Thilak K. Mudalige

Capillary Electrophoresis/Inductively-Coupled Plasma-Mass Spectrometry: Development and Optimization of a High Resolution Analytical Tool for the Size-Based Characterization of Nanomaterials in Dietary Supplements

We report the development and optimization of a system consisting of capillary electrophoresis (CE) interfaced with inductively coupled plasma mass spectrometry (ICPMS) for rapid and high resolution speciation and characterization of metallic (e.g., gold, platinum, and palladium) nanoparticles in a dietary supplement. Multiple factors, including surfactant type and concentration, pH of running buffer, and applied voltage, were investigated to optimize the separation conditions. It was found that by using the anionic surfactant sodium dodecyl benzenesulfonate (SDBS) in the running buffer the separation resolution was significantly improved, allowing for easy distinction of adjacent size fractions in a gold nanoparticle mixture with very small size differences (e.g., 5, 15, 20, and 30 nm). The type and concentration of the surfactant was found to be critical in obtaining sufficient separation while applied voltage and pH values of the running buffers largely affected the elution times by varying the electroosmotic flow. Quantum dots were used as mobility markers to eliminate the run-to-run variation. The diameters of the nanoparticles followed a linear relationship with their relative electrophoretic mobility, and size information on unknown samples could be extrapolated from a standard curve. The accuracy and precision of this method was confirmed using 10 and 30 nm gold nanoparticle standard reference materials. Furthermore, the method was successfully applied to the analysis of commercially available metallic nanoparticle-based dietary supplements, as evidenced by good agreement between the particle sizes calculated by CE/ICPMS and transmission electron microscopy (TEM).

Asymmetric Flow-Field Flow Fractionation Hyphenated ICP-MS as an Alternative to Cloud Point Extraction for Quantification of Silver Nanoparticles and Silver Speciation: Application for Nanoparticles with a Protein Corona

Production and application of nanoparticles in consumer products is at an all-time high due to the emerging field of nanotechnology. Direct detection and quantification of trace levels of nanoparticles within consumer products is very challenging and problematic. Although multiple methodologies are available for this purpose, each method has its own set of limitations. Herein, we developed an analytical platform consisting of asymmetric flow-field flow fractionation (AF4) coupled with inductively coupled plasma mass spectroscopy (ICP-MS) for the speciation and quantification of silver ions and silver nanoparticles at the ng/kg level (ppt). AF4 is utilized to concentrate the nanoparticles, and ICP-MS acts as the detector. The protein corona that forms upon exposure of nanoparticles to bovine serum albumin was utilized as a nanoparticle stabilization and AF4 recovery enhancement mechanism. Speciation of silver ions and nanoparticles was achieved with the assistance of penicillamine as a complexation ligand. The effect of nanoparticle size, surface coating, and ionization state toward the detection and quantification of the developed methodology was evaluated. The detection limit was found to be 4 ng/kg with the application of a 5 mL sample loop. Further application of this developed methodology on environmentally relevant samples was demonstrated by the analysis of Arkansas River water spiked with silver nanoparticles and nanoparticle spiked into humic acid solution (50 mg/L) at an environmentally relevant level.

Simple Functionalization Strategies for Enhancing Nanoparticle Separation and Recovery with Asymmetric Flow Field Flow Fractionation

Due to the increasing use of engineered nanomaterials in consumer products, regulatory agencies and other research organizations have determined that the development of robust, reliable, and accurate methodologies to characterize nanoparticles in complex matrices is a top priority. Of particular interest are methods that can separate and determine the size of nanomaterials in samples that contain polydisperse and/or multimodal nanoparticle populations. Asymmetric-flow field flow fractionation (AF4) has shown promise for the separation of nanoparticles with wide size range distributions; however, low analyte recoveries and decreased membrane lifetimes, due to membrane fouling, have limited its application. Herein, we report straightforward strategies to minimize membrane fouling and improve nanoparticle recovery by functionalizing the surface of the nanoparticles, as well as that of the AF4 membranes. Gold nanoparticles (AuNP) were stabilized through functionalization with a phosphine molecule, whereas the surface of the membranes was coated with a negatively charged polystyrenesulfonate polymer. Improved nanoparticle separation, recoveries of 99.1 (±0.5) %, and a detection limit of 6 μg/kg were demonstrated by analyzing AuNP reference materials of different sizes (e.g., 10, 30, and 60 nm), obtained from the National Institute of Standards and Technology (NIST). Furthermore, the stability of the polymer coating and its specificity toward minimizing membrane fouling were demonstrated.

