Publications by authors named "Stefania Rapino"

36 Publications

A Plant Bioreactor for the Synthesis of Carbon Nanotube Bionic Nanocomposites.

Front Bioeng Biotechnol 2020 5;8:560349. Epub 2020 Nov 5.

Dipartimento di Chimica "Giacomo Ciamician," Alma mater Studiorum-Università di Bologna, Bologna, Italy.

Bionic composites are an emerging class of materials produced exploiting living organisms as reactors to include synthetic functional materials in their native and highly performing structures. In this work, single wall carboxylated carbon nanotubes (SWCNT-COOH) were incorporated within the roots of living plants of . This biogenic synthetic route produced a bionic composite material made of root components and SWCNT-COOH. The synthesis was possible exploiting the transport processes existing in the plant roots. Scanning electrochemical microscopy (SECM) measurements showed that SWCNT-COOH entered the vascular bundles of roots localizing within xylem vessels. SWCNT-COOH preserved their electrical properties when embedded inside the root matrix, both at a microscopic level and a macroscopic level, and did not significantly affect the mechanical properties of roots.
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http://dx.doi.org/10.3389/fbioe.2020.560349DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7676904PMC
November 2020

Insights into the mechanism of coreactant electrochemiluminescence facilitating enhanced bioanalytical performance.

Nat Commun 2020 05 29;11(1):2668. Epub 2020 May 29.

Department of Chemistry Giacomo Ciamician, University of Bologna, via Selmi 2, 40126, Bologna, Italy.

Electrochemiluminescence (ECL) is a powerful transduction technique with a leading role in the biosensing field due to its high sensitivity and low background signal. Although the intrinsic analytical strength of ECL depends critically on the overall efficiency of the mechanisms of its generation, studies aimed at enhancing the ECL signal have mostly focused on the investigation of materials, either luminophores or coreactants, while fundamental mechanistic studies are relatively scarce. Here, we discover an unexpected but highly efficient mechanistic path for ECL generation close to the electrode surface (signal enhancement, 128%) using an innovative combination of ECL imaging techniques and electrochemical mapping of radical generation. Our findings, which are also supported by quantum chemical calculations and spin trapping methods, led to the identification of a family of alternative branched amine coreactants, which raises the analytical strength of ECL well beyond that of present state-of-the-art immunoassays, thus creating potential ECL applications in ultrasensitive bioanalysis.
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http://dx.doi.org/10.1038/s41467-020-16476-2DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7260178PMC
May 2020

Light-Triggered Electron Transfer between a Conjugated Polymer and Cytochrome C for Optical Modulation of Redox Signaling.

iScience 2020 May 22;23(5):101091. Epub 2020 Apr 22.

Center for Nano Science and [email protected], Istituto Italiano di Tecnologia, via Pascoli 70/3, 20133 Milano, Italy. Electronic address:

Protein reduction/oxidation processes trigger and finely regulate a myriad of physiological and pathological cellular functions. Many biochemical and biophysical stimuli have been recently explored to precisely and effectively modulate intracellular redox signaling, due to the considerable therapeutic potential. Here, we propose a first step toward an approach based on visible light excitation of a thiophene-based semiconducting polymer (P3HT), demonstrating the realization of a hybrid interface with the Cytochrome c protein (CytC), in an extracellular environment. By means of scanning electrochemical microscopy and spectro-electrochemistry measurements, we demonstrate that, upon optical stimulation, a functional interaction between P3HT and CytC is established. Polymer optical excitation locally triggers photoelectrochemical reactions, leading to modulation of CytC redox activity, either through an intermediate step, involving reactive oxygen species formation, or via a direct photoreduction process. Both processes are triggered by light, thus allowing excellent spatiotemporal resolution, paving the way to precise modulation of protein redox signaling.
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http://dx.doi.org/10.1016/j.isci.2020.101091DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7240120PMC
May 2020

Structural and electrochemical characterization of lawsone-dependent production of tellurium-metal nanoprecipitates by photosynthetic cells of Rhodobacter capsulatus.

Bioelectrochemistry 2020 Jun 8;133:107456. Epub 2020 Jan 8.

Dept. of Pharmacy and Biotechnology FaBiT, University of Bologna, Bologna, Italy. Electronic address:

Cells of the facultative photosynthetic bacterium Rhodobacter capsulatus exploit the simultaneous presence in the cultural medium of the toxic oxyanion tellurite (TeO) and the redox mediator lawsone (2-hydroxy-1,4-naphthoquinone) by reducing tellurite to metal Te nanoprecipitates (TeNPs) outside the cells. Here we have studied the mechanism by which lawsone interacts with metabolically active cells and analysed both structure and composition of the TeNPs collected from the growth medium of phototrophycally grown R. capsulatus. High Resolution Transmission Electron Microscopy (HR-TEM) images and Energy-Dispersive X-ray (EDX) microanalysis of TeNPs showed a central core of polycrystalline tellurium interspersed in an organic matrix with a predominant protein-based composition. The main proteins from Te nanostructures were identified by Liquid Chromatography tandem-Mass Spectrometry and were all correlated with the cell outer membrane composition. The interaction of reduced lawsone with tellurite and with the bacterial cells was probed by Cyclic Voltammetry and Scanning ElectroChemical Microscopy (SECM). We concluded that lawsone is required for the reduction of tellurite to metal Te in a reaction mechanism dependent on reducing equivalents deriving from the cell photosynthetic metabolism. SECM experiments demonstrate that lawsone, by diffusing inside the bacterial cells, is effectively available at the membrane site of the photosynthetic electron transport chain.
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http://dx.doi.org/10.1016/j.bioelechem.2020.107456DOI Listing
June 2020

