Recuento de Streptococcus mutans sobre restauraciones realizadas con adhesivo con y sin carga de nanopartículas de óxido de zinc y cobre / Streptococcus mutans count on restorations made with adhesive with and without loading zinc and copper oxide nanoparticles

Objetivo: Determinar las propiedades antimicrobianas de la incorporación de nanopartículas de óxido de zinc y cobre en un adhesivo de grabado y lavado total sobre Streptococcus mutans en pacientes con restauraciones de resina compuesta confeccionadas con adhesivo cargado. Métodos: Estudio experimental, randomizado, la muestra estuvo conformada por 25 pacientes, de ambos sexos, pertenecientes al posgrado de Ortodoncia de la Facultad de Odontología de la Universidad de Chile, en los cuales se confirmó presencia de Streptococcus mutans en saliva. Se confeccionaron restauraciones de resina compuesta oclusales, en premolares superiores con indicación de exodoncia por el tratamiento de ortodoncia, con adhesivo cargado (cuya composición fue 5/0,2 por ciento ZnO y Cu, respectivamente) y control (sin presencia de nanopartículas en su composición), según el listado de aleatorización. Se tomaron muestras microbiológicas en tres tiempos con la técnica de la cubeta (antes, 1 semana y 4 semanas posterior a la confección de las restauraciones). Se obtuvieron, aislaron e identificaron colonias de Streptococcus mutans a partir de las muestras obtenidas. Se usó el test de Mann-Whitney mediante el paquete estadístico SPSS v.21 Resultados: El promedio del recuento de UFC de Streptococcus mutans en el grupo experimental fue mayor posterior a la confección de las restauraciones de resina compuesta. Los resultados de la identificación molecular por PCR demuestran la presencia de Streptococcus mutans en 20 de 25 muestras. Conclusiones: No existen diferencias en el recuento de Streptococcus mutans antes y después de la aplicación del adhesivo sobre las restauraciones de resina compuesta(AU)

Objective: To determine the antimicrobial properties of the incorporation of zinc and copper oxide nanoparticles in an etching and total wash adhesive on Streptococcus mutans in patients with composite resin restorations made with loaded adhesive. Methods: Experimental and randomized trial, the sample were 25 patients, of both sexes, belonging to the FOUCH Orthodontic postgraduate program, in whom the presence of Streptococcus mutans in saliva was confirmed. Occlusal composite resin restorations were made in upper premolars with indication of extraction by orthodontic treatment, with loaded adhesive (whose composition is 5 / 0.2% ZnO and Cu respectively) and control (without the presence of nanoparticles in their composition), according to the scrambling listing. Microbiological samples were taken in three stages with the cuvette technique (before, 1 week and 4 weeks after the restoration was made). Colonies of Streptococcus mutans were obtained, isolated and identified from the samples obtained. The statistical analysis used the SPSS v.21 software, the data was analyzed by Mann Whitney test Results: The average CFU count of Streptococcus mutans in the experimental group (adhesive modified with zinc oxide and copper nanoparticles) was higher after the fabrication of composite resin restorations. The results of molecular identification by PCR demonstrate the presence of Streptococcus mutans in 20 of 25 samples. Conclusions: There are no differences in the count of Streptococcus mutans before and after the application of the adhesive on the composite resin restorations(AU)

Design and development of porous terracotta disc: An eco-friendly novel control agent for mosquito larvae.

In the present work, we synthesized silver nanoparticles supported by rice husk by hydrothermal treatment, as-synthesized silver nanoparticles rice husk (AgNPs-RH) bio-composite mixed with potter clay thoroughly, molded, dried into a disc-shaped before firing and applying as a point of use larvicidal agent. As designed, porous terracotta disc (PTD) infused with AgNPs-RH-biocomposite were characterized by UV spectrophotometer, Fourier-transform infrared spectroscopy, transmission electron microscopy, X-ray diffraction analysis and energy-dispersive X-ray spectroscopy. The amount of silver ions released from the PTD was also found to be within the prescribed limit of 0.1 ppm-level. Later we dropped the PTD and tested its larvicidal activity against the IVth instar larva stage of Aedes, Anopheles and Culex species. We found 100% larvicidal mortality in 24 h of exposure to the designed PTD and the amount of silver released from the porous disc was found to be 0.0343 ppm. Further from the histopathological studies of dead larvae revealed that the silver ions from the PTD have substantially damaged the exoskeleton of larvae.