Arsenic Speciation in Rice by Capillary Electrophoresis/Inductively Coupled Plasma Mass Spectrometry: Enzyme-Assisted Water-Phase Microwave Digestion

We report an analytical methodology for the quantification of common arsenic species in rice and rice cereal using capillary electrophoresis coupled with inductively coupled plasma mass spectrometry (CE-ICPMS). An enzyme (i.e., α-amylase)-assisted water-phase microwave extraction procedure was used to extract four common arsenic species, including dimethylarsinic acid (DMA), monomethylarsonic acid (MMA), arsenite [As(III)], and arsenate [As(V)] from the rice matrices. The addition of the enzyme α-amylase during the extraction process was necessary to reduce the sample viscosity, which subsequently increased the injection volume and enhanced the signal response. o-Arsanilic acid (o-ASA) was added to the sample solution as a mobility marker and internal standard. The obtained repeatability [i.e., relative standard deviation (RSD %)] of the four arsenic analytes of interest was less than 1.23% for elution time and 2.91% for peak area. The detection limits were determined to be 0.15-0.27 ng g(-1). Rice standard reference materials SRM 1568b and CRM 7503-a were used to validate this method. The quantitative concentrations of each organic arsenic and summed inorganic arsenic were found within 5% difference of the certified values of the two reference materials.

Detection and Characterization of SiO2 and TiO2 Nanostructures in Dietary Supplements

Nanomaterials are beginning to enter our daily lives through various consumer products as the result of technology commercialization. The development of methodologies to detect the presence of nanomaterials in consumer products is an essential element in understanding our exposure. In this study, we have developed methods for the separation and characterization of silicon dioxide (SiO2) and titanium dioxide (TiO2) nanostructures in dietary supplements marketed in products specifically targeted for women. A total of 12 commercial products claiming the inclusion of SiO2 and TiO2, but not making any claims regarding the particle size, were randomly selected for purchase through various retailers. To isolate nanostructures from these products, a simple methodology that combines acid digestion and centrifugation was utilized. Once isolated, the chemical composition, size, morphology, and crystal structure were characterized using mass spectroscopy, light scattering, electron microscopy, and X-ray diffraction techniques. SiO2 and TiO2 nanostructures were detected in 11 of 12 products using these methods. Many of the isolated nanoscale materials showed a high degree of aggregation; however, identified individual structures had at least one dimension below 100 nm. These robust methods can be used for routine monitoring of commercial products for nanoscale oxides of silica and titanium.

Rapid determination of silver in nanobased liquid dietary supplements using a portable X-ray fluorescence analyzer.

This paper reports a rapid and straightforward method for the quantitation of total Ag content in nanobased commercially available liquid dietary supplements using a portable X-ray fluorescence (pXRF) analyzer. Figures of merits were evaluated by analyzing a series of AgNO3 standards. This method was shown to have a detection limit of 3 ppm, a quantitation limit of 10 ppm, and a broad linear range from the detection limit to 10000 ppm (1%). Accurate detection and quantitation of Ag content in well-characterized Ag nanoparticle samples and in nanobased liquid dietary supplements were achieved with good correlation (i.e., percentage difference average values under 15%) between the total Ag concentration obtained by the pXRF analyzer and by inductively coupled plasma mass spectrometry (ICP-MS). Furthermore, accurate quantitation of Ag in the presence of high concentrations of potential spectral interferences was also demonstrated.

Rapid determination of plasmonic nanoparticle agglomeration status in blood

Plasmonic nanomaterials as drug delivery or bio-imaging agents are typically introduced to biological systems through intravenous administration. However, the potential for agglomeration of nanoparticles in biological systems could dramatically affect their pharmacokinetic profile and toxic potential. Development of rapid screening methods to evaluate agglomeration is urgently needed to monitor the physical nature of nanoparticles as they are introduced into blood. Here, we establish novel methods using darkfield microscopy with hyperspectral detection (hsDFM), single particle inductively-coupled plasma mass spectrometry (spICP-MS), and confocal Raman microscopy (cRM) to discriminate gold nanoparticles (AuNPs) and their agglomerates in blood. Rich information about nanoparticle agglomeration in situ is provided by hsDFM monitoring of the plasmon resonance of primary nanoparticles and their agglomerates in whole blood; cRM is an effective complement to hsDFM to detect AuNP agglomerates in minimally manipulated samples. The AuNPs and the particle agglomerates were further distinguished in blood for the first time by quantification of particle mass using spICP-MS with excellent sensitivity and specificity. Furthermore, the agglomeration status of synthesized and commercial NPs incubated in blood was successfully assessed using the developed methods. Together, these complementary methods enable rapid determination of the agglomeration status of plasmonic nanomaterials in biological systems, specifically blood.

