Dendrimer Stabilized Nanoalloys for Ink-Jet Printing of Surface-Enhanced Raman Scattering Substrates

Research on paper substrates prepared by inkjet deposition of metal nanoparticles for sensing applications has become a hot topic in recent years; however, the design of such substrates based on the deposition of alloy nanoparticles remains less explored. Herein, we report for the first time the inkjet printing of dendrimer-stabilized colloidal metal nanoalloys for the preparation of paper substrates for surface-enhanced Raman scattering (SERS) spectroscopy. To this end, nanoassemblies containing variable molar ratios of Au:Ag were prepared in the presence of poly(amidoamine) dendrimer (PAMAM), resulting in plasmonic properties that depend on the chemical composition of the final materials. The dendrimer-stabilized Au:Ag:PAMAM colloids exhibit high colloidal stability, making them suitable for the preparation of inks for long-term use in inkjet printing of paper substrates. Moreover, the pre-treatment of paper with a polystyrene (PS) aqueous emulsion resulted in hydrophobic substrates with improved SERS sensitivity, as illustrated in the analytical detection of tetramethylthiuram disulfide (thiram pesticide) dissolved in aqueous solutions. We suggest that the interactions established between the two polymers (PAMAM and PS) in an interface region over the cellulosic fibres, resulted in more exposed metallic surfaces for the adsorption of the analyte molecules. The resulting hydrophobic substrates show long-term plasmonic stability with high SERS signal retention for at least ninety days.publishe

pH-responsive hybrid nanoassemblies for cancer treatment: formulation development, optimization, and in vitro therapeutic performance

Current needs for increased drug delivery carrier efficacy and specificity in cancer necessitate the adoption of intelligent materials that respond to environmental stimuli. Therefore, we developed and optimized pH-triggered drug delivery nanoassemblies that exhibit an increased release of doxorubicin (DOX) in acidic conditions typical of cancer tissues and endosomal vesicles (pH 5.5) while exhibiting significantly lower release under normal physiological conditions (pH 7.5), indicating the potential to reduce cytotoxicity in healthy cells. The hybrid (polymeric/lipid) composition of the lyotropic non-lamellar liquid crystalline (LNLCs) nanoassemblies demonstrated high encapsulation efficiency of the drug (>90%) and high drug loading content (>7%) with colloidal stability lasting at least 4 weeks. Confocal microscopy revealed cancer cellular uptake and DOX-loaded LNLCs accumulation near the nucleus of human hepatocellular carcinoma cells, with a large number of cells appearing to be in apoptosis. DOX-loaded LNLCs have also shown higher citotoxicity in cancer cell lines (MDA-MB 231 and HepG2 cell lines after 24 h and in NCI-H1299 cell line after 48 h) when compared to free drug. After 24 h, free DOX was found to have higher cytotoxicity than DOX-loaded LNLCs and empty LNLCs in the normal cell line. Overall, the results demonstrate that DOX-loaded LNLCs have the potential to be explored in cancer therapy.This research was funded by FCT/MCTES—Foundation for Science and Technology I.P. from the Minister of Science, Technology, and Higher Education (PIDDAC) and European Regional Development Fund (ERDF) by the COMPETE—Programa Operacional Factores de Competitividade (POFC) through the project CONCERT [POCI-01-0145-FEDER-032651 and PTDC/NAN-MAT/326512017]. This work was also supported by the Portuguese Foundation for Science and Technology (FCT) in the framework of the Strategic Funding UIDB/04650/2020 and UIDB/04050/2020 (CF-UM-UP and CBMA). The authors are grateful for the support of the BioISI center grant (UIDB/04046/2020 and UIDP/04046/2020), financed by FCT. R.M. acknowledges FCT I.P. for funding within the Scientific Employment Stimulus instrument (CEECIND/00526/2018)

Homogeneous point mutation detection by quantum dot-mediated two-color fluorescence coincidence analysis

