an endogenous polyamine, is employed with a dual role: as cross-linking agent for alginate (ALG) and as antioxidant and anti-inflammatory compound. In particular, micro/nanogels based on the ionic interaction between ALG and SP were obtained via ionotropic gelation. Different ALG concentrations and viscosity grades and different SP concentrations were considered. The influence of such variables on micro/nanogels size was investigated by means of a Design of Experiments (DoE) approach (full factorial design). The formation of micro/nanogels was proved by Scanning Electron Microscope (SEM) analysis and by rheological and profilometry measurements. Fourier Transform Infrared (FTIR) measurements performed on nanogels of optimal composition confirmed SP-ALG interaction. The addition of trehalose as cryoprotectant agent to nanogel dispersion was considered in view of the employment of freeze-drying process to obtain a stable product. Moreover, in vitro studies on Schwann cells proved the ability of SP of expressing antioxidant and anti-inflammatory properties, even if involved in the formation of nanogels., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2022 Elsevier B.V. All rights reserved.)

Indocyanine green (ICG) is a safe dye widely used in the biomedical field. Its photodynamic effect (PDT), originating from laser irradiation at 803 nm, opens interesting perspectives in theranostic applications. To overcome its low water stability, ICG can be shielded with nanoparticles (NPs). In this work, previously developed NPs based on poly lactic-co-glycolic acid (PLGA) coated with chitosan oleate (CS-OA) and loaded with resveratrol as a hydrophobic model drug have been proposed as an ICG carrier. These systems have been selected for their observed immunostimulatory properties. The possible loading of the dye by adsorption onto NP surface by electrostatic interaction was studied here in comparison with the encapsulation into the PLGA core. The ICG-chitosan (CS) interaction has been characterized by spectrophotometry, spectroscopy and in-cell in vitro assays. Fluorescence quenching was observed due to the ionic interaction between ICG and CS and was studied considering the dye:polymer stoichiometry and the effect of the NP dilution in cell culture medium (DMEM). The NP systems have been compared in vitro, assessing their behaviour in Caco-2 cell lines. A reduction in cell viability was observed after irradiation of ICG associated with NPs, evident also for the samples loaded by adsorption. These findings open the opportunity to exploit the association of PDT's effect on ICG with the properties of CS-OA coated NPs, whose immunostimulatory effect can be associated with PDT mechanism in cancer therapy.

Formaldehyde (FA) is ubiquitous in the atmospheric environment. It is generally the dominant atmospheric carbonyl compound. Due to its well-known carcinogenicity, FA is a compound that arises the attention in the scientific community. In studies concerning the toxicological effects of FA on humans, animals, and the environment, testing and calibration of air sampling systems and analytical instruments are pivotal. Therefore, the preparation of controllable standard gaseous atmospheres containing FA at levels known with precision and accuracy is essential. This review summarizes the procedures for generating the FA atmosphere, given that operative solutions have been evolving recently. Furthermore, an overview on the available system to collect and store gaseous standard is reported. The progressively implemented FA generation techniques, together with commercially-available instruments, are herein described, classified, and compared.

Producing mesenchymal stem cell (MSC)-secretome for dose escalation studies and clinical practice requires scalable and good manufacturing practice (GMP)-compliant production procedures and formulation into a standardized medicinal product. Starting from a method that combines ultrafiltration and freeze-drying to transform MSC-secretome into a pharmaceutical product, the lyosecretome, this work aims to: (i) optimize the lyosecretome formulation; (ii) investigate sources of variability that can affect the robustness of the manufacturing process; (iii) modify the ultrafiltration step to obtain a more standardized final product. Design of experiments and principal component analysis of the data were used to study the influence of batch production, lyophilization, mannitol (M)/sucrose (S) binary mixture, selected as cryoprotectant excipients, and the total amount of excipients on the extracellular vesicles (EV) particle size, the protein and lipid content and the in vitro anti-elastase. The different excipients ratios did not affect residual moisture or EV particle size; simultaneously, proteins and lipids were better preserved in the freeze-dried product using the maximum total concentration of excipients (1.5% w / v ) with a M:S ratio of about 60% w / w . The anti-elastase activity was instead better preserved using 0.5% w / w of M as excipient. The secretome batch showed to be the primary source of variability; therefore, the manufacturing process has been modified and then validated: the final product is now concentrated to reach a specific protein (and lipid) concentration instead of cell equivalent concentration. The new standardization approach led to a final product with more reproducible quali-quantitative composition and higher biological activity.