Capillary electrophoresis coupled with inductively coupled mass spectrometry as an alternative to cloud point extraction based methods for rapid quantification of silver ions and surface coated silver nanoparticles

Speciation and accurate quantification of ionic silver and metallic silver nanoparticles are critical to investigate silver toxicity and to determine the shelf-life of products that contain nano silver under various storage conditions. We developed a rapid method for quantification of silver ions and silver nanoparticles using capillary electrophoresis (CE) interfaced with inductively-coupled plasma mass spectrometry (ICPMS). The addition of 2-mercaptopropionylglycine (tiopronin) to the background electrolyte was used to facilitate the chromatographic separation of ionic silver and maintain the oxidation state of silver. The obtained limits of detection were 0.05 μg kg(-1) of silver nanoparticles and 0.03 μg kg(-1) of ionic silver. Nanoparticles of varied sizes (10-110 nm) with different surface coating, including citrate acid, lipoic acid, polyvinylpyrrolidone and bovine serum albumin (BSA) were successfully analyzed. Particularly good recoveries (>93%) were obtained for both ionic silver and silver nanoparticle in the presence of excess amount of BSA. The method was further tested with six commercially available dietary supplements which varied in concentration and matrix components. The summed values of silver ions and silver nanoparticles correlated well with the total silver concentration determined by ICPMS after acid digestion. This method can serve as an alternative to cloud point extraction technique when the extraction efficiency for protein coated nanoparticles is low.

Polydopamine-Coated Manganese Complex/Graphene Nanocomposite for Enhanced Electrocatalytic Activity Towards Oxygen Reduction.

Platinum electrodes are commonly used electrocatalysts for oxygen reduction reactions (ORR) in fuel cells. However, this material is not economical due to its high cost and scarcity. We prepared an Mn(III) catalyst supported on graphene and further coated with polydopamine, resulting in superior ORR activity compared to the uncoated PDA structures. During ORR, a peak potential at 0.433 V was recorded, which is a significant shift compared to the uncoated material’s −0.303 V (both versus SHE). All the materials reduced oxygen in a wide pH range via a four-electron pathway. Rotating disk electrode and rotating ring disk electrode studies of the polydopamine-coated material revealed ORR occurring via 4.14 and 4.00 electrons, respectively. A rate constant of 6.33 × 106 mol−1s−1 was observed for the polydopamine-coated material–over 4.5 times greater than the uncoated nanocomposite and superior to those reported for similar carbon-supported metal catalysts. Simply integrating an inexpensive bioinspired polymer coating onto the Mn-graphene nanocomposite increased ORR performance significantly, with a peak potential shift of over +730 mV. This indicates that the material can reduce oxygen at a higher rate but with lower energy usage, revealing its excellent potential as an ORR electrocatalyst in fuel cells.

MicroRNA-152-mediated dysregulation of hepatic transferrin receptor 1 in liver carcinogenesis

Over-expression of transferrin receptor 1 (TFRC) is observed in hepatocellular carcinoma (HCC); however, there is a lack of conclusive information regarding the mechanisms of this dysregulation. In the present study, we demonstrated a significant increase in the levels of TFRC mRNA and protein in preneoplastic livers from relevant experimental models of human hepatocarcinogenesis and in human HCC cells. Additionally, using the TCGA database, we demonstrated an over-expression of TFRC in human HCC tissue samples and a markedly decreased level of microRNA-152 (miR-152) when compared to non-tumor liver tissue. The results indicated that the increase in levels of TFRC in human HCC cells and human HCC tissue samples may be attributed, in part, to a post-transcriptional mechanism mediated by a down-regulation of miR-152. This was evidenced by a strong inverse correlation between the level of TFRC and the expression of miR-152 in human HCC cells (r = −0.99, p = 4. 7 × 10−9), and was confirmed by in vitro experiments showing that transfection of human HCC cell lines with miR-152 effectively suppressed TFRC expression. This suggests that miR-152-specific targeting of TFRC may provide a selective anticancer therapeutic approach for the treatment of HCC.