Steven J. Christopher

An accurate and sensitive method for the determination of methylmercury in biological specimens using GC-ICP-MS with solid phase microextraction

A highly sensitive and selective method has been developed for the determination of methylmercury in biological specimens and NIST Standard Reference Materials (SRMs). The procedure involves microwave extraction with acetic acid, followed by derivatization and headspace solid-phase microextraction (SPME) with a polydimethylsiloxane (PDMS)-coated silica fiber. Optimization of conditions including gas chromatograph injection temperature, microwave extraction power and microwave extraction time are presented. The identification and quantification (via the method of standard additions) of the extracted compounds is carried out by capillary gas chromatography with inductively coupled plasma mass spectrometric detection (GC-ICP-MS) using a unique heated interface that was designed for this work. The SPME-GC-ICP-MS method was validated for the determination of methylmercury (MeHg) concentrations in a variety of biological Standard Reference Materials (SRMs), ranging from 13.2 ng g−1 in SRM 1566b Oyster Tissue, to 397 ng g−1 in SRM 1946 Lake Superior Fish Tissue. Additionally, this method was applied to the determination of MeHg in seabird eggs (common murres, Uria aalge and thick-billed murres, Uria lomvia) collected from colonies on Little Diomede and Bogoslof islands in the Bering Sea and Saint Lazaria Island in the Gulf of Alaska and cryogenically banked in the Marine Environmental Specimen Bank. The results obtained demonstrate that SPME-GC-ICP-MS is a sensitive technique for the determination of methylmercury at trace and ultra-trace levels in a variety of natural matrices with high reproducibility and accuracy. In all instances, the sample-to-sample variability was typically 2% relative standard deviation (RSD) and the method detection limit for methylmercury was 4.2 pg g−1 (as Hg), based on a 0.5 g tissue sample of SRM 1566b Oyster Tissue.

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.

Diamondback terrapins, Malaclemys terrapin, as a sentinel species for monitoring mercury pollution of estuarine systems in South Carolina and Georgia, USA

Total mercury concentrations were measured in diamondback terrapin blood and scutes collected from four sites in South Carolina, USA, and at a superfund site in Brunswick, Georgia, USA. There was a strong correlation between mercury concentrations in the two terrapin body compartments (Kendalls tau = 0.79, p < 0.001). Mercury concentrations in terrapin scute and blood and in salt marsh periwinkles, Littoraria irrorata, were significantly higher in Brunswick (scute x = 3810.2 ng/g, blood x = 746.2 ng/g) than from all other sites (scute x = 309.5 ng/g, blood x = 43.2 ng/g, p < 0.001). Seasonal fluctuations of total mercury in the blood and scutes of terrapins collected in the Ashley River, South Carolina, were significantly lower in August than in April, June, or October in blood (p < 0.001); however, scute concentrations did not vary seasonally. Overall, we found higher concentrations of mercury in the scutes of females than males (n = 32, p < 0.05). Larger females may preferentially prey on larger food items, like large periwinkles, which had significantly higher mercury levels in their body tissues than smaller periwinkles (p < 0.001). Methylmercury levels in terrapin scutes were measured, revealing that 90% of the total mercury stored in this compartment was in the organic form. A methylmercury biomagnification factor of 173.5 was calculated from snails to terrapin scutes, and we found that mercury levels in scutes were representative of the mercury levels in other compartments of the ecosystem. These findings show that terrapin scutes are good predictors of mercury pollution and that this species could be used as a bioindicator for assessing mercury contamination of estuarine systems.

Development and certification of green tea-containing standard reference materials.