Specific, Surface-Driven, and High-Affinity Interactions of Fluorescent Hyaluronan with PEGylated Nanomaterials.

ACS Appl Mater Interfaces 2020 Feb 29;12(6):6806-6813. Epub 2020 Jan 29.

Dipartimento di Chimica "Giacomo Ciamician" , Università di Bologna , via Selmi 2 , 40126 Bologna , Italy.

Hybrid nanomaterials are a subject of extensive research in nanomedicine, and their clinical application is reasonably envisaged in the near future. However, the fate of nanomaterials in biological environments poses serious limitations to their application; therefore, schemes to monitor them and gain control on their toxicity could be of great help for the development of the field. Here, we propose a probe for PEGylated nanosurfaces based on hyaluronic acid (HA) functionalized with rhodamine B (RB). We show that the high-affinity interaction of this fluorogenic hyaluronan (HA-RB) with nanoparticles exposing PEGylated surfaces results in their sensing, labeling for super-resolution imaging, and synergistic cellular internalization. HA-RB forms nanogels that interact with high affinity-down to the picomolar range-with silica nanoparticles, selectively when their surface is covered by a soft and amphiphilic layer. This surface-driven interaction triggers the enhancement of the luminescence intensity of the dyes, otherwise self-quenched in HA-RB nanogels. The sensitive labeling of specific nanosurfaces also allowed us to obtain their super-resolution imaging via binding-activated localization microscopy (BALM). Finally, we show how this high-affinity interaction activates a synergistic cellular uptake of silica nanoparticles and HA-RB nanogels, followed by a differential fate of the two partner nanomaterials inside cells.
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http://dx.doi.org/10.1021/acsami.9b17974DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7993635PMC
February 2020

Oxygen Redox Reaction in Ionic Liquid and Ionic Liquid-like Based Electrolytes: A Scanning Electrochemical Microscopy Study.

J Phys Chem Lett 2019 Jun 4;10(12):3333-3338. Epub 2019 Jun 4.

Department of Chemistry Giacomo Ciamician , Alma Mater Studiorum Bologna University , 40126 Bologna , Italy.

Improving the stability of the cathode interface is one of the critical issues for the development of high-performance Li/O batteries. The most critical feature to address is the development of electrolytes that mitigate side reactions that bring about cathode passivation. It is well-known that the superoxide anion (O) plays a critical role. Here, we propose scanning electrochemical microscopy (SECM) as an analytical tool to screen the electrolyte of Li/O batteries. We demonstrate that by using SECM it is possible to evaluate the stability of O and of the cathode to the passivation process occurring during the oxygen redox reaction. Specifically, we report a study carried out at a glassy carbon electrode in 1-butyl-1-methylpyrrolidinium bis(trifluoromethanesulfonyl)imide (PYRTFSI) and lithium bis(trifluoromethanesulfonyl)imide (LiTFSI) and in tetraethylene glycol dimethyl ether with LiTFSI, the latter ranging from the salt-in-solvent to solvent-in-salt regions.
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http://dx.doi.org/10.1021/acs.jpclett.9b00774DOI Listing
June 2019

Electrochemical monitoring of reactive oxygen/nitrogen species and redox balance in living cells.

Anal Bioanal Chem 2019 Jul 23;411(19):4365-4374. Epub 2019 Apr 23.

Department of Chemistry "Giacomo Ciamician", University of Bologna, Via F. Selmi 2, 40126, Bologna, Italy.

Levels of reactive oxygen species (ROS) and reactive nitrogen species (RNS) in cells and cell redox balance are of great interest in live cells as they are correlated to several pathological and physiological conditions of living cells. ROS and RNS detection is limited due to their spatially restricted abundance: they are usually located in sub-cellular areas (e.g., in specific organelles) at low concentration. In this work, we will review and highlight the electrochemical approach to this bio-analytical issue. Combining electrochemical methods and miniaturization strategies, specific, highly sensitive, time, and spatially resolved measurements of cellular oxidative stress and redox balance analysis are possible. Graphical abstract In this work, we highlight and review the use of electrochemistry for the highly spatial and temporal resolved detection of ROS/RNS levels and of redox balance in living cells. These levels are central in several pathological and physiological conditions and the electrochemical approach is a vibrant bio-analytical trend in this field.
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http://dx.doi.org/10.1007/s00216-019-01734-0DOI Listing
July 2019

Stable and Biocompatible Monodispersion of C in Water by Peptides.

