Development of a kelp powder (Thallus laminariae) Standard Reference Material.

A Standard Reference Material (SRM) of seaweed, SRM 3232 Kelp Powder (Thallus laminariae) has been developed to support food and dietary supplement measurements in compliance with the Food Safety Modernization Act (FSMA) and the Dietary Supplement Health and Education Act of 1994 (DSHEA). The material was characterized for nutritional minerals, arsenic species, isomers of vitamin K1, proximates, and toxic elements. Kelp is a rich source of vitamins and minerals, and it is an excellent source of dietary iodine. Kelp also contains a large amount of arsenic, which is toxic as inorganic species but much less so as organic species. To capture the dietary profile of kelp, certified values were issued for As, Ca, Cd, Cr, Cu, Fe, Hg, I, K, Mg, Mn, Mo, Na, Pb, and Zn. Reference values for proximates were assigned. For the first time, a certified value for iodine, reference values for isomers of vitamin K1, and reference values for arsenic species including arsenosugars were assigned in a seaweed. SRM 3232 fills a gap in Certified Reference Materials (CRMs) needed for quality assurance and method validation in the compositional measurements of kelp and similar seaweeds used as food and as dietary supplements. Graphical Absract Arsenic species and isomers of vitamin K1 were determined in the development of SRM 3232 Kelp Powder (Thallus laminariae).

On neutron activation analysis with γγ coincidence spectrometry.

A new γγ coincidence system has been set up at NIST. It is operated with a digital data finder supported by new software developed at NIST. The system is used to explore possible enhancements in instrumental neutron activation analysis (INAA) and study applicability to neutron capture prompt gamma activation analysis (PGAA). The performance of the system is tested with certified reference materials for efficiency calibration and quantitative performance. Comparisons of INAA results based on conventional gamma-ray spectrometry data with INAA results based on coincidence data obtained from the same samples show improvements in the counting uncertainties and demonstrates the quantitative accuracy of the new system.

SI traceable determination of arsenic species in kelp (Thallus laminariae).

A dietary supplement, kelp contains a significant amount of arsenic that is mostly arsenosugars. The determination of arsenosugars is difficult due to a lack of arsenosugar calibration standard, because arsenosugar compounds are not commercially available. This work reports the determination of arsenicals in a kelp extract with traceability to the International System of Units (SI). The hydrophilic fraction of arsenicals was reproducibly extracted from a candidate Standard Reference Material (SRM) 3232 Kelp Powder (Thallus Laminariae) in development at the National Institute of Standards and Technology (NIST). Arsenosugars and dimethylarsinic acid (DMA) were separated into fractions using analytical liquid chromatography (LC) cation and anion columns. The arsenic in the fractions was determined using instrumental neutron activation analysis (INAA). Cation exchange separation was used for INAA determination of arsenosugar 3-[5'-deoxy-5'-(dimethylarsinoyl)-ß-ribofuranosyloxy]propylene glycol (As(328)) for the first time, while DMA, arsenosugars 3-[5'-deoxy-5'-(dimethylarsinoyl)-ß-ribofuranosyloxy]-2-hydroxypropyl 2,3-dihydroxypropyl hydrogen phosphate (As(482)), and 3-[5'-deoxy-5'-(dimethylarsinoyl)-ß-ribofuranosyloxy]-2-hydroxypropanesulfonic acid (As(392)) were determined following anion exchange separation. The contents of DMA, As(328), As(482), and As(392) were 0.41 mg kg-1 ± 0.09 mg kg-1, 1.10 mg kg-1 ± 0.25 mg kg-1, 5.23 mg kg-1 ± 0.46 mg kg-1, and 13.17 mg kg-1 ± 0.67 mg kg-1, respectively. Separately, components of arsenic species in the kelp extract including DMA, As(328), and inorganic arsenic were determined using LC-inductively coupled plasma mass spectrometry. Results of DMA and As(328) were 0.485 mg kg-1 ± 0.024 mg kg-1 and 1.14 mg kg-1 ± 0.03 mg kg-1, respectively, which were in good agreement with those determined by INAA in fractions of LC eluent. The most toxic species, AsIII and AsV were found to be < 0.07 mg kg-1 and 0.231 mg kg-1 ± 0.018 mg kg-1, respectively. Results were traceable to SI.