AbstractA suite of three green tea-containing Standard Reference Materials (SRMs) has been issued by the National Institute of Standards and Technology (NIST): SRM 3254 Camellia sinensis (Green Tea) Leaves, SRM 3255 Camellia sinensis (Green Tea) Extract, and SRM 3256 Green Tea-Containing Solid Oral Dosage Form. The materials are characterized for catechins, xanthine alkaloids, theanine, and toxic elements. As many as five methods were used in assigning certified and reference values to the constituents, with measurements carried out at NIST and at collaborating laboratories. The materials are intended for use in the development and validation of new analytical methods, and for use as control materials as a component in the support of claims of metrological traceability. FigureGreen Tea - Camellia sinensis

Consideration and influence of complexed forms of mercury species on the reactivity patterns determined by speciated isotope dilution model approaches: A case for natural biological reference materials

The processes driving the inadvertent transformations of inorganic mercury (iHg) and methylmercury (MeHg) in cryogenically homogenized fresh-frozen (FF) and freeze-dried (FD) biological standard reference materials (SRM) were investigated using alkaline digestion, derivatization and gas chromatography inductively coupled plasma mass spectrometry (GC-ICP-MS). Labile inorganic mercury (201iHg) and methylmercury (Me202Hg) isotopic standards and their cysteine-complexed analogs (201Hg(Cys)2 and Me202HgCys) were used in a double-spike speciated isotope dilution (SID) model to document the influence of complexing ligands on the equilibration, the reactivity and the transformation processes between isotopic mercury species and their endogenous analogs. Cysteine-complexed and labile Me202Hg spiking standards displayed similar equilibration processes in both classes of materials, leading to accurate MeHg determinations with negligible methylation transformations. Labile 201iHg standards provided accurate iHg concentration results in both material series, although an apparent demethylation reaction specific of FF materials was detected, with a negligible effect in FD materials. Cysteine-complexed 201iHg standards led to higher but inaccurate iHg concentrations and higher demethylation yields in both materials. This comparison illustrated a significant influence of complexing ligands on the equilibration processes between labile and/or complexed 201iHg species and their endogenous analogs. The derivatization step was found to catalyze these non-equilibrium conditions with different derivatization yields between labile and complexed iHg species, whereas no differences were observed for MeHg species. When complexation disparities existed in solution between 201iHg species and their endogenous analogs, this process specifically affected the determination of 200/201iHg ratios relative to 200/202iHg, 200/201MeHg and 200/202iHg ratios, which are all involved in the double-spike SID model to establish transformation yields and mercury species concentrations. This process and the influence of demethylating agents were presumably responsible for the apparent demethylation reaction only observed in FF materials. This result gave rise to several questions on the influence of freeze-drying procedures on the lability/complexation patterns of mercury species and/or on the activity of demethylating agents. A complementary analytical step consisting of maximizing the equilibrium conditions between isotopic and endogenous mercury species together with inhibiting the activity of demethylating agent is proposed. When applying these conditions, the two classes of materials displayed negligible transformation reactions, improving their commutability for the simultaneous determination of iHg and MeHg. The approach proposed in this work allowed for the first time documentation of the potential and limitations of speciated isotope dilution procedures to handle the problem of lability/complexation and ligand interactions in natural biological matrices when considering multiple-spike SID approaches.

Improved Calibration Strategy for Measurement of Trace Elements in Biological and Clinical Whole Blood Reference Materials via Collision-Cell Inductively Coupled Plasma Mass Spectrometry

A multi-element quantification strategy based on the method of standard additions incorporating internal standards and collision cell inductively coupled plasma mass spectrometry (ICP-MS) is presented. Approaches to experimental design are discussed in the context of streamlining analytical measurement protocols employing ratio-based standard additions quantification schemes for the certification of multiple elements in Certified Reference Materials, including reduction of the number of required analytical samples and measurement of analytes and internal standards at alternate quadrupole mass resolution settings. This strategy was implemented for the measurement of As, Se, Fe, Mn, Rb, Cu, and Zn levels in a candidate fish tissue NIST Standard Reference Material and measurement of Cd and Pb in two clinical, whole blood Certified Reference Materials. A simple approach to calculating analytical uncertainties for concentration data, as determined using standard additions calibrations, is presented which utilizes regression and prediction uncertainties and quotient propagation of error formulae.

Novel separation for the determination of cadmium by isotope dilution ICP-MS in samples containing high concentrations of molybdenum and tin

A four-step matrix reduction procedure, beginning with thiourea-based solid phase extraction, was developed for the determination of cadmium in difficult sample matrixes by inductively coupled plasma mass spectrometry. The separation scheme reduced the concentrations of tin and molybdenum, significant spectral interference sources for cadmium, from mg kg−1 levels to μg kg−1 and sub-μg kg−1 levels respectively, making possible interference-free measurement of cadmium and facilitating isotope dilution quantification. Cadmium recovery through the matrix reduction procedure was 78%. The effectiveness of the method was shown by application to NIST SRMs 1568a Rice Flour and 2703 Marine Sediment.

