SCREENING FOR OXIDATIVE STRESS ELICITED BY ENGINEERED NANOMATERIALS: EVALUATION OF ACELLULAR DCFH ASSAY

The DCFH assay is commonly used for measuring free radicals generated by engineered nanomaterials (ENM), a well-established mechanism of ENM toxicity. Concerns exist over susceptibility of the DCFH assay to: assay conditions, adsorption of DCFH onto ENM, fluorescence quenching and light scattering. These effects vary in magnitude depending on ENM physiochemical properties and concentration. A rigorous evaluation of this method is still lacking. The objective was to evaluate performance of the DCFH assay for measuring ENM-induced free radicals. A series of diverse and well-characterized ENM were tested in the acellular DCFH assay. We investigated the effect of sonication conditions, dispersion media, ENM concentration, and the use of horseradish peroxidase (HRP) on the DCFH results. The acellular DCFH assay suffers from high background signals resulting from dye auto-oxidation and lacks sensitivity and robustness. DCFH oxidation is further enhanced by HRP. The number of positive ENM in the assay and their relative ranking changed as a function of experimental conditions. An inverse dose relationship was observed for several Carbon-based ENM. Overall, these findings indicate the importance of having standardized assays for evaluating ENM toxicity and highlights limitations of the DCFH assay for measuring ENM-induced free radicals

Polyvinyl alcohol as a biocompatible alternative for the passivation of gold nanorods

The functionalization of gold nanorods (GNRs) with polymers is essential for both their colloidal stability and biocompatibility. However, a bilayer of the toxic cationic surfactant cetyl trimethylammonium bromide (CTAB) adsorbed on the nanorods complicates this process. Herein, we report on a strategy for the biocompatible functionalization of GNRs with a hydrophobic polymeric precursor, polyvinyl acetate, which is then transformed into its hydrophilic analogue, polyvinyl alcohol. This polymer was chosen due to its well-established biocompatibility, tunable “stealth” properties, tunable hydrophobicity, and high degree of functionality. The biocompatibility of the functionalized GNRs was tested by exposing them to primary human blood monocyte derived macrophages; the advantages of tunable hydrophobicity were demonstrated with the long-term stable encapsulation of a model hydrophobic drug molecule

Tandem Hydroperoxyl-Alkylperoxyl Radical Quenching by an Engineered Nanoporous Cerium Oxide Nanoparticle Macrostructure (NCeONP): Toward Efficient Solid-State Autoxidation Inhibitors

The use of nanomaterials as inhibitors of the autoxidation of organic materials is attracting tremendous interest in petrochemistry, food storage, and biomedical applications. Metal oxide materials and CeO2 in particular represent one of the most investigated inorganic materials with promising radical trapping and antioxidant abilities. However, despite the importance, examples of the CeO2 material’s ability to retard the autoxidation of organic substrates are still lacking, together with a plausible chemical mechanism for radical trapping. Herein, we report the synthesis of a new CeO2-derived nanoporous material (NCeONP) with excellent autoxidation inhibiting properties due to its ability to catalyze the cross-dismutation of alkyl peroxyl (ROO•) and hydroperoxyl (HOO•) radicals, generated in the system by the addition of the pro-aromatic hydrocarbon γ-terpinene. The antioxidant ability of NCeONP is superior to that of other nanosized metal oxides, including TiO2, ZnO, ZrO2, and pristine CeO2 nanoparticles. Studies of the reaction with a sacrificial reductant allowed us to propose a mechanism of inhibition consisting of H atom transfer from HOO• to the metal oxides (MOx + HOO• → MOx-H• + O2), followed by the release of the H atom to an ROO• radical (MOx-H• + ROO• → MOx + ROOH). Besides identifying NCeONP as a promising material for developing effective antioxidants, our study provides the first evidence of a radical mechanism that can be exploited to develop novel solid-state autoxidation inhibitors

Multicore–Shell PNIPAm-<i>co</i>-PEGMa Microcapsules for Cell Encapsulation

The overall goal of this study was to fabricate multifunctional core–shell microcapsules with biological cells encapsulated within the polymer shell. Biocompatible temperature responsive microcapsules comprised of silicone oil droplets (multicores) and yeast cells embedded in a polymer matrix (shell) were prepared using a novel microarray approach. The cross-linked polymer shell and silicone multicores were formed in situ via photopolymerization of either poly(<i>N</i>-isopropylacryamide)(PNIPAm) or PNIPAm, copolymerized with poly(ethylene glycol monomethyl ether monomethacrylate) (PEGMa) within the droplets of an oil-in-water-in-oil double emulsion. An optimized recipe yielded a multicore–shell morphology, which was characterized by optical and laser scanning confocal microscopy (LSCM) and theoretically confirmed by spreading coefficient calculations. Spreading coefficients were calculated from interfacial tension and contact angle measurements as well as from the determination of the Hamaker constants and the pair potential energies. The effects of the presence of PEGMa, its molecular weight (<i>M</i>_n 300 and 1100 g/mol), and concentration (10, 20, and 30 wt %) were also investigated, and they were found not to significantly alter the morphology of the microcapsules. They were found, however, to significantly improve the viability of the yeast cells, which were encapsulated within PNIPAm-based microcapsules by direct incorporation into the monomer solutions, prior to polymerization. Under LSCM, the fluorescence staining for live and dead cells showed a 30% viability of yeast cells entrapped within the PNIPAm matrix after 45 min of photopolymerization, but an improvement to 60% viability in the presence of PEGMa. The thermoresponsive behavior of the microcapsules allows the silicone oil cores to be irreversibly ejected, and so the role of the silicone oil is 2-fold. It facilitates multifunctionality in the microcapsule by first being used as a template to obtain the desired core–shell morphology, and second it can act as an encapsulant for oil-soluble drugs. It was shown that the encapsulated oil droplets were expelled above the volume phase transition temperature of the polymer, while the collapsed microcapsule remained intact. When these microcapsules were reswollen with an aqueous solution, it was observed that the hollow compartments refilled. In principle, these hollow-core microcapsules could then be filled with water-soluble drugs that could be delivered in vivo in response to temperature