Bioconjug Chem 2019 03 24;30(3):808-814. Epub 2019 Jan 24.

Dipartimento di Chimica "Giacomo Ciamician" , Alma Mater Studiorum - Università di Bologna , Via Francesco Selmi, 2 - 40126 Bologna , Italy.

The lack of solubility in water and the formation of aggregates hamper many opportunities for technological exploitation of C. Here, different peptides were designed and synthesized with the aim of monomolecular dispersion of C in water. Phenylalanines were used as recognizing moieties, able to interact with C through π-π stacking, while a varying number of glycines were used as spacers, to connect the two terminal phenylalanines. The best performance in the dispersion of C was obtained with the FGGGF peptidic nanotweezer at a pH of 12. A full characterization of this adduct was carried out. The peptides disperse C in water with high efficiency, and the solutions are stable for months both in pure water and in physiological environments. NMR measurements demonstrated the ability of the peptides to interact with C. AFM measurements showed that C is monodispersed. Electrospray ionization mass spectrometry determined a stoichiometry of [email protected](FGGGF). Molecular dynamics simulations showed that the peptides assemble around the C cage, like a candy in its paper wrapper, creating a supramolecular host able to accept C in the cavity. The peptide-wrapped C is fully biocompatible and the C "dark toxicity" is eliminated. [email protected](FGGGF) shows visible light-induced reactive oxygen species (ROS) generation at physiological saline concentrations and reduction of the HeLa cell viability in response to visible light irradiation.
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http://dx.doi.org/10.1021/acs.bioconjchem.8b00916DOI Listing
March 2019

Surface-Confined Electrochemiluminescence Microscopy of Cell Membranes.

J Am Chem Soc 2018 11 29;140(44):14753-14760. Epub 2018 Oct 29.

University of Bordeaux , Bordeaux INP, ISM, UMR CNRS 5255 , 33607 Pessac , France.

Herein is reported a surface-confined microscopy based on electrochemiluminescence (ECL) that allows to image the plasma membrane of single cells at the interface with an electrode. By analyzing photoluminescence (PL), ECL and AFM images of mammalian CHO cells, we demonstrate that, in contrast to the wide-field fluorescence, ECL emission is confined to the immediate vicinity of the electrode surface and only the basal membrane of the cell becomes luminescent. The resulting ECL microscopy reveals details that are not resolved by classic fluorescence microscopy, without any light irradiation and specific setup. The thickness of the ECL-emitting regions is ∼500 nm due to the unique ECL mechanism that involves short-lifetime electrogenerated radicals. In addition, the reported ECL microscopy is a dynamic technique that reflects the transport properties through the cell membranes and not only the specific labeling of the membranes. Finally, disposable transparent carbon nanotube (CNT)-based electrodes inkjet-printed on classic microscope glass coverslips were used to image cells in both reflection and transmission configurations. Therefore, our approach opens new avenues for ECL as a surface-confined microscopy to develop single cell assays and to image the dynamics of biological entities in cells or in membranes.
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http://dx.doi.org/10.1021/jacs.8b08080DOI Listing
November 2018

Enhancing radiosensitivity of melanoma cells through very high dose rate pulses released by a plasma focus device.

PLoS One 2018 29;13(6):e0199312. Epub 2018 Jun 29.

University of Bologna, Department of Physics and Astronomy, Bologna, Italy.

Radiation therapy is a useful and standard tumor treatment strategy. Despite recent advances in delivery of ionizing radiation, survival rates for some cancer patients are still low because of recurrence and radioresistance. This is why many novel approaches have been explored to improve radiotherapy outcome. Some strategies are focused on enhancement of accuracy in ionizing radiation delivery and on the generation of greater radiation beams, for example with a higher dose rate. In the present study we proposed an in vitro research of the biological effects of very high dose rate beam on SK-Mel28 and A375, two radioresistant human melanoma cell lines. The beam was delivered by a pulsed plasma device, a "Mather type" Plasma Focus for medical applications. We hypothesized that this pulsed X-rays generator is significantly more effective to impair melanoma cells survival compared to conventional X-ray tube. Very high dose rate treatments were able to reduce clonogenic efficiency of SK-Mel28 and A375 more than the X-ray tube and to induce a greater, less easy-to-repair DNA double-strand breaks. Very little is known about biological consequences of such dose rate. Our characterization is preliminary but is the first step toward future clinical considerations.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0199312PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6025851PMC
April 2019

Electrochemical imaging of cells and tissues.

Chem Sci 2018 May 9;9(20):4546-4554. Epub 2018 Apr 9.