Determination of trace sulfur in biodiesel and diesel standard reference materials by isotope dilution sector field inductively coupled plasma mass spectrometry.

A method is described for quantification of sulfur at low concentrations on the order of mgkg(-1) in biodiesel and diesel fuels using isotope dilution and sector field inductively coupled plasma mass spectrometry (ID-SF-ICP-MS). Closed vessel microwave-assisted digestion was employed using a diluted nitric acid and hydrogen peroxide decomposition medium to reduce sample dilution volumes. Medium resolution mode was employed to eliminate isobaric interferences at (32)S and (34)S related to polyatomic phosphorus and oxygen species, and sulfur hydride species. The method outlined yielded respective limits of detection (LOD) and limits of quantification (LOQ) of 0.7 mg kg(-1) S and 2.5 mg kg(-1) S (in the sample). The LOD was constrained by instrument background counts at (32)S but was sufficient to facilitate value assignment of total S mass fraction in NIST SRM 2723b Sulfur in Diesel Fuel Oil at 9.06±0.13 mg kg(-1). No statistically significant difference at a 95% confidence level was observed between the measured and certified values for certified reference materials NIST SRM 2773 B100 Biodiesel (Animal-Based), CENAM DRM 272b and NIST SRM 2723a Sulfur in Diesel Fuel Oil, validating method accuracy.

Determination of trace level cadmium in SRM 3280 Multivitamin/Multielement Tablets via isotope dilution inductively coupled plasma mass spectrometry

Cadmium was quantified at 80.15±0.86 ng/g (mean±95% expanded uncertainty) in NIST SRM 3280 Multivitamin/Multielement Tablets, using isotope dilution mass spectrometry. The method described utilized various precipitation and solid-phase extraction separation methodologies to isolate Cd from Sn and Mo, present respectively, at 11.1±0.9 mg/kg and 70.7±4.5 mg/kg in the tablet matrix. This allowed for measurement of (111)Cd/(113)Cd and (111)Cd/(114)Cd isotope ratios using both quadrupole collision cell technology inductively coupled plasma mass spectrometry (Q-CCT-ICP-MS) and sector field (SF)-ICP-MS equipped with a desolvating nebulizer system to mitigate the MoO(+) and MoOH(+) molecular ion interferences that typically affect the envelope of Cd isotopes.

Influence of mercury and selenium chemistries on the progression of cardiomyopathy in pygmy sperm whales, Kogia breviceps.

More than half of pygmy sperm whales (Kogia breviceps) that strand exhibit signs of cardiomyopathy (CMP). Many factors may contribute to the development of idiopathic CMP in K. breviceps, including genetics, infectious agents, contaminants, biotoxins, and dietary intake (e.g. selenium, mercury, and pro-oxidants). This study assessed trace elements in K. breviceps at various stages of CMP progression using fresh frozen liver and heart samples collected from individuals that stranded along US Atlantic and Gulf coasts between 1993 and 2007. Standard addition calibration and collision cell inductively coupled plasma mass spectrometry (ICP-MS) were employed for total Se analysis and pyrolysis atomic absorption (AA) was utilized for total Hg analysis to examine if the Se/Hg detoxification pathway inhibits the bioavailability of Se. Double spike speciated isotope dilution gas chromatography ICP-MS was utilized to measure methyl Hg and inorganic Hg. Immunoblot detection and colorimetric assays were used to assess protein oxidation status. Data collected on trace elements, selenoproteins, and oxidative status were evaluated in the context of animal life history and other complementary histological information to gain insight into the biochemical pathways contributing to the development of CMP in K. breviceps. Cardiomyopathy was only observed in adult pygmy sperm whales, predominantly in male animals. Both Hg:Se molar ratios and overall protein oxidation were greater in males than females and increased with progression of CMP.