Daniel Grasseschi

A novel functionalisation process for glucose oxidase immobilisation in poly(methyl methacrylate) microchannels in a flow system for amperometric determinations.

Different materials like glass, silicon and poly(methyl methacrylate) (PMMA) are being used to immobilise enzymes in microchannels. PMMA shows advantages such as its low price, biocompatibility and attractive mechanical and chemical properties. Despite this, the introduction of reactive functional groups on PMMA is still problematic, either because of the complex chemistry or extended reaction time involved. In this paper, a new methodology was developed to immobilise glucose oxidase (GOx) in PMMA microchannels, with the benefit of a rapid immobilisation process and a very simple route. The new procedure involves only two steps, based on the reaction of 5.0% (w/w) polyethyleneimine (PEI) with PMMA in a dimethyl sulphoxide medium, followed by the immobilisation of glucose oxidase using a solution containing 100U enzymes and 1.0% (v/v) glutaraldehyde. The reactors prepared in this way were evaluated by a flowing system with amperometric detection (+0.60V) based on the oxidation of the H2O2 produced by the reactor. The microreactor proposed here was able to work with high bioconversion and a frequency of 60 samples h(-1), with detection and quantification limits of 0.50 and 1.66µmol L(-1), respectively. Michaelis-Menten parameters (Vmax and KM) were calculated as 449±47.7nmol min(-1) and 7.79±0.98mmol. Statistical evaluations were done to validate the proposed methodology. The content of glucose in natural and commercial coconut water samples was evaluated using the developed method. Comparison with spectrophotometric measurements showed that both methodologies have a very good correlation (tcalculated, 0.05, 4=1.35

Unraveling the nature of Turkevich gold nanoparticles: the unexpected role of the dicarboxyketone species

By monitoring the synthesis of the Turkevich gold nanoparticles, under appropriate conditions, it was possible to probe the formation of the dicarboxyketone intermediate species, revealing their unexpected strong interaction with the gold nanoparticles. The dicarboxyketone species exhibited a contrasting spectral and kinetics behaviour in relation to citrate stabilized products, explaining several existing controversial points, such as the lack of reproducibility and variable SERS response. In these species, because of the covalent interaction, the chemical mechanisms involved in SERS predominate over the electromagnetic contribution observed for the citrate stabilized gold nanoparticles. New interesting aspects were found, such as a greater stabilization and strong SERS response observed even for the non-aggregated nanoparticles.

Unveiling the Structure of Polytetraruthenated Nickel Porphyrin by Raman Spectroelectrochemistry

The structure of polytetraruthenated nickel porphyrin was unveiled for the first time by electrochemistry, Raman spectroelectrochemistry, and a hydroxyl radical trapping assay. The electrocatalytic active material, precipitated on the electrode surface after successive cycling of [NiTPyP{Ru(bipy)2Cl}4](4+) species in strong aqueous alkaline solution (pH 13), was found to be a peroxo-bridged coordination polymer. The electropolymerization process involves hydroxyl radicals (as confirmed by the characteristic set of DMPO/(•)OH adduct EPR peaks) as reaction intermediates, electrocatalytically generated in the 0.80-1.10 V range, that induce the formation of Ni-O-O-Ni coordination polymers, as evidenced by Raman spectroelectrochemistry and molecular modeling studies. The film growth is halted above 1.10 V due to the formation of oxygen gas bubbles.

Hyperspectral dark-field microscopy of gold nanodisks

The light scattering properties of hexagonal and triangular gold nanodisks were investigated by means of Cytoviva hyperspectral dark-field microscopy, exploring the huge enhancement of the scattered waves associated with the surface plasmon resonance (SPR) effect. Thanks to the high resolution capability of the dark-field microscope, the SPR effect turned it possible to probe the individual nanoparticles directly from their hyperspectral images, extrapolating the classical optical resolution limit, and providing their corresponding extinction spectra. Blue spectral shifts involving the in-plane dipolar modes were observed for the hexagonal gold nanodisks in relation to the triangular ones, allowing their spectroscopic differentiation in the dark-field images.

How relevant can the SERS effect in isolated nanoparticles be

Gold nanoparticles electrostatically stabilized with negatively charged ruthenium complexes were individually monitored based on their characteristic plasmon bands, by means of hyperspectral dark field microscopy. Very strong SERS enhancements were observed for the isolated gold nanoparticles at exciting wavelengths in resonance with the surface plasmon band, using confocal Raman microscopy, revealing a contrasting behaviour between the agglomerated and non-agglomerated systems. The results highlighted the relevant role played by the surface enhanced resonance Raman mechanisms (SERRS) in isolated nanoparticles, more than compensating for the lack of local hot spots, in relation to the agglomerated systems.

Gold nanoparticles functionalised with Ru–dicarboxybipyridine–trimercaptotriazine: SERS effect and application in plasmonic dye solar cells

A dicarboxybipyridine–trimercaptotriazine ruthenium complex, primarily designed for dye–sensitised solar cells (DSSC), has been successfully employed in the generation of electrostatically stabilised gold colloids, because of its high negative charge and capability of binding to gold nanoparticles via the sulphur groups. Strong SERS enhancements have been obtained for the isolated gold nanoparticles, as monitored by dark field hyperspectral and confocal Raman microscopy. A preliminary design of TiO2 solar cell incorporating the ruthenium dye in association with the gold nanoparticles, has been tested, in the presence of a cobalt complex mediator. The results were considered rather promising, in spite of the slight decay of efficiency in relation to the original dye cell, owing to lixiviation processes and electron recombination effects at the interface.

Confocal Raman microscopy and hyperspectral dark field microscopy imaging of chemical and biological systems

Hyperspectral imaging can provide accurate information on the distribution of the chemical species in materials and biological samples, based on the analysis of their electronic and vibrational profiles. In special, confocal Raman microscopy is one of the best ways to access the chemical distribution of molecules, especially under resonance Raman or SERS conditions. On the other hand, enhanced dark field optical microscopy can be employed for hyperspectral imaging in the visible and near-IR region, while extending the optical resolution up to the nanoscale dimension. It allows the detection of gold or silver single nanoparticles, as well as spectral monitoring from the characteristic surface plasmon bands. These two hyperspectral microscopies can be conveniently combined to provide nanoscale electronic and vibrational information of the species present in a wide variety of chemical and biological systems. A case study focusing on the improvement of the classical spot-test analysis of nickel(II) ions with dithizone is here detailed. A great enhancement of sensitivity in the detection of nickel(II) ions, by at least 4 orders of magnitude, has been observed in this work. Hyperspectral measurements allowed the mapping of the gold nanoparticles (AuNP) distribution on cellulose fibers and on glass, and the evaluation of their extinction and SERS spectra for analytical purposes.