Reshan A. Fernando

Exposure to Hexavalent Chromium Resulted in Significantly Higher Tissue Chromium Burden Compared With Trivalent Chromium Following Similar Oral Doses to Male F344/N Rats and Female B6C3F1 Mice

In National Toxicology Program 2-year studies, hexavalent chromium [Cr(VI)] administered in drinking water was clearly carcinogenic in male and female rats and mice, resulting in small intestine epithelial neoplasms in mice at a dose equivalent to or within an order of magnitude of human doses that could result from consumption of chromium-contaminated drinking water, assuming that dose scales by body weight(3/4) (body weight raised to the 3/4 power). In contrast, exposure to trivalent chromium [Cr(III)] at much higher concentrations may have been carcinogenic in male rats but was not carcinogenic in mice or female rats. As part of these studies, total chromium was measured in tissues and excreta of additional groups of male rats and female mice. These data were used to infer the uptake and distribution of Cr(VI) because Cr(VI) is reduced to Cr(III) in vivo, and no methods are available to speciate tissue chromium. Comparable external doses resulted in much higher tissue chromium concentrations following exposure to Cr(VI) compared with Cr(III), indicating that a portion of the Cr(VI) escaped gastric reduction and was distributed systemically. Linear or supralinear dose responses of total chromium in tissues were observed following exposure to Cr(VI), indicating that these exposures did not saturate gastric reduction capacity. When Cr(VI) exposure was normalized to ingested dose, chromium concentrations in the liver and glandular stomach were higher in mice, whereas kidney concentrations were higher in rats. In vitro studies demonstrated that Cr(VI), but not Cr(III), is a substrate of the sodium/sulfate cotransporter, providing a partial explanation for the greater absorption of Cr(VI).

Development, validation, and application of an ultra-performance liquid chromatography–sector field inductively coupled plasma mass spectrometry method for simultaneous determination of six organotin compounds in human serum

Organotin compounds (OTCs) are heavily employed by industry for a wide variety of applications, including the production of plastics and as biocides. Reports of environmental prevalence, differential toxicity between OTCs, and poorly characterized human exposure have fueled the demand for sensitive, selective speciation methods. The objective of this investigation was to develop and validate a rapid, sensitive, and selective analytical method for the simultaneous determination of a suite of organotin compounds, including butyl (mono-, di-, and tri-substituted) and phenyl (mono-, di-, and tri-substituted) species in human serum. The analytical method utilized ultra-performance liquid chromatography (UPLC) coupled with sector field inductively coupled plasma mass spectrometry (SF-ICP-MS). The small (sub-2 µm) particle size of the UPLC column stationary phase and the sensitivity of the SF-ICP-MS enabled separation and sensitive determination of the analyte suite with a runtime of approximately 3 min. Validation activities included demonstration of method linearity over the concentration range of approximately 0.250-13.661 ng mL(-1), depending on the species; intraday precision of less than 21%, interday precision of less than 18%, intraday accuracy of -5.3% to 19%, and interday accuracy of -14% to 15% for all species; specificity, and matrix impact. In addition, sensitivity, and analyte stability under different storage scenarios were evaluated. Analyte stability was found to be limited for most species in freezer, refrigerator, and freeze-thaw conditions. The validated method was then applied for the determination of the OTCs in human serum samples from women participating in the Snart-Foraeldre/MiljØ (Soon-Parents/Environment) Study. The concentration of each OTC ranged from below the experimental limit of quantitation to 10.929 ng tin (Sn) mL(-1) serum. Speciation values were confirmed by a total Sn analysis.

Development and validation of a high-throughput method for the determination of titanium dioxide in rodent lung and lung-associated lymph node tissues

Abstract There is a need to determine the tissue burdens of inhaled titanium dioxide due to the widespread industrial use of this compound. A high-throughput method for the determination of TiO2 as titanium in rodent lung and lung-associated lymph node tissues by inductively coupled plasma optical emission spectrometry (ICP-OES) was developed and validated. Samples were digested in open-vessel, disposable centrifuge tubes in the presence of nitric and hydrofluoric acids. The determined method quantitation limit (MQL) was 4.2 µg TiO2/g tissue. Replicate control samples prepared at this level were determined with a high level of accuracy (100%) and precision (2% RSD). After validation, the described method was applied to determine the concentration of TiO2 (as titanium) in 600 rodent tissue samples spanning a 16-month time period. During this time period, laboratory control samples (n = 48) were analyzed to assess the long-term precision and accuracy of the method. The average recovery of titanium in these samples was 102±4.8%.

Validation of a Method for the Determination of Total Chromium in Rat Feces by Inductively Coupled Plasma Optical Emission Spectrometry

Abstract The validation of a method for the determination of total chromium in Fischer-344 rat feces by inductively coupled plasma optical emission spectrometry following a rapid, atmospheric-pressure microwave digestion is described. The performance of the method was evaluated over the concentration range of 5.00 to 200 µg Cr/g feces. Data for method linearity, accuracy, precision, digest stability, and storage stability are presented along with limit of detection and limit of quantitation data. Data from a cross-validation method for B6C3F1 mouse feces are also presented. Following validation, the method was applied to analyze samples collected in support of two chronic toxicological investigations.

