Bryant C. Nelson

Copper Oxide Nanoparticle Mediated DNA Damage in Terrestrial Plant Models

Engineered nanoparticles, due to their unique electrical, mechanical, and catalytic properties, are presently found in many commercial products and will be intentionally or inadvertently released at increasing concentrations into the natural environment. Metal- and metal oxide-based nanomaterials have been shown to act as mediators of DNA damage in mammalian cells, organisms, and even in bacteria, but the molecular mechanisms through which this occurs are poorly understood. For the first time, we report that copper oxide nanoparticles induce DNA damage in agricultural and grassland plants. Significant accumulation of oxidatively modified, mutagenic DNA lesions (7,8-dihydro-8-oxoguanine; 2,6-diamino-4-hydroxy-5-formamidopyrimidine; 4,6-diamino-5-formamidopyrimidine) and strong plant growth inhibition were observed for radish (Raphanus sativus), perennial ryegrass (Lolium perenne), and annual ryegrass (Lolium rigidum) under controlled laboratory conditions. Lesion accumulation levels mediated by copper ions and macroscale copper particles were measured in tandem to clarify the mechanisms of DNA damage. To our knowledge, this is the first evidence of multiple DNA lesion formation and accumulation in plants. These findings provide impetus for future investigations on nanoparticle-mediated DNA damage and repair mechanisms in plants.

Selection of Column and Gradient Elution System for the Separation of Catechins in Green Tea Using High-Performance Liquid Chromatography

A study of a variety of stationary phases and elution conditions for the liquid chromatographic (LC) determination of six biologically active green tea catechins has resulted in the development of two well-defined, reproducible systems for such analyses which overcome limitations of previously described methods. Comparison of six reversed-phase columns indicates that deactivated stationary phases, which utilize ultrapure silica and maximize coverage of the silica support, provide significantly improved separation and chromatographic efficiencies for catechin analyses using LC, compared to conventional monomeric or polymeric C18 columns. Evaluation of elution conditions used for the separations reveals that the presence of acid in the mobile phase (0.05% trifluoroacetic acid) is essential for both the complete resolution of the catechins present in tea and the efficient chromatography of these compounds. The efficacy of one of the developed systems was demonstrated by the quantitative measurement of the six biologically active catechins in aqueous infusions of green tea (Camellia sinensis). Overall precision values for the analyses were within the range 0.3-1% (relative standard deviation).

Determination of tea catechins

An overview of analytical methods for the measurement of biologically important tea catechins is presented. Liquid chromatography and capillary electrophoresis are the most cited techniques for catechin separation, identification and quantitation. Liquid chromatography with ultraviolet detection is frequently used; however, mass spectrometry, electrochemical, fluorescence and chemiluminescence detection are also utilized in cases where more sensitive or selective detection is needed. Two modes of capillary electrophoresis, capillary zone electrophoresis and micellar electrokinetic capillary chromatography, have been employed for the determination of catechins. Both modes of capillary electrophoresis are based on ultraviolet detection. Additional analytical techniques, such as gas chromatography, thin-layer chromatography, paper chromatography, spectrophotometry, biosensing, chemiluminescence and nuclear magnetic resonance spectroscopy have also been utilized for the determination of catechins and are reviewed herein.

Mechanisms and measurements of nanomaterial-induced oxidative damage to DNA

Many of the current investigations on the environmental and human health risks of engineered nanomaterials focus on their short-term acute toxicity. However, the long-term chronic effects of nanomaterials on living systems, and in particular, on the genetic components of living systems, also warrant attention. An increasing number of nanomaterial safety studies include an assessment of genotoxicity as part of the overall risk evaluation. The potential of nanomaterials to directly or indirectly promote the formation of reactive oxygen species is one of the primary steps in their genotoxic repertoire. The subsequent modification of genomic DNA by reactive oxygen species could lead to the development of mutagenesis, carcinogenesis, or other age-related diseases if the DNA damage is not repaired. This review focuses on the interactions of nanomaterials with DNA and specifically on the capacity of some nanomaterials to induce oxidative damage to DNA. A critical assessment of the analytical methodology and the potential biochemical mechanisms involved in nanomaterial induction of oxidative damage to DNA is presented, results obtained for the various studies with each nanomaterial are compared, and recommendations for future research are discussed. Researchers should consider, among other experimental recommendations, (1) the application of more chromatography-based and mass-spectrometry-based analytical techniques to the assessment of oxidative damage to DNA to facilitate an enhanced understanding of DNA damage mechanisms and (2) the verification of cellular viability before conducting genotoxicity assays to reduce the impact of fragmented DNA, formed as a consequence of cell death, on DNA damage measurements.

The role of iron redox state in the genotoxicity of ultrafine superparamagnetic iron oxide nanoparticles.

Ultrafine superparamagnetic iron oxide nanoparticles (USPION) hold great potential for revolutionising biomedical applications such as MRI, localised hyperthermia, and targeted drug delivery. Though evidence is increasing regarding the influence of nanoparticle physico-chemical features on toxicity, data however, is lacking that assesses a range of such characteristics in parallel. We show that iron redox state, a subtle though important physico-chemical feature of USPION, dramatically modifies the cellular uptake of these nanoparticles and influences their induction of DNA damage. Surface chemistry was also found to have an impact and evidence to support a potential mechanism of oxidative DNA damage behind the observed responses has been demonstrated. As human exposure to ferrofluids is predicted to increase through nanomedicine based therapeutics, these findings are important in guiding the fabrication of USPION to ensure they have characteristics that support biocompatibility.