Laboratory of Physical and Analytical Electrochemistry (LEPA) , École Polytechnique Fédéderale de Lausanne , EPFL Valais Valais , Rue de l'Industrie 17 , CP 440 , 1951 Sion , Switzerland . Email:

The technological and experimental progress in electrochemical imaging of biological specimens is discussed with a view on potential applications for skin cancer diagnostics, reproductive medicine and microbial testing. The electrochemical analysis of single cell activity inside cell cultures, 3D cellular aggregates and microtissues is based on the selective detection of electroactive species involved in biological functions. Electrochemical imaging strategies, based on nano/micrometric probes scanning over the sample and sensor array chips, respectively, can be made sensitive and selective without being affected by optical interference as many other microscopy techniques. The recent developments in microfabrication, electronics and cell culturing/tissue engineering have evolved in affordable and fast-sampling electrochemical imaging platforms. We believe that the topics discussed herein demonstrate the applicability of electrochemical imaging devices in many areas related to cellular functions.
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http://dx.doi.org/10.1039/c8sc01035hDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5969511PMC
May 2018

Proteins as supramolecular hosts for C: a true solution of C in water.

Nanoscale 2018 May;10(21):9908-9916

Dipartimento di Chimica "G. Ciamician", Alma Mater Studiorum - Università di Bologna, via F. Selmi 2, 40126 Bologna, Italy.

Hybrid systems have great potential for a wide range of applications in chemistry, physics and materials science. Conjugation of a biosystem to a molecular material can tune the properties of the components or give rise to new properties. As a workhorse, here we take a [email protected] hybrid. We show that lysozyme recognizes and disperses fullerene in water. AFM, cryo-TEM and high resolution X-ray powder diffraction show that the C60 dispersion is monomolecular. The adduct is biocompatible, stable in physiological and technologically-relevant environments, and easy to store. Hybridization with lysozyme preserves the electrochemical properties of C60. EPR spin-trapping experiments show that the [email protected] hybrid produces ROS following both type I and type II mechanisms. Due to the shielding effect of proteins, the adduct generates significant amounts of 1O2 also in aqueous solution. In the case of type I mechanism, the protein residues provide electrons and the hybrid does not require addition of external electron donors. The preparation process and the properties of [email protected]yme are general and can be expected to be similar to other [email protected] systems. It is envisaged that the properties of the [email protected] hybrids will pave the way for a host of applications in nanomedicine, nanotechnology, and photocatalysis.
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http://dx.doi.org/10.1039/c8nr02220hDOI Listing
May 2018

Functional Biocompatible Matrices from Mussel Byssus Waste.

ACS Biomater Sci Eng 2018 Jan 14;4(1):57-65. Epub 2017 Dec 14.

Dipartimento di Chimica "Giacomo Ciamician", Alma Mater Studiorum Università di Bologna, via Selmi 2, 40126 Bologna, Italy.

The mussel byssus is a biorenewable, protein-based material produced by marine mussels, which has attracted the interest of material scientists because of its remarkable mechanical and self-healing properties. Large quantities of byssus waste material from mussel mariculture are produced every year, which have great potential as a raw starting material for producing sustainable advanced materials. In this work, we developed a facile and scalable method to synthesize whole byssus-based porous matrices that retain part of the hierarchical organization of the pristine material at the nanoscale. The resulting material is biocompatible and maintains important native byssus features: metal ion chelation (≥12 mg/g), collagen domains, and hierarchical organization, with tunable properties controlled via metal ion content. Furthermore, these biocompatible matrices showed a dye absorbing efficiency (up to 64 mg/g for anionic dyes) that was similar to or higher than that of the pristine byssus, a proof of preservation of structural motifs. These findings indicate that biorenewable matrices originating from byssus waste could have potential use in biomedical engineering and applied material science.
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http://dx.doi.org/10.1021/acsbiomaterials.7b00743DOI Listing
January 2018

Single Cell Electrochemiluminescence Imaging: From the Proof-of-Concept to Disposable Device-Based Analysis.

J Am Chem Soc 2017 11 8;139(46):16830-16837. Epub 2017 Nov 8.

University of Bordeaux, Bordeaux INP, ISM, UMR CNRS 5255, 33607 Pessac, France.

We report here the development of coreactant-based electrogenerated chemiluminescence (ECL) as a surface-confined microscopy to image single cells and their membrane proteins. Labeling the entire cell membrane allows one to demonstrate that, by contrast with fluorescence, ECL emission is only detected from fluorophores located in the immediate vicinity of the electrode surface (i.e., 1-2 μm). Then, to present the potential diagnostic applications of our approach, we selected carbon nanotubes (CNT)-based inkjet-printed disposable electrodes for the direct ECL imaging of a labeled plasma receptor overexpressed on tumor cells. The ECL fluorophore was linked to an antibody and enabled to localize the ECL generation on the cancer cell membrane in close proximity to the electrode surface. Such a result is intrinsically associated with the unique ECL mechanism and is rationalized by considering the limited lifetimes of the electrogenerated coreactant radicals. The electrochemical stimulus used for luminescence generation does not suffer from background signals, such as the typical autofluorescence of biological samples. The presented surface-confined ECL microscopy should find promising applications in ultrasensitive single cell imaging assays.
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http://dx.doi.org/10.1021/jacs.7b09260DOI Listing
November 2017

Glucose and Lactate Miniaturized Biosensors for SECM-Based High-Spatial Resolution Analysis: A Comparative Study.

