Marco Roman

Active Silver Nanoparticles for Wound Healing

In this preliminary study, the silver nanoparticle (Ag NP)-based dressing, Acticoat™ Flex 3, has been applied to a 3D fibroblast cell culture in vitro and to a real partial thickness burn patient. The in vitro results show that Ag NPs greatly reduce mitochondrial activity, while cellular staining techniques show that nuclear integrity is maintained, with no signs of cell death. For the first time, transmission electron microscopy (TEM) and inductively coupled plasma mass spectrometry (ICP-MS) analyses were carried out on skin biopsies taken from a single patient during treatment. The results show that Ag NPs are released as aggregates and are localized in the cytoplasm of fibroblasts. No signs of cell death were observed, and the nanoparticles had different distributions within the cells of the upper and lower dermis. Depth profiles of the Ag concentrations were determined along the skin biopsies. In the healed sample, most of the silver remained in the surface layers, whereas in the unhealed sample, the silver penetrated more deeply. The Ag concentrations in the cell cultures were also determined. Clinical observations and experimental data collected here are consistent with previously published articles and support the safety of Ag NP-based dressing in wound treatment.

Characterization and evaluation of silver release from four different dressings used in burns care

For centuries silver and silver compounds have been in use to control infection and avoid septicaemia in the care of burns and chronic wounds. Renewed interest has resulted in a number of Ag based dressings that are now widely used in burns centres. Despite extensive use, a systematic study of the chemical composition, release kinetics and biochemical action of these products has yet to be published. In this work we have characterized the morphology of four commercial Ag dressings by scanning electron microscopy and the silver content was determined to range between 1.39 mg/cm(2) and 0.03 mg/cm(2). Release kinetics in three different matrices (ultra pure water, normal saline solution and a human serum substitute) were determined. The highest rates were found in serum substitute, with a maximum of 4099 μg/(hcm(2)) to a minimum of 0.0001 μg/(hcm(2)). Our results show that the mean inhibitory concentrations are exceeded for most common pathogens in serum substitute and sterile water, but the presence of high Cl(-) concentrations tend to inactivate the dressings.

Hydrodynamic chromatography coupled to single-particle ICP-MS for the simultaneous characterization of AgNPs and determination of dissolved Ag in plasma and blood of burn patients

AbstractSilver nanoparticles (AgNPs) are increasingly used in medical devices as innovative antibacterial agents, but no data are currently available on their chemical transformations and fate in vivo in the human body, particularly on their potential to reach the circulatory system. To study the processes involving AgNPs in human plasma and blood, we developed an analytical method based on hydrodynamic chromatography (HDC) coupled to inductively coupled plasma mass spectrometry (ICP-MS) in single-particle detection mode. An innovative algorithm was implemented to deconvolute the signals of dissolved Ag and AgNPs and to extrapolate a multiparametric characterization of the particles in the same chromatogram. From a single injection, the method provides the concentration of dissolved Ag and the distribution of AgNPs in terms of hydrodynamic diameter, mass-derived diameter, number and mass concentration. This analytical approach is robust and suitable to study quantitatively the dynamics and kinetics of AgNPs in complex biological fluids, including processes such as agglomeration, dissolution and formation of protein coronas. The method was applied to study the transformations of AgNP standards and an AgNP-coated dressing in human plasma, supported by micro X-ray fluorescence (μXRF) and micro X-ray absorption near-edge spectroscopy (μXANES) speciation analysis and imaging, and to investigate, for the first time, the possible presence of AgNPs in the blood of three burn patients treated with the same dressing. Together with our previous studies, the results strongly support the hypothesis that the systemic mobilization of the metal after topical administration of AgNPs is driven by their dissolution in situ. Graphical AbstractSimplified scheme of the combined analytical approach adopted for studying the chemical dynamics of AgNPs in human plasma/blood

Development and application of methods for the determination of silver in polymeric dressings used for the care of burns

Open vessel and microwave digestion methods have been developed for the determination of total silver in six commercial dressing used for the treatment of skin burns. An extraction method using TMAH has also been developed to determine the amount of silver present in the exudates found on the surface after dressing removal so an estimation of the patient dose can be made. All microwave methods had a quantitative recovery, whereas the open vessel had recoveries that ranged from 80 to 100%. The silver concentrations were determined by inductively coupled plasma mass spectrometry using an external calibration. In the absence of suitable reference materials, isotope dilution analysis was applied to validate the accuracy of results obtained by external calibration. All the products had a total Ag content that agreed with the values declared by the producer, which ranged from 10 to 0.2% Ag by weight. One of the methods was applied to the indirect determination of Ag released in vivo by Acticoat™ Flex 3, a dressing composed of silver nanoparticles on a polymer net. Silver levels were determined in used dressings after application to patients with partial thickness skin burns. A maximum of 62% of the silver was found to have been released onto the patient where hemopurulent exudate occurred, indicating that the dressing was virtually exhausted after 3 days of use. We conclude that the Ag released into the patients tissues is closely correlated with the local severity of the wound.