This report describes a new genotyping method capable of detecting low-abundant point mutations in a homogeneous, separation-free format. The method is based on integration of oligonucleotide ligation with a semiconductor quantum dot (QD)-mediated two-color fluorescence coincidence detection scheme. Surface-functionalized QDs are used to capture fluorophore-labeled ligation products, forming QD-oligonucleotide nanoassemblies. The presence of such nanoassemblies and thereby the genotype of the sample is determined by detecting the simultaneous emissions of QDs and fluorophores that occurs whenever a single nanoassembly flows through the femtoliter measurement volume of a confocal fluorescence detection system. The ability of this method to detect single events enables analysis of target signals with a multiple-parameter (intensities and count rates of the digitized target signals) approach to enhance assay sensitivity and specificity. We demonstrate that this new method is capable of detecting zeptomoles of targets and achieve an allele discrimination selectivity factor >10(5)

Photochromic Molecular Materials for the Controlled Organization of Nanoparticles

The displacement of polystyrene nanoparticles promoted by bulk azo mass transfer using interferential light illumination will be discussed. For this a variety of different push pull small azo molecules were synthesis and characterized, changing the polarity, hydrophobicity and bulky groups around the azo moiety. These small dyes appear to orientate quicker than similarly report azo polymers, of entanglement of the azobenzene group in the polymeric backbone. These dyes were used to form surface relief grating on the thin films due to the bulk mass transfer. This mass transfer was exploited to align polystyrene nanoparticles in the crests and troughs of the gratings. Secondly, the fabrication of azo nano objects will be discussed, here the photochromic studies showed the azo nanospheres had a larger photochemical conversion then their corresponding thin films. These nanoobjects were then successfully coated with gold nanoparticles by using azo compounds with dithiane or thioacetate ligands that are known to complex gold. The hybrid structure was proved by Raman spectroscopy where surface enhanced resonance effects have been evidenced upon excitation in the gold localized surface plasmon bands. Such assemblies, following a reverse architecture as the azo is situated in the core and not on the periphery, represents the first example of high-payload photochromic nanoparticles

Development and characterisation of advanced MRI contrast agents

Magnetic resonance imaging (MRI) is a non-invasive imaging technique that has emerged as one of the most powerful diagnostic tools in clinical medicine. Paramagnetic contrast agents (CAs), usually Gd-chelates, are often employed during an MRI scan to achieve better image contrast between a diseased and normal tissue. They work by modulating the longitudinal and transverse relaxation times of water protons within the tissues. Most of the current commercially available CAs, which can be broadly classified as low molecular weight CAs, are small, fast reorientating molecules with restricted specificity and targeting ability and relatively moderate efficiency (called relaxivity in the case of MRI CAs). Also, there have been reports linking their use with nephrogenic systemic fibrosis, a serious disorder that occurs in patients with chronic kidney disease and acute renal failure. The suspected cause is the dissociation of the Gd-ligand complex due to their slow clearance as a result of renal failure, suggesting the need for more stable CAs. This has spurred interest in developing stable high molecular weight CAs. Such CAs possess slow molecular reorientation, resulting in much better contrast activities compared to their low molecular weight counterparts. They are also passively targeted towards tumours due to enhanced permeation and retention. A useful approach for increasing the molecular weight (or size) of MRI CAs is to impart amphiphilic character to them and let them self-assemble to form supramolecular nanoparticles (hereafter called nanoassemblies) upon dispersion in an aqueous solution. Supramolecular MRI CAs have all the benefits of high molecular weight CAs and can also provide high payloads of Gd(III) ions at the target site. In this thesis, highly ordered supramolecular nanoassemblies, made of paramagnetic amphiphilic chelates, were developed and investigated as advanced MRI CAs. To begin with, relaxation theory based on the dipolar and scalar relaxation mechanisms in the two spin system was pedagogically (re)derived to understand the theoretical framework of a paramagnetic MRI CA. Based on this, novel paramagnetic amphiphilic chelates were designed and synthesised with the ability to form lamellar or inverse bicontinuous cubic nanoassemblies by themselves. Selected paramagnetic amphiphiles were also incorporated, at various concentrations, within non-ionic external lipid matrices to assist the formation of highly ordered mesophases. Differential scanning calorimetry, thermogravimetric analysis, polarised optical microscopy and synchrotron small angle X-ray scattering (SAXS) were employed to characterise the neat amphiphiles and examine the self-assembly structures of their bulk phases. Upon dispersion in an aqueous solution, cryogenic transmission electron microscopy, variable temperature SAXS, and dynamic light scattering were used to investigate the morphology, structure and size of the dispersed nanoassemblies. To assess their potential as MRI CAs, the in-vitro relaxivities of the nanoassemblies were measured at various magnetic field strengths (ranging from 0.47 – 11.74 T) and in some cases, contrast enhancement was directly observed by performing in-vitro MRI experiments. For nanoassemblies made solely from the paramagnetic amphiphiles, molecular parameters such as reorientational correlation time and water exchange rate were determined by comparing the theoretical models to the variable temperature 17O transverse relaxation time measurements, and the variable magnetic field 1H longitudinal relaxation time measurements (also known as nuclear magnetic relaxation dispersions profiles). It was found that paramagnetic amphiphilic chelates containing branched hydrophobic chains can be incorporated at a higher concentration within the inverse bicontinuous cubic phases of an external lipid compared to paramagnetic amphiphilic chelates with unsaturated unbranched chains. In general, the in-vitro relaxivities (and in some cases image contrast) of the nanoassemblies were found to be higher than Magnevist, a commercially available CA, at both low and high magnetic field strengths. The nanoassemblies made of mixed amphiphiles (paramagnetic amphiphilic chelates and non-ionic external lipids) showed better relaxivities than those made solely from paramagnetic amphiphiles at high magnetic field strength. Liposomal nanoassemblies made from sole paramagnetic amphiphiles with branched chains and monoamide conjugates as the chelating head groups showed faster water exchange than the previously reported paramagnetic liposomes (with unbranched chains and bisamide conjugates as head groups). Nevertheless, their relaxation enhancement was still found to be limited by water exchange. Improved relaxivities were largely due to the slow molecular reorientation. Finally, a novel and efficient method of determining relaxivities was also developed. According to this method, relaxivity of a paramagnetic MRI CA can be determined by relating the two NMR observables: induced chemical shifts and enhanced relaxation rates, both of which are dependent on the concentration of the paramagnetic MRI CA