ACS Sens 2017 Sep 24;2(9):1310-1318. Epub 2017 Aug 24.

Chemistry Department "Giacomo Ciamician", University of Bologna , Via Selmi 2, 40126 Bologna, Italy.

With the aim of developing miniaturized enzymatic biosensors suitable for in vitro diagnostic applications, such as monitoring of metabolites at single cell level, glucose and lactate biosensors were fabricated by immobilizing enzymes (glucose oxidase and lactate oxidase, respectively) on 10 μm Pt ultramicroelectrodes. These electrodes are meant to be employed as probes for scanning electrochemical microscopy (SECM), which is a unique technique for high-spatial-resolution electrochemical-based analysis. The use of enzymatic moieties improves sensitivity, time scale response, and information content of the microprobes; however, protein immobilization is a key step in the biosensor preparation that greatly affects the overall performance. A crucial aspect is the miniaturization of the sensing, preserving their sensitivity. In this work, we investigated the most common enzyme immobilization techniques. Several fabrication routes are reported and the main figures of merit, such as sensitivity, detection limit, response time, reproducibility, spatial resolution, biosensor efficiency, permeability, selectivity, and the ability to block electro-active interfering species, are investigated and compared. With the intent of using the microprobes for in vitro functional imaging of single living cells, we carefully evaluate the spatial resolution achieved by our modified electrodes on 2D SECM imaging. Metabolic activity of single MCF10A cells were obtained by monitoring the glucose concentrations in close proximity of single living cell, using the UME-based biosensor probes prepared. A voltage-switch approach was implemented to disentangle the topographical contribution of the cells enabling quantitative measurements of cellular uptakes.
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http://dx.doi.org/10.1021/acssensors.7b00324DOI Listing
September 2017

Co-axial heterostructures integrating palladium/titanium dioxide with carbon nanotubes for efficient electrocatalytic hydrogen evolution.

Nat Commun 2016 12 12;7:13549. Epub 2016 Dec 12.

Department of Chemistry 'Giacomo Ciamician', University of Bologna and INSTM, via Selmi 2, Bologna 40126, Italy.

Considering the depletion of fossil-fuel reserves and their negative environmental impact, new energy schemes must point towards alternative ecological processes. Efficient hydrogen evolution from water is one promising route towards a renewable energy economy and sustainable development. Here we show a tridimensional electrocatalytic interface, featuring a hierarchical, co-axial arrangement of a palladium/titanium dioxide layer on functionalized multi-walled carbon nanotubes. The resulting morphology leads to a merging of the conductive nanocarbon core with the active inorganic phase. A mechanistic synergy is envisioned by a cascade of catalytic events promoting water dissociation, hydride formation and hydrogen evolution. The nanohybrid exhibits a performance exceeding that of state-of-the-art electrocatalysts (turnover frequency of 15000 H per hour at 50 mV overpotential). The Tafel slope of ∼130 mV per decade points to a rate-determining step comprised of water dissociation and formation of hydride. Comparative activities of the isolated components or their physical mixtures demonstrate that the good performance evolves from the synergistic hierarchical structure.
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http://dx.doi.org/10.1038/ncomms13549DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5159813PMC
December 2016

Complex Media and Enzymatic Kinetics.

Anal Chem 2016 06 19;88(11):5790-6. Epub 2016 May 19.

Dipartimento di Chimica "G. Ciamician", Università di Bologna , V. F. Selmi 2, 40126, Bologna, Italy.

Enzymatic reactions in complex environments often take place with concentrations of enzyme comparable to that of substrate molecules. Two such cases occur when an enzyme is used to detect low concentrations of substrate/analyte or inside a living cell. Such concentrations do not agree with standard in vitro conditions, aimed at satisfying one of the founding hypotheses of the Michaelis-Menten reaction scheme, MM. It would be desirable to generalize the classical approach and show its applicability to complex systems. A permeable micrometrically structured hydrogel matrix was fabricated by protein cross-linking. Glucose oxidase enzyme (GOx) was embedded in the matrix and used as a prototypical system. The concentration of H2O2 was monitored in time and fitted by an accurate solution of the enzymatic kinetic scheme, which is expressed in terms of simple functions. The approach can also find applications in digital microfluidics and in systems biology where the kinetics response in the linear regimes often employed must be replaced.
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http://dx.doi.org/10.1021/acs.analchem.6b00343DOI Listing
June 2016

"Active" drops as phantom models for living cells: a mesoscopic particle-based approach.

Soft Matter 2016 Apr;12(15):3538-44

Dipartimento di Chimica "G. Ciamician", Università di Bologna, V. F. Selmi 2, 40126, Bologna, Italia.