Assessment of PGAA capability for low-level measurements of H in Ti alloys.

The accuracy of low-level hydrogen measurements with prompt gamma-ray activation analysis (PGAA) depends on identifying and accounting for all background H signals, including interfering signals. At the cold-neutron (CN)PGAA facility at the NIST Center for Neutron Research, the sources of background H signals were investigated in the context of titanium-based matrices containing low-levels of H (<300 mg H per kg Ti) with the measurements of prepared standards (mixtures of polyvinyl chloride and titanium oxide) and Ti alloy (Ti6Al4V) samples. The sensitivity ratio, defined as the ratio of the H signal to the Ti signal per unit mass ratio of H in Ti, was determined (1) with the measurements of prepared standards and (2) based on partial gamma-ray production cross sections and full-energy detection efficiencies. The resulting calibrations from these two approaches agreed within experimental uncertainty. A series of Ti alloy NIST Standard Reference Materials (SRMs) previously certified for the H content (SRMs 2452, 2453, 2453a, 2454) were used as test cases, with the mass fractions determined based on the sensitivity ratios derived from method 1 and method 2, respectively. The results agreed with the certified values within experimental uncertainties, validating the analysis performed on the new instrument with newly-prepared standards at low H mass fractions (method 1), and with the standard-independent analysis (method 2). Various sample mounting improvements were made to lower the background H signal. Spectral interferences near the H peak were identified as potential sources of bias and as a limiting factor in the detection limit of H in Ti alloy samples.

Development of two fine particulate matter standard reference materials (<4 µm and <10 µm) for the determination of organic and inorganic constituents.

Two new Standard Reference Materials (SRMs), SRM 2786 Fine Particulate Matter (<4 µm) and SRM 2787 Fine Particulate Matter (<10 µm) have been developed in support of the US Environmental Protection Agency's National Ambient Air Quality Standards for particulate matter (PM). These materials have been characterized for the mass fractions of selected polycyclic aromatic hydrocarbons (PAHs), nitrated PAHs, brominated diphenyl ether (BDE) congeners, hexabromocyclododecane (HBCD) isomers, sugars, polychlorinated dibenzo-p-dioxin (PCDD) and dibenzofuran (PCDF) congeners, and inorganic constituents, as well as particle-size characteristics. These materials are the first Certified Reference Materials available to support measurements of both organic and inorganic constituents in fine PM. In addition, values for PAHs are available for RM 8785 Air Particulate Matter on Filter Media. As such, these SRMs will be useful as quality control samples for ensuring compatibility of results among PM monitoring studies and will fill a void to assess the accuracy of analytical methods used in these studies. Graphical Abstract Removal of PM from filter for the preparation of SRM 2786 Fine Particulate Matter.

An approach for identification and determination of arsenic species in the extract of kelp.

The National Institute of Standards and Technology is developing a kelp powder standard reference material (SRM) in support of dietary supplement measurements. Edible seaweeds such as kelp and laver consumed as diet or dietary supplement contain tens of mg/kg arsenic. The speciation information of arsenic in the seaweed should be provided because the total arsenic alone does not fully address the safety issue of the dietary supplement as the value assignment is originally intended. The inability to avail all arsenic species for value assignment measurements prevented the certification of arsenic species in the candidate SRM; however, approximately 70 % of total arsenic extracted with a 1:1 volume fraction of methanol:water mixture allowed arsenic speciation values to be assigned to a procedure-defined extract, which may be used for method validation in research to improve upon current extraction and measurement practices. Arsenic species in kelp and laver were identified using electrospray ionization ion trap time of flight mass spectrometry (ESI-IT-TOF). Arsenosugars As(328), As(482), and As(392) were found in the kelp candidate SRM while As(328) and As(482) were found in GBW 08521, a certified reference material (CRM) of laver produced by the National Institute of Metrology of China (NIM). A discovery that the digests of kelp and laver contained only dimethylarsinic acid led to the conclusion that the seaweeds did not contain detectible levels of arsenobetaine, arsenocholine or trimethylarsine oxide that could overlap with the peaks of arsenosugars in the separation. The mean ± s of (5.68 ± 0.28) mg/kg and (13.43 ± 0.31) mg/kg found for As(482) and As(392) in kelp, respectively, using instrumental neutron activation analysis (INAA) demonstrated that value assignment measurement of arsenosugars was possible without arsenosugar calibration standards.