Trace Element Content of Senna Study Material and Selected Senna-Based Dietary Supplements as Determined by Inductively Coupled Plasma-Optical Emission Spectrometry and Inductively Coupled Plasma-Mass Spectrometry

Abstract The efficacy of senna laxative products and the worldwide trend towards self-medication with dietary supplements of a natural origin have fueled several investigations into the potential health effects associated with their use. The goal of this work was to determine the trace element content [aluminum (Al), arsenic (As), barium (Ba), calcium (Ca), cadmium (Cd), chromium (Cr), copper (Cu), iron (Fe), magnesium (Mg), manganese (Mn), sodium (Na), lead (Pb), strontium (Sr), vanadium (V), and zinc (Zn)] of a bulk senna sample that was procured for use as a study material in one such investigation. In addition, seven commercially available senna-based dietary supplements were purchased and analyzed along with the study material. Samples were digested using high-purity acids and oxidants and were analyzed by inductively coupled plasma-optical emission spectrometry (ICP-OES) and inductively coupled plasma-mass spectrometry (ICP-MS). Differences between the trace element content of the analyzed products were observed. Lead was detected in three of the tested samples (ranging from 0.686 to 1.12 μg g−1), and arsenic was detected in seven samples (ranging from 0.063 to 0.280 μg g−1). Recovery data for fortified control samples and standard reference materials (NIST 1573a Tomato Leaves, NIST 1547 Peach Leaves) indicated accurate and precise method performance.

DEVELOPMENT AND VALIDATION OF A METHOD FOR THE DETERMINATION OF MERCURY IN SMALL RAT BRAIN TISSUE SAMPLES BY COLD VAPOR ATOMIC FLUORESCENCE SPECTROMETRY

ABSTRACT A rugged, sensitive, semi-automated method for the determination of trace-level mercury in small rat brain tissue samples by cold vapor atomic fluorescence spectrometry (CVAFS) is described. Small tissue aliquots (100 mg) are digested for 6–8 h at an elevated temperature in a mixture of nitric and sulfuric acids. Following digestion, samples are oxidized with potassium bromate/bromide at room temperature and treated with a continuous flow of stannous chloride to generate mercury vapor. The experimentally determined method quantitation limit (MQL) in the rat brain matrix is 10 pg Hg/mL, which corresponds to 1.5 ng Hg/g brain tissue. Replicate matrix spike preparations (n = 4) at this level were determined with a high level of accuracy (89.8%) and precision (6.9% RSD). Replicates (n = 36) of a certified reference material (DOLT-2, trace metals in dogfish liver) were prepared and analyzed over a four year period in order to assess the ruggedness and accuracy of the method. The average recovery for mercury in this material was 99.9% of the certified level.

Development and Optimization of a Procedure for the Determination of Indium-Tin Oxide Particle Size and Concentration in Cellular Media

Indium-tin oxide (ITO) is a solid mixture often comprised of approximately 90% indium oxide and 10% tin oxide by weight. ITO is employed as a transparent conductive coating for flat panel, liquid crystal, and plasma displays and is typically deposited after sintering as a thin film on a substrate through a variety of processes. Unfortunately, there have been published reports of workers who have been adversely impacted through exposure to this compound. When these reports are coupled with the industrial utility of ITO, future toxicological investigations are likely. Comprehensive characterization of ITO test materials would be an essential component of these studies. Ideally, characterization would include assessment of ITO particle size as administered during dosing because this physicochemical parameter can significantly impact solubility. The objective of this task was to develop, optimize, and apply a dynamic light scattering (DLS) sample preparation and measurement protocol for determination of particle size for a suite of sintered/non-sintered ITO samples prepared in cellular growth media at nominal 0.3 mg/mL doses. Sonication time was evaluated from 15 to 90 minutes over a 24-hour, continuous DLS measurement period. Resulting suspensions from samples subjected to 30 minute sonication times were stable with respect to hydrodynamic particle size between 10 and 24 hours after sonication. Throughout the 24-hour DLS measurement period, suspension aliquots were analyzed by inductively coupled plasma mass spectrometry to determine indium concentration. The developed protocol enabled investigators to use ITO suspensions of known hydrodynamic particle size and concentration in their in vitro cellular study.

Characterization of Zinc Carbonate Basic as a Source of Zinc in a Rodent Study Investigating the Effects of Dietary Deficiency or Excess

ABSTRACT Zinc deficiency and excess can result in adverse health outcomes. There is conflicting evidence regarding whether excess or deficient zinc in the diet can contribute to carcinogenicity. The objective of this study was to characterize zinc carbonate basic for use as a source of dietary zinc in a rodent toxicity and carcinogenicity study investigating the effects of zinc deficiency and excess. Because of the complex chemistries of zinc carbonate basic compounds, inconsistent nomenclature, and literature and reference spectra gaps, it was necessary to use multiple analytical techniques, including Karl Fischer titration, combustion analysis, inductively coupled plasma–optical emission spectrometry, X-ray diffraction, infrared spectroscopy, X-ray fluorescence spectrometry, and thermogravimetric analysis to characterize the test article. Based on the collective evidence and through the process of elimination, the test article was found to be composed mainly of zinc carbonate basic with zinc oxide as a minor component. The zinc content was determined to be 56.6% (w/w) with heavy metals such as arsenic, cadmium, mercury, and lead below the limit of quantitation of less than or equal to 0.01%. The test material was stable at ambient temperature. Based on the work described in this manuscript, the test article was suitable for use as a source of zinc in studies of deficiency and excess in the diet.