Drops and biological cells share some morphological features and visco-elastic properties. The modelling of drops by mesoscopic non-atomistic models has been carried out to a high degree of success in recent years. We extend such treatment and discuss a simple, drop-like model to describe the interactions of the outer layer of cells with the surfaces of materials. Cells are treated as active mechanical objects that are able to generate adhesion forces. They appear with their true size and are made of "parcels of fluids" or beads. The beads are described by (very) few quantities/parameters related to fundamental chemical forces such as hydrophilicity and lipophilicity that represent an average of the properties of a patch of material or an area of the cell(s) surface. The investigation of adhesion dynamics, motion of individual cells, and the collective behavior of clusters of cells on materials is possible. In the simulations, the drops become active soft matter objects and different from regular droplets they do not fuse when in contact, their trajectories are not Brownian, and they can be forced "to secrete" molecules, to name some of the properties targeted by the modeling. The behavior that emerges from the simulations allows ascribing some cell properties to their mechanics, which are related to their biological features.
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http://dx.doi.org/10.1039/c5sm02686eDOI Listing
April 2016

Calcite Single Crystals as Hosts for Atomic-Scale Entrapment and Slow Release of Drugs.

Adv Healthc Mater 2015 Jul 1;4(10):1510-6. Epub 2015 Jun 1.

Dipartimento di Chimica "Giacomo Ciamician", Alma Mater Studiorum Università di Bologna, via Selmi 2, 40126, Bologna, Italy.

Doxorubicin (DOX)/CaCO3 single crystals act as pH responsive drug carrier. A biomimetic approach demonstrates that calcite single crystals are able, during their growth in the presence of doxorubicin, to entrap drug molecules inside their lattice along specific crystallographic directions. Alterations in lattice dimensions and microstructural parameters are determined by means of high-resolution synchrotron powder diffraction measurements. Confocal microscopy confirms that doxorubicin is uniformly embedded in the crystal and is not simply adsorbed on the crystal surface. A slow release of DOX was obtained preferentially in the proximity of the crystals, targeting cancer cells.
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http://dx.doi.org/10.1002/adhm.201500170DOI Listing
July 2015

A novel neuroferritinopathy mouse model (FTL 498InsTC) shows progressive brain iron dysregulation, morphological signs of early neurodegeneration and motor coordination deficits.

Neurobiol Dis 2015 Sep 4;81:119-33. Epub 2014 Nov 4.

San Raffaele Scientific Institute, Via Olgettina 60, 20132 Milano, Italy; Vita-Salute San Raffaele University, Via Olgettina 58, 20132 Milano, Italy. Electronic address:

Neuroferritinopathy is a rare genetic disease with a dominant autosomal transmission caused by mutations of the ferritin light chain gene (FTL). It belongs to Neurodegeneration with Brain Iron Accumulation, a group of disorders where iron dysregulation is tightly associated with neurodegeneration. We studied the 498-499InsTC mutation which causes the substitution of the last 9 amino acids and an elongation of extra 16 amino acids at the C-terminus of L-ferritin peptide. An analysis with cyclic voltammetry on the purified protein showed that this structural modification severely reduces the ability of the protein to store iron. In order to analyze the impact of the mutation in vivo, we generated mouse models for the some pathogenic human FTL gene in FVB and C57BL/6J strains. Transgenic mice in the FVB background showed high accumulation of the mutated ferritin in brain where it correlated with increased iron deposition with age, as scored by magnetic resonance imaging. Notably, the accumulation of iron-ferritin bodies was accompanied by signs of oxidative damage. In the C57BL/6 background, both the expression of the mutant ferritin and the iron levels were lower than in the FVB strain. Nevertheless, also these mice showed oxidative alterations in the brain. Furthermore, post-natal hippocampal neurons obtained from these mice experienced a marked increased cell death in response to chronic iron overload and/or acute oxidative stress, in comparison to wild-type neurons. Ultrastructural analyses revealed an accumulation of lipofuscin granules associated with iron deposits, particularly enriched in the cerebellum and striatum of our transgenic mice. Finally, experimental subjects were tested throughout development and aging at 2-, 8- and 18-months for behavioral phenotype. Rotarod test revealed a progressive impaired motor coordination building up with age, FTL mutant old mice showing a shorter latency to fall from the apparatus, according to higher accumulation of iron aggregates in the striatum. Our data show that our 498-499InsTC mouse models recapitulate early pathological and clinical traits of the human neuroferritinopathy, thus providing a valuable model for the study of the disease. Finally, we propose a mechanistic model of lipofuscine formation that can account for the etiopathogenesis of human neuroferritinopathy.
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http://dx.doi.org/10.1016/j.nbd.2014.10.023DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4642750PMC
September 2015

Playing peekaboo with graphene oxide: a scanning electrochemical microscopy investigation.

Chem Commun (Camb) 2014 Nov;50(86):13117-20

Dipartimento di Chimica G. Ciamician, Via F. Selmi 2, 40126 Bologna, Italy.