Comparative physicochemical and biological characterization of NIST Interim Reference Material PM2.5 and SRM 1648 in human A549 and mouse RAW264.7 cells.

The epidemiological association between exposure to fine particulate matter (PM2.5) and adverse health effects is well-known. Here we report the size distribution, metals content, endotoxin content, and biological activity of National Institute of Standards and Technology (NIST) Interim Reference Material (RM) PM2.5. Biological activity was measured in vitro by effects on cell viability and the release of four inflammatory immune mediators, from human A549 alveolar epithelial cells or murine RAW264.7 monocytes. A dose range covering three orders of magnitude (1-1000µg/mL) was tested, and biological activity was compared to an existing Standard Reference Material (SRM) for urban PM (NIST SRM 1648). Robust release of IL-8 and MCP-1 from A549 cells was observed in response to IRM PM2.5 exposures. Significant TNF-α, but not IL-6, secretion from RAW264.7 cells was observed in response to both IRM PM2.5 and SRM 1648 particle types. Cytokine or chemokine release at high doses often occurred in the presence of cytotoxicity, likely as a result of externalization of preformed mediator. Our results are consistent with a local cytotoxic and pro-inflammatory mechanism of response to exposure to inhaled ambient PM2.5 and reinforce the continued relevance of in vitro assays for mechanistic research in PM toxicology. Our study furthers the goal of developing reference samples of environmentally relevant particulate matter of various sizes that can be used for hypothesis testing by multiple investigators.

Certification of elements in and use of standard reference material 3280 multivitamin/multielement tablets.

Standard Reference Material 3280 Multivitamin/ Multielement Tablets was issued by the National Institute of Standards and Technology in 2009, and has certified and reference mass fraction values for 13 vitamins, 26 elements, and two carotenoids. Elements were measured using two or more analytical methods at NIST with additional data contributed by collaborating laboratories. This reference material is expected to serve a dual purpose: to provide quality assurance in support of a database of dietary supplement products and to provide a means for analysts, dietary supplement manufacturers, and researchers to assess the appropriateness and validity of their analytical methods and the accuracy of their results.

Determination of moisture content of single-wall carbon nanotubes.

Several techniques were evaluated for the establishment of reliable water/moisture content of single-wall carbon nanotubes. Karl Fischer titration (KF) provides a direct measure of the water content and was used for benchmarking against results obtained by conventional oven drying, desiccation over anhydrous magnesium perchlorate as well as by thermogravimetry and prompt gamma-ray activation analysis. Agreement amongst results was satisfactory with the exception of thermogravimetry, although care must be taken with oven drying as it is possible to register mass gain after an initial moisture loss if prolonged drying time or elevated temperatures (120 °C) are used. Thermogravimetric data were precise but a bias was evident that could be accounted for by considering the non-selective loss of mass as volatile carbonaceous components. Simple drying over anhydrous magnesium perchlorate for a minimum period of 8-10 days is recommended if KF is not available for this measurement.

Methods for the separation and quantification of arsenic species in SRM 2669: arsenic species in frozen human urine.

Two independent liquid chromatography inductively coupled plasma-mass spectrometry (LC/ICP-MS) methods for the separation of arsenic species in urine have been developed with quantification by standard additions. Seven arsenic species have been quantified in a new NIST frozen human urine Standard Reference Material (SRM) 2669 Arsenic Species in Frozen Human Urine, Levels 1 and 2. The species measured were: arsenite (As(III)), arsenate (As(V)), monomethylarsonate (MMA), dimethylarsinate (DMA), arsenobetaine (AB), arsenocholine (AC), and trimethylarsine oxide (TMAO). The purity of each arsenic standard used for quantification was measured as well as the arsenic species impurities determined in each standard. Analytical method limits of detection (L(D)) for the various species in both methods ranged from 0.2 to 0.8 microg L(-1) as arsenic. The results demonstrate that LC/ICP-MS is a sensitive, reproducible, and accurate technique for the determination of low-level arsenic species in urine. Measurements of the arsenic species 3 years after initial production of the SRM demonstrate the stability of the arsenic species in the urine reference material.