Laccase-Functionalized Graphene Oxide Assemblies as Efficient Nanobiocatalysts for Oxidation Reactions

Multi-layer graphene oxide-enzyme nanoassemblies were prepared through the multi-point covalent immobilization of laccase from Trametes versicolor (TvL) on functionalized graphene oxide (fGO). The catalytic properties of the fGO-TvL nanoassemblies were found to depend on the number of the graphene oxide-enzyme layers present in the nanostructure. The fGO-TvL nanoassemblies exhibit an enhanced thermal stability at 60 degrees C, as demonstrated by a 4.7-fold higher activity as compared to the free enzyme. The multi-layer graphene oxide-enzyme nanoassemblies can efficiently catalyze the oxidation of anthracene, as well as the decolorization of an industrial dye, pinacyanol chloride. These materials retained almost completely their decolorization activity after five reaction cycles, proving their potential as efficient nano- biocatalysts for various applications

Immobilization of Inorganic Nanoparticles on Responsive Polymer Brushes

Exploitation of well defined responsive polymer brushes for direct and controlled immobilization of metal/semiconductor nanoparticles on macroscopic surfaces has been demonstrated. The employed approach offers the possibility of the organization of a variety of inorganic nanoparticles by irreversible bonding and homogenous distribution on an underlying substrate. The immobilization process has been realized by chemical grafting of a variety of polymer brushes on a suitable substrate followed by the attachment of pre-/in-situ formed nanoparticles exploiting the chemical/physical interactions between surface functionalities of nanoparticles and polymer chain segments. A number of polymer brushes including poly (acrylic acid), polystyrene, poly (2-vinyl pyridine) and poly (N-isopropyl acrylamide) brushes have been prepared on silicon substrate by the “grafting to” approach. A variety of inorganic nanoparticles such as quantum dots (CdTe) noble metals (gold and silver) and magnetic (Fe3O4) were immobilized on macroscopic surfaces to impart them photo luminescent, catalytic or magnetic properties. In addition, responsiveness of grafted polymer brushes in terms of variation in thickness (due to changes in chain conformation) as a function of external stimuli such as solvent and pH allowed to use the resulting polymer brush-nanoparticles nanoassemblies in the fabrication of nanosensors. The design of fabricated nanosensors is based on the modulation in the interparticle distance of immobilized nanoparticles due to swelling/deswelling of underlined polymer brushes in response to some external trigger