Assessment of strategies for the formation of stable suspensions of titanium dioxide nanoparticles in aqueous media suitable for the analysis of biological fluids.

Due to their omnipresence in consumer products, there is a growing concern about the potential effects of nanoparticles on human health. Toxicological assessment and NP end-product studies require proper quantification of these materials in biological fluids. However, their quantifications in these media require stable predispersed NP solutions in aqueous media to enable the fortification in the matrices of interest or the preparation of calibration standards. In this study, a sample preparation scheme was developed by studying various dispersion media (polyvinylpyrrolidone and polyethylene glycol) and sonication strategies (bath and ultrasonic probe) to ensure homogeneous dispersion of titanium dioxide nanoparticles. Optimization of the various parameters was performed using SRM NIST 1898 NP reference material, composed of rutile and anatase phases. Number-based size distribution for titanium dioxide NPs was determined by dynamic light scattering and single-particle inductively coupled plasma mass spectrometry to evaluate the procedure efficiency. Changes in mean size and most frequent size distribution were also studied to determine if the agglomeration of nanoparticles occurs at the various dispersion conditions tested. Among the different dispersion parameters tested herein, the use of polyvinylpyrrolidone combined with a sonication process generated by a probe leads to a significant improvement in terms of suspension efficiency and stability over 72 h. The dispersion efficiency of the proposed methodology was assessed by single-particle inductively coupled plasma mass spectrometry with spiked biological fluids such as urine and blood. Graphical abstract.

Biocompatibility enhancement of graphene oxide-silver nanocomposite by functionalisation with polyvinylpyrrolidone.

Several materials such as silver are used to enhance graphene oxide (GO) sheets antimicrobial activity. However, these toxic materials decrease its biocompatibility and hinder its usage in many biological applications. Therefore, there is an urgent need to develop nanocomposites that can preserve both the antimicrobial activity and biocompatibility simultaneously. This work highlights the importance of functionalisation of GO sheets using Polyvinylpyrrolidone (PVP) and decorating them with silver nanoparticles (AgNPs) in order to enhance their antimicrobial activity and biocompatibility at the same time. The structural and morphological characterisations were performed by UV-Visible, Fourier transform infrared (FTIR), and Raman spectroscopic techniques, X-ray diffraction (XRD), and high-resolution transmission electron microscopy (HR-TEM). The antimicrobial activities of the prepared samples against Staphylococcus aureus, Pseudomonas aeruginosa, and Candida albicans were studied. The cytotoxicity of prepared materials was tested against BJ1 normal skin fibroblasts. The results indicated that the decoration with AgNPs showed a significant increase in the antimicrobial activity of GO and FGO sheets, and functionalisation of GO sheets and GO-Ag nanocomposite with PVP improved the cell viability about 40 and 35%, respectively.

Toxic Effects of Size-tunable Gold Nanoparticles on Caenorhabditis elegans Development and Gene Regulation.

We utilized size-tunable gold nanoparticles (Au NPs) to investigate the toxicogenomic responses of the model organism Caenorhabditis elegans. We demonstrated that the nematode C. elegans can uptake Au NPs coated with or without 11-mercaptoundecanoic acid (MUA), and Au NPs are detectable in worm intestines using X-ray microscopy and confocal optical microscopy. After Au NP exposure, C. elegans neurons grew shorter axons, which may have been related to the impeded worm locomotion behavior detected. Furthermore, we determined that MUA to Au ratios of 0.5, 1 and 3 reduced the worm population by more than 50% within 72 hours. In addition, these MUA to Au ratios reduced the worm body size, thrashing frequency (worm mobility) and brood size. MTT assays were employed to analyze the viability of cultured C. elegans primary neurons exposed to MUA-Au NPs. Increasing the MUA to Au ratios increasingly reduced neuronal survival. To understand how developmental changes (after MUA-Au NP treatment) are related to changes in gene expression, we employed DNA microarray assays and identified changes in gene expression (e.g., clec-174 (involved in cellular defense), cut-3 and fil-1 (both involved in body morphogenesis), dpy-14 (expressed in embryonic neurons), and mtl-1 (functions in metal detoxification and homeostasis)).