Characterization of SiGe films for use as a National Institute of Standards and Technology Microanalysis Reference Material (RM 8905).

Bulk silicon-germanium (SiGe) alloys and two SiGe thick films (4 and 5 microm) on Si wafers were tested with the electron probe microanalyzer (EPMA) using wavelength dispersive spectrometers (WDS) for heterogeneity and composition for use as reference materials needed by the microelectronics industry. One alloy with a nominal composition of Si0.86Ge0.14 and the two thick films with nominal compositions of Si0.90Ge0.10 and Si0.75Ge0.25 on Si, evaluated for micro- and macroheterogeneity, will make good microanalysis reference materials with an overall expanded heterogeneity uncertainty of 1.1% relative or less for Ge. The bulk Ge composition in the Si0.86Ge0.14 alloy was determined to be 30.228% mass fraction Ge with an expanded uncertainty of the mean of 0.195% mass fraction. The thick films were quantified with WDS-EPMA using both the Si0.86Ge0.14 alloy and element wafers as reference materials. The Ge concentration was determined to be 22.80% mass fraction with an expanded uncertainty of the mean of 0.12% mass fraction for the Si0.90Ge0.10 wafer and 43.66% mass fraction for the Si0.75Ge0.25 wafer with an expanded uncertainty of the mean of 0.25% mass fraction. The two thick SiGe films will be issued as National Institute of Standards and Technology Reference Materials (RM 8905).

Improvements in determinations using the Cu-64 annihilation gamma rays.

The method of gammagamma coincidence counting has been applied to the determination of Cu via the (64)Cu annihilation gamma rays. Preliminary experiments show that at least an order of magnitude reduction in (24)Na interference may be obtained by employing the 511-511 keV coincidence peak rather than the singles 511-keV peak. The effect of the sample matrix on the yield of (24)Na pair-production events was investigated by a combination of experimental measurements and Monte Carlo calculations.

Involvement of TLR2 and TLR4 in inflammatory immune responses induced by fine and coarse ambient air particulate matter.

Induction of proinflammatory mediators by alveolar macrophages exposed to ambient air particulate matter has been suggested to be a key factor in the pathogenesis of inflammatory and allergic diseases in the lungs. However, receptors and mechanisms underlying these responses have not been fully elucidated. In this study, we examined whether TLR2, TLR4, and the key adaptor protein, MyD88, mediate the expression of proinflammatory cytokines and chemokines by mouse peritoneal macrophages exposed to fine and coarse PM. TLR2 deficiency blunted macrophage TNF-alpha and IL-6 expression in response to fine (PM2.5), while not affecting cytokine-inducing ability of coarse NIST Standard Reference Material (SRM 1648) particles. In contrast, TLR4(-/-) macrophages showed inhibited cytokine expression upon stimulation with NIST SRM 1648 but exhibited normal responses to PM2.5. Preincubation with polymyxin B markedly suppressed the capacity of NIST SRM 1648 to elicit TNF-alpha and IL-6, indicating endotoxin as a principal inducer of cytokine responses. Overexpression of TLR2 in TLR2/4-deficient human embryonic kidney 293 cells imparted PM2.5 sensitivity, as judged by IL-8 gene expression, whereas NIST SRM 1648, but not PM2.5 elicited IL-8 expression in 293/TLR4/MD-2 transfectants. Engagement of TLR4 by NIST SRM 1648 induced MyD88-independent expression of the chemokine RANTES, while TLR2-reactive NIST IRM PM2.5 failed to up-regulate this response. Consistent with the shared use of MyD88 by TLR2 and TLR4, cytokine responses of MyD88(-/-) macrophages to both types of air PM were significantly reduced. These data indicate differential utilization of TLR2 and TLR4 but shared use of MyD88 by fine and coarse air pollution particles.

Characterization of a suite of ginkgo-containing standard reference materials.

A suite of three ginkgo-containing dietary supplement Standard Reference Materials (SRMs) has been issued by the National Institute of Standards and Technology (NIST) with certified values for flavonoid aglycones, ginkgolides, bilobalide, and selected toxic trace elements. The materials represent a range of matrices (i.e., plant, extract, and finished product) that provide different analytical challenges. The constituents have