Separation of blackcurrant anthocyanins by capillary zone electrophoresis

The four major anthocyanins present in juice of the blackcurrant (Ribes nigrum) may be completely separated by capillary zone electrophoresis under strongly acidic conditions. The separation, resolution and peak shapes of the anthocyanins are critically influenced by the pH of the running buffer and the presence of an organic solvent. Fused-silica and polyacrylamide-coated capillary columns were evaluated for their ability to resolve the closely migrating analytes. Optimum qualitative separation was achieved on a fused-silica capillary with a phosphate running buffer containing 30% (v/v) acetonitrile at an apparent pH of 1.5.

Antioxidant Cerium Oxide Nanoparticles in Biology and Medicine.

Previously, catalytic cerium oxide nanoparticles (CNPs, nanoceria, CeO2-x NPs) have been widely utilized for chemical mechanical planarization in the semiconductor industry and for reducing harmful emissions and improving fuel combustion efficiency in the automobile industry. Researchers are now harnessing the catalytic repertoire of CNPs to develop potential new treatment modalities for both oxidative- and nitrosative-stress induced disorders and diseases. In order to reach the point where our experimental understanding of the antioxidant activity of CNPs can be translated into useful therapeutics in the clinic, it is necessary to evaluate the most current evidence that supports CNP antioxidant activity in biological systems. Accordingly, the aims of this review are three-fold: (1) To describe the putative reaction mechanisms and physicochemical surface properties that enable CNPs to both scavenge reactive oxygen species (ROS) and to act as antioxidant enzyme-like mimetics in solution; (2) To provide an overview, with commentary, regarding the most robust design and synthesis pathways for preparing CNPs with catalytic antioxidant activity; (3) To provide the reader with the most up-to-date in vitro and in vivo experimental evidence supporting the ROS-scavenging potential of CNPs in biology and medicine.

Development of a Standard Reference Material for Metabolomics Research

The National Institute of Standards and Technology (NIST), in collaboration with the National Institutes of Health (NIH), has developed a Standard Reference Material (SRM) to support technology development in metabolomics research. SRM 1950 Metabolites in Human Plasma is intended to have metabolite concentrations that are representative of those found in adult human plasma. The plasma used in the preparation of SRM 1950 was collected from both male and female donors, and donor ethnicity targets were selected based upon the ethnic makeup of the U.S. population. Metabolomics research is diverse in terms of both instrumentation and scientific goals. This SRM was designed to apply broadly to the field, not toward specific applications. Therefore, concentrations of approximately 100 analytes, including amino acids, fatty acids, trace elements, vitamins, hormones, selenoproteins, clinical markers, and perfluorinated compounds (PFCs), were determined. Value assignment measurements were performed by NIST and the Centers for Disease Control and Prevention (CDC). SRM 1950 is the first reference material developed specifically for metabolomics research.

Manganese-induced oxidative DNA damage in neuronal SH-SY5Y cells: Attenuation of thymine base lesions by glutathione and N-acetylcysteine

Manganese (Mn) is an essential trace element required for normal function and development. However, exposure to this metal at elevated levels may cause manganism, a progressive neurodegenerative disorder with neurological symptoms similar to idiopathic Parkinsons disease (IPD). Elevated body burdens of Mn from exposure to parental nutrition, vapors in mines and smelters and welding fumes have been associated with neurological health concerns. The underlying mechanism of Mn neurotoxicity remains unclear. Accordingly, the present study was designed to investigate the toxic effects of Mn(2+) in human neuroblastoma SH-SY5Y cells. Mn(2+) caused a concentration dependent decrease in SH-SY5Y cellular viability compared to controls. The LD50 value was 12.98 μM Mn(2+) (p<0.001 for control vs. 24h Mn treatment). Both TUNEL and annexin V/propidium iodide (PI) apoptosis assays confirmed the induction of apoptosis in the cells following exposure to Mn(2+) (2 μM, 62 μM or 125 μM). In addition, Mn(2+) induced both the formation and accumulation of DNA single strand breaks (via alkaline comet assay analysis) and oxidatively modified thymine bases (via gas chromatography/mass spectrometry analysis). Pre-incubation of the cells with characteristic antioxidants, either 1mM N-acetylcysteine (NAC) or 1mM glutathione (GSH) reduced the level of DNA strand breaks and the formation of thymine base lesions, suggesting protection against oxidative cellular damage. Our findings indicate that (1) exposure of SH-SY5Y cells to Mn promotes both the formation and accumulation of oxidative DNA damage, (2) SH-SY5Y cells with accumulated DNA damage are more likely to die via an apoptotic pathway and (3) the accumulated levels of DNA damage can be abrogated by the addition of exogenous chemical antioxidants. This is the first known report of Mn(2+)-induction and antioxidant protection of thymine lesions in this SH-SY5Y cell line and contributes new information to the potential use of antioxidants as a therapeutic strategy for protection against Mn(2+)-induced oxidative DNA damage.

Capillary liquid chromatography/electrospray mass spectrometry for the separation and detection of catechins in green tea and human plasma

The separation and detection of biologically active green tea catechins has been accomplished using capillary liquid chromatography/electrospray mass spectrometry (cLC/ESI-MS). Microscale determination (approximately 20 ng) of all six catechins in a green tea infusion, and the most extensively studied catechin, (-)epigallocatechin gallate (EGCG), in human plasma is demonstrated by cLC/ESI-MS with selected ion monitoring of protonated molecular ions. The overall quality of the analysis is shown to be dependent on the use of a capillary column with a deactivated, monomeric C18 stationary phase. The high chromatographic separation efficiency of this packed-capillary column, combined with the high sensitivity and selectivity afforded by the mass spectrometer as detector, provide a reliable approach to the analysis of picomolar quantities of these interesting compounds in complex matrices.