Selenium speciation in rat colon tissues

A study of selenium (Se) speciation in rat colon tissues is presented. Four different procedures for the extraction of Se compounds were evaluated in terms of recovery and species preservation. Total Se in tissue and extracts was determined by ICP-MS and isotope dilution analysis. The selected and optimized protocol allowed an extraction of 43% of Se, while continuously bubbling nitrogen in the solution during the procedure was mandatory to prevent the oxidative degradation of selenoproteins. Speciation analysis was then performed on the extracts using size exclusion- and anion exchange-HPLC for species separation. A number of Se compounds were detected in rat colon extracts, and individually quantified by coupling HPLC-ICP-MS and species-unspecific on-line (post-column) isotope dilution analysis. Among the isolated selenospecies, the two major proteins glutathione peroxidase type 2 and thioredoxin reductase type 1 have been potentially identified by their molecular weight using MALDI-TOF-MS.

Feedback mechanisms between snow and atmospheric mercury: Results and observations from field campaigns on the Antarctic plateau

The Antarctic Plateau snowpack is an important environment for the mercury geochemical cycle. We have extensively characterized and compared the changes in surface snow and atmospheric mercury concentrations that occur at Dome C. Three summer sampling campaigns were conducted between 2013 and 2016. The three campaigns had different meteorological conditions that significantly affected mercury deposition processes and its abundance in surface snow. In the absence of snow deposition events, the surface mercury concentration remained stable with narrow oscillations, while an increase in precipitation results in a higher mercury variability. The Hg concentrations detected confirm that snowfall can act as a mercury atmospheric scavenger. A high temporal resolution sampling experiment showed that surface concentration changes are connected with the diurnal solar radiation cycle. Mercury in surface snow is highly dynamic and it could decrease by up to 90% within 4/6 h. A negative relationship between surface snow mercury and atmospheric concentrations has been detected suggesting a mutual dynamic exchange between these two environments. Mercury concentrations were also compared with the Br concentrations in surface and deeper snow, results suggest that Br could have an active role in Hg deposition, particularly when air masses are from coastal areas. This research presents new information on the presence of Hg in surface and deeper snow layers, improving our understanding of atmospheric Hg deposition to the snow surface and the possible role of re-emission on the atmospheric Hg concentration.

Electrochemical preparation of standard solutions of Pb(II) ions in ionic liquid for analysis of hydrophobic samples: The olive oil case

In this paper we present an electrochemical approach to prepare standard solutions of metal ions in a room temperature ionic liquid (IL), which can find useful application for analysis in hydrophobic matrices. The method, developed here for the case of lead ions, is based on the galvanostatic dissolution of a lead anode dipped directly in a suitable IL, namely tri-hexyl(tetradecyl)phosphonium bis (trifluoromethylsulfonyl) imide ([P14,6,6,6]+[NTf2]-). After each oxidation step, the metal dissolution process in the IL solutions was monitored by cyclic voltammetric measurements at a glassy carbon disk electrode. The results indicated that the peak current relevant to the reduction of the electro-generated Pb(II) increased linearly while increasing the oxidation time. By varying the oxidation time from 200 to 6000s, a set of Pb(II)/[P14,6,6,6]+[NTf2]- solutions at concentrations ranging between 10 and 300μgg-1 was prepared. To validate the efficiency of the electrochemical procedure to produce metal ion standard solutions, the Pb content was quantified by developing a microwave digestion procedure specifically suitable for the IL medium, followed by ICP-QMS analysis in the digested standards. The results indicated a satisfactory agreement between concentrations found by ICP-QMS and calculated from electrochemical data, with a coulometric efficiency of Pb(II) generation in ionic liquid ≥95.6%. Finally, the applicability of the Pb(II)/IL solutions as standards for analyses in hydrophobic media was tested by determining, by ICP-QMS, the Pb content in an extra-virgin olive oil spiked with known amounts of a Pb(II)/IL standard. Satisfactory Pb recoveries, ≥96%, were measured.