Potential of metabolic engineering in bacterial nanosilver synthesis.

The widespread applications of silver nanoparticles in present days demand an industrial-scale production process. The ability of bacteria to synthesise silver nanoparticles can be exploited to overcome many shortcomings associated with conventional production processes, such as high cost and nanoparticle toxicity. However, lack of a standardised protocol and suboptimal yield remain a major obstacle for bacterial synthesis route. A potential, yet unexplored, solution to this problem could be envisioned through rewiring of the metabolic network to direct cellular resources towards the product of interest. Mathematical modelling of metabolic pathway is the key to understand and manipulate the cellular metabolism for enhanced production of desired metabolite(s). The present study provides a perspective on the scope of metabolic engineering approaches to enhance bacterial synthesis of silver nanoparticles.

Exposure of engineered nanomaterials to plants: Insights into the physiological and biochemical responses-A review.

Recent investigations show that carbon-based and metal-based engineered nanomaterials (ENMs), components of consumer goods and agricultural products, have the potential to build up in sediments and biosolid-amended agricultural soils. In addition, reports indicate that both carbon-based and metal-based ENMs affect plants differently at the physiological, biochemical, nutritional, and genetic levels. The toxicity threshold is species-dependent and responses to ENMs are driven by a series of factors including the nanomaterial characteristics and environmental conditions. Effects on the growth, physiological and biochemical traits, production and food quality, among others, have been reported. However, a complete understanding of the dynamics of interactions between plants and ENMs is not clear enough yet. This review presents recent publications on the physiological and biochemical effects that commercial carbon-based and metal-based ENMs have in terrestrial plants. This document focuses on crop plants because of their relevance in human nutrition and health. We have summarized the mechanisms of interaction between plants and ENMs as well as identified gaps in knowledge for future investigations.

Precision improvement in dark-field microscopy imaging by using gold nanoparticles as an internal reference: a combined theoretical and experimental study.

Low accuracy is a big obstacle in the dark-field microscopy imaging (iDFM) technique in practical applications. In order to reduce the deviations and fluctuations in the observed or snapped scattered light in the iDFM technique caused by unavoidable measurement errors, bare gold nanoparticles (AuNPs) were introduced as an internal reference (IR). The feasibility of using AuNPs as the IR in iDFM in theory was verified. The function of the IR in improving the precision of the acquired data through post data analysis was identified by three kinds of experiments: monitoring the oxidation process of silver nanoparticles (AgNPs) at room temperature, quantifying the level of glucose with AgNPs used as probes and quantifying the change in the light intensity of AuNPs after the plasmon resonance energy transfer (PRET) between AuNPs and tetramethylrhodamine (TAMRA).

Biotechnological aspects of ZnO nanoparticles: overview on synthesis and its applications.

The physicochemical methods of the synthesis of zinc nanoparticles (ZnO NPs) and some detailed studies on ZnO toxicity mechanism and biokinetics have been reported. However, some of these physical and chemical methods of synthesis are expensive and can also have toxic substances absorbed onto them. Hence, eco-friendly synthesis of nanoparticles due to their easier process, cheaper availability, and high stability is dominating new research. In particular, ZnO NPs which are now being synthesized through major biological systems involved in this are bacteria, fungi, and plant extracts; this has increased studies in various applications in the biological field. In this review, we have elaborated on various natural source-mediated syntheses of ZnO NPs and their role in various biological activities like antimicrobial, anticandidal, larvicidal, cytotoxic, and photocatalytic activities. Apart from these applications, ZnO NPs are also reported to help to prevent dust formation, for several years, on oil paintings.

Acute and subchronic oral toxicity studies in rats with nanoscale and pigment grade titanium dioxide particles.