Scanning electrochemical microscopy (SECM) can image graphene oxide (GO) flakes on insulating and conducting substrates. The contrast between GO and the substrate is controlled by the electrostatic interactions that are established between the charges of the molecular redox mediator and the charges present in the sheet/substrate. SECM also allows quantitative measurement - at the nano/microscale - of the charge transfer kinetics between single monolayer sheets and agent molecules.
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http://dx.doi.org/10.1039/c4cc06368fDOI Listing
November 2014

Localization of proteins in paint cross-sections by scanning electrochemical microscopy as an alternative immunochemical detection technique.

Anal Chim Acta 2014 Jun 2;831:31-7. Epub 2014 May 2.

Department of Chemistry "G. Ciamician", University of Bologna, Via Selmi, Bologna 2 40126, Italy. Electronic address:

The qualitative identification of proteinaceous substances, as well as their location within a complex paint stratigraphy, is one of the most challenging issues in the characterization of painting materials. Nevertheless, information on paint components represent a crucial task for studies concerning both the ancient painting techniques adopted and the state of conservation, being fundamental investigations for the selection of appropriate conservation actions. The present research was aimed at developing a new detection approach for the immunochemical localization of ovalbumin in paint cross-sections based on the use of scanning electrochemical microscopy (SECM). The immunochemical analyses were performed using an anti-ovalbumin primary antibody and a secondary antibody labelled with horseradish peroxidase (HRP). SECM measurements were performed in feedback mode using benzoquinone (BQ)/hydroquinone (H2Q) redox couple. In presence of hydrogen peroxide (H2O2), HRP catalyzes the re-oxidation of H2Q to BQ and the increment of BQ concentration in correspondence of the target protein was detected by SECM through the electrochemical reduction of the regenerated BQ at the microelectrode. Indeed, the localization of ovalbumin was possible thanks to a clear discrimination of SECM currents, achieved by the comparison of the measurements recorded before and after H2O2 administration, based on the HRP on/off approach. The method was evaluated both on samples from standard mocks-up and on a historical sample, collected from a Renaissance wood painting. The obtained results were promising, foreseeing a wider application of SECM on cultural heritage researches.
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http://dx.doi.org/10.1016/j.aca.2014.04.058DOI Listing
June 2014

Inhibition of microbial growth by carbon nanotube networks.

Nanoscale 2013 Oct 9;5(19):9023-9. Epub 2013 Aug 9.

Dept. Biology and Biotechnology, Sapienza-University of Rome, Rome, Italy.

In the last years carbon nanotubes have attracted increasing attention for their potential applications in the biomedical field as diagnostic and therapeutic nano tools. Here we investigate the antimicrobial activity of different fully characterized carbon nanotube types (single walled, double walled and multi walled) on representative pathogen species: Gram-positive Staphylococcus aureus, Gram-negative Pseudomonas aeruginosa and the opportunistic fungus Candida albicans. Our results show that all the carbon nanotube types possess a highly significant antimicrobial capacity, even though they have a colony forming unit capacity and induction of oxidative stress in all the microbial species to a different extent. Moreover, scanning electron microscopy analysis revealed that the microbial cells were wrapped or entrapped by carbon nanotube networks. Our data taken together suggest that the reduced capacity of microbial cells to forming colonies and their oxidative response could be related to the cellular stress induced by the interactions of pathogens with the CNT network.
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http://dx.doi.org/10.1039/c3nr02091fDOI Listing
October 2013

Electrochemical detection of H2O2 formation in isolated mitochondria.

Methods Enzymol 2013 ;526:123-34

Department of Experimental Oncology, European Institute of Oncology, Milan, Italy.

Mitochondrial respiration produces both complete and partially reduced oxygen species that are involved in physiological and pathological processes. Indeed, unspecific oxidative damage induced by excessive mitochondrial reactive oxygen species (ROS) plays a role in aging and several diseases, whereas low amounts of ROS act in physiological signaling processes. The exact molecular species, the rate, and the conditions of mitochondrial ROS release are not clearly evaluable by current methods based on oxidation sensitive markers. Recently, electrochemical analysis of biological samples has improved. Following latest methodology, we implemented a novel electrochemical assay for the investigation of aerobic metabolism in isolated mitochondria through simultaneous measurement of O2 consumption and H2O2 production. Our experiments confirm active H2O2 production by respiring mouse liver mitochondria and show that ATP synthase activation increases the rate of H2O2, suggesting that state 3 mitochondria might induce the cell through oxidative signals.
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http://dx.doi.org/10.1016/B978-0-12-405883-5.00007-7DOI Listing
January 2014

Knitting the catalytic pattern of artificial photosynthesis to a hybrid graphene nanotexture.

ACS Nano 2013 Jan 27;7(1):811-7. Epub 2012 Dec 27.

Center of Excellence for Nanostructured Materials (CENMAT) and INSTM, unit of Trieste, Dipartimento di Scienze Chimiche e Farmaceutiche, University of Trieste, Piazzale Europa 1, I-34127 Trieste, Italy.

The artificial leaf project calls for new materials enabling multielectron catalysis with minimal overpotential, high turnover frequency, and long-term stability. Is graphene a better material than carbon nanotubes to enhance water oxidation catalysis for energy applications? Here we show that functionalized graphene with a tailored distribution of polycationic, quaternized, ammonium pendants provides an sp(2) carbon nanoplatform to anchor a totally inorganic tetraruthenate catalyst, mimicking the oxygen evolving center of natural PSII. The resulting hybrid material displays oxygen evolution at overpotential as low as 300 mV at neutral pH with negligible loss of performance after 4 h testing. This multilayer electroactive asset enhances the turnover frequency by 1 order of magnitude with respect to the isolated catalyst, and provides a definite up-grade of the carbon nanotube material, with a similar surface functionalization. Our innovation is based on a noninvasive, synthetic protocol for graphene functionalization that goes beyond the ill-defined oxidation-reduction methods, allowing a definite control of the surface properties.
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http://dx.doi.org/10.1021/nn305313qDOI Listing
January 2013

Electrochemical study of hydrogen peroxide formation in isolated mitochondria.

Bioelectrochemistry 2012 Jun 3;85:21-8. Epub 2011 Dec 3.

Experimental Oncology Dept. European Institute of Oncology, Via Adamello 16, 20139 Milan, Italy.

Mitochondrial respiration generates reactive oxygen species that are involved in physiological and pathological processes. The majority of methods, with exception of electron paramagnetic resonance, used to evaluate the identity, the rate and the conditions of the reactive oxygen species produced by mitochondria, are mainly based on oxidation sensitive markers. Following latest electrochemical methodology, we implemented a novel electrochemical assay for the investigation of aerobic metabolism in preparations of isolated mitochondria through simultaneous measurement of O₂ consumption and reactive species production. This electrochemical assay reveals active H₂O₂ production by respiring mouse liver mitochondria, and shows that ATP synthase activation and moderate depolarization increase the rate of H₂O₂ formation, suggesting that ATP synthesizing (state 3) mitochondria might contribute to oxidative stress or signaling.
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http://dx.doi.org/10.1016/j.bioelechem.2011.11.005DOI Listing
June 2012

Highly electroconductive multiwalled carbon nanotubes as potentially useful tools for modulating calcium balancing in biological environments.

Nanomedicine 2012 Apr 30;8(3):299-307. Epub 2011 Jun 30.

Institute of Translational Pharmacology, Consiglio Nazionale delle Ricerche (CNR), Rome, Italy.

Aiming to explore the mechanisms modulating cell-carbon nanotube interactions, we investigated whether Ca(2+) ion balancing between intra- and extracellular environments could be affected by multiwalled carbon nanotubes (MWCNTs). We analyzed the effects induced by two different kinds of MWCNTs (as prepared and annealed at 2400°C) on the intracellular Ca(2+) ion levels in rat electrically sensitive cells and on the intercellular junction integrity of rat adenocarcinoma colon cells and platelet aggregation ability, which depend on the Ca(2+) concentration in the medium. MWCNTs, purified by annealing and more electroconductive as compared to nonannealed MWCNTs, affected Ca(2+) ion balancing between extra- and intracellular environments and induced changes on Ca(2+) ion-dependent cellular junctions and platelet aggregation, behaving as the calcium chelator ethylene glycol tetraacetic acid. This could be due to the sorption of cationic Ca(2+) ions on CNTs surface because of the excess of negatively charged electrons on the aromatic units formed on MWCNTs after annealing. From the ClinicAL Editor: The authors investigated whether Ca(2+) ion balance between intra- and extracellular space can be modulated by multiwalled carbon nanotubes (MWCNTs). Annealed nanotubes induced changes on Ca(2+) dependent cellular junctions and platelet aggregation, behaving similary to ethylene glycol tetraacetic acid, an established calcium chelator.
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http://dx.doi.org/10.1016/j.nano.2011.06.018DOI Listing
April 2012

Efficient water oxidation at carbon nanotube-polyoxometalate electrocatalytic interfaces.

Nat Chem 2010 Oct 8;2(10):826-31. Epub 2010 Aug 8.

Center of Excellence for Nanostructured Materials (CENMAT), INSTM, Dipartimento di Scienze Farmaceutiche, Università di Trieste, Piazzale Europa 1, 34127 Trieste, Italy.

Water is the renewable, bulk chemical that nature uses to enable carbohydrate production from carbon dioxide. The dream goal of energy research is to transpose this incredibly efficient process and make an artificial device whereby the catalytic splitting of water is finalized to give a continuous production of oxygen and hydrogen. Success in this task would guarantee the generation of hydrogen as a carbon-free fuel to satisfy our energy demands at no environmental cost. Here we show that very efficient and stable nanostructured, oxygen-evolving anodes are obtained by the assembly of an oxygen-evolving polyoxometalate cluster (a totally inorganic ruthenium catalyst) with a conducting bed of multiwalled carbon nanotubes. Our bioinspired electrode addresses the one major challenge of artificial photosynthesis, namely efficient water oxidation, which brings us closer to being able to power the planet with carbon-free fuels.
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http://dx.doi.org/10.1038/nchem.761DOI Listing
October 2010