Data generated using standardized testing protocols for toxicity studies generally provide reproducible and reliable results for establishing safe levels and formulating risk assessments. The findings of three OECD guideline-type oral toxicity studies of different duration in rats are summarized in this publication; each study evaluated different titanium dioxide (TiO2) particles of varying sizes and surface coatings. Moreover, each study finding demonstrated an absence of any TiO2 -related hazards. To briefly summarize the findings: 1) In a subchronic 90-day study (OECD TG 408), groups of young adult male and female rats were dosed with rutile-type, surface-coated pigment-grade TiO2 test particles (d50 = 145 nm - 21% nanoparticles by particle number criteria) by oral gavage for 90 days. The no-adverse-effect level (NOAEL) for both male and female rats in this study was 1000 mg/kg bw/day, the highest dose tested. The NOAEL was determined based on a lack of TiO2 particle-related adverse effects on any in-life, clinical pathology, or anatomic/microscopic pathology parameters; 2) In a 28-day repeated-dose oral toxicity study (OECD TG 407), groups of young adult male rats were administered daily doses of two rutile-type, uncoated, pigment-grade TiO2 test particles (d50 = 173 nm by number) by daily oral gavage at a dose of 24,000 mg/kg bw/day. There were no adverse effects measured during or following the end of the exposure period; and the NOAEL was determined to be 24,000 mg/kg bw/day; 3) In an acute oral toxicity study (OECD TG 425), female rats were administered a single oral exposure of surface-treated rutile/anatase nanoscale TiO2 particles (d50 = 73 nm by number) with doses up to 5000 mg/kg and evaluated over a 14-day post-exposure period. Under the conditions of this study, the oral LD50 for the test substance was >5000 mg/kg bw. In summary, the results from these three toxicity studies - each with different TiO2 particulate-types, demonstrated an absence of adverse toxicological effects. Apart from reporting the findings of these three studies, this publication also focuses on additional critical issues associated with particle and nanotoxicology studies. First, describing the detailed methodology requirements and rigor upon which the standardized OECD 408 guideline subchronic oral toxicity studies are conducted. Moreover, an attempt is made to reconcile the complex issue of particle size distribution as it relates to measurements of nanoscale and pigment-grade TiO2 particles. Clearly this has been a confusing issue and often misrepresented in the media and the scientific literature. It is clear that the particle-size distribution for pigment-grade TiO2, contains a small ("tail") component of nanoscale particles (i.e., 21% by particle number and <1% by weight in the test material used in the 90-day study). However, this robust particle characterization finding should not be confused with mislabeling the test materials as exclusively in the nanoscale range. Moreover, based upon the findings presented herein, there appears to be no significant oral toxicity impact contributed by the nanoscale component of the TiO2 Test Material sample in the 90-day study. Finally, it seems reasonable to conclude that the study findings should be considered for read-across purposes to food-grade TiO2 particles (e.g., E171), as the physicochemical characteristics are quite similar.

The gold standard: gold nanoparticle libraries to understand the nano-bio interface.

Since the late 1980s, researchers have prepared inorganic nanoparticles of many types--including elemental metals, metal oxides, metal sulfides, metal selenides, and metal tellurides--with excellent control over size and shape. Originally many researchers were primarily interested in exploring the quantum size effects predicted for such materials. Applications of inorganic nanomaterials initially centered on physics, optics, and engineering but have expanded to include biology. Many current nanomaterials can serve as biochemical sensors, contrast agents in cellular or tissue imaging, drug delivery vehicles, or even as therapeutics. In this Account we emphasize that the understanding of how nanomaterials will function in a biological system relies on the knowledge of the interface between biological systems and nanomaterials, the nano-bio interface. Gold nanoparticles can serve as excellent standards to understand more general features of the nano-bio interface because of its many advantages over other inorganic materials. The bulk material is chemically inert, and well-established synthetic methods allow researchers to control its size, shape, and surface chemistry. Gold's background concentration in biological systems is low, which makes it relatively easy to measure it at the part-per-billion level or lower in water. In addition, the large electron density of gold enables relatively simple electron microscopic experiments to localize it within thin sections of cells or tissue. Finally, gold's brilliant optical properties at the nanoscale are tunable with size, shape, and aggregation state and enable many of the promising chemical sensing, imaging, and therapeutic applications. Basic experiments with gold nanoparticles and cells include measuring the toxicity of the particles to cells in in vitro experiments. The species other than gold in the nanoparticle solution can be responsible for the apparent toxicity at a particular dose. Once the identity of the toxic agent in nanoparticle solutions is known, researchers can employ strategies to mitigate toxicity. For example, the surfactant used at high concentration in the synthesis (0.1 M) of gold nanorods remains on their surface in the form of a bilayer and can be toxic to certain cells at 200 nM concentrations. Several strategies can alleviate the toxic response. Polyelectrolyte layer-by-layer wrapping can cover up the surfactant bilayer, or researchers can exchange the surfactant with chemically similar molecules. Researchers can also replace the surfactant with a biocompatible thiol or use a polymerizable surfactant that can be "stitched" onto the nanorods and reduce its lability. In all these cases, however, proteins or other molecules from the cellular media cover the engineered surface of the nanoparticles, which can drastically change the charges and functional groups on the nanoparticle surface.

Preparation and quality control of silver nanoparticle-antibody conjugate for use in electrochemical immunoassays.

Metal nanoparticle-antibody conjugates are often used as optical or electrochemical markers in applications like immunohistochemistry, lateral flow tests, biosensors and immunoassays. In order to serve that role, an antibody needs to be immobilized on the surface of the nanoparticle. This is easily done, as proteins bind to gold and silver nanoparticles spontaneously. However, this immobilization process might result in nanoparticle aggregation or the loss of the bioactivity of the conjugated antibodies. In this work the optimization of antibody immobilization on silver colloid in order to obtain conjugates with the best possible activity is investigated. The parameters investigated were the type of immobilization buffer, its molarity and pH, the nanoparticle/antibody ratio and also blocking and washing protocols to reduce non-specific binding. The functionality of the obtained conjugates was tested with electrochemical immunoassay. It was found out that the optimum environment for immobilization of an anti-myoglobin antibody on silver nanoparticles was 0.2M boric acid pH 6.5 with 10 µg of antibody loading per 1 mL of silver colloid. For an anti-troponin antibody it was 0.1M boric acid pH 7.5 also with 10 µg/mL of antibody loading. The main problem for silver conjugation was the tendency of silver nanoparticles to aggregate during the immobilization process, but by choosing the optimum conditions the aggregation problem was completely removed. Here it is demonstrated that by using the conjugates prepared with an optimized protocol an increase in the sensitivity of the assay 10 times can be achieved. The electrochemical immunoassay described here can be used as a test for quality control of conjugates and for the estimation of batch-to-batch variability.

Standardisation of metalloimmunoassay protocols for assessment of silver nanoparticle antibody conjugates.

The application of silver nanoparticles to electrochemical metalloimmunoassays has previously been reported (Szymanski et al., 2009) and is now used in the electrochemical Argento immunoassay platform via UK Patent No. 2458420. The development of an immunoassay in this format requires the optimisation of the antibody to silver nanoparticle conjugation process. Issues such as pH, antibody concentration and other factors can affect the assay performance. In order to determine the effect of these variables it is necessary to understand and control the effects of other factors that may affect the assay signal. In this study a number of conditions which affect the assay signal were identified and methods developed to minimise variability in the assay signal, resulting in a standardised method allowing easy comparisons of silver nanoparticle conjugates prepared and assayed at different times.

Optimization of immunolabeled plasmonic nanoparticles for cell surface receptor analysis.

Noble metal nanoparticles hold great potential as optical contrast agents due to a unique feature, known as the plasmon resonance, which produces enhanced scattering and absorption at specific frequencies. The plasmon resonance also provides a spectral tunability that is not often found in organic fluorophores or other labeling methods. The ability to functionalize these nanoparticles with antibodies has led to their development as contrast agents for molecular optical imaging. In this review article, we present methods for optimizing the spectral agility of these labels. We discuss synthesis of gold nanorods, a plasmonic nanoparticle in which the plasmonic resonance can be tuned during synthesis to provide imaging within the spectral window commonly utilized in biomedical applications. We describe recent advances in our group to functionalize gold and silver nanoparticles using distinct antibodies, including EGFR, HER-2 and IGF-1, selected for their relevance to tumor imaging. Finally, we present characterization of these nanoparticle labels to verify their spectral properties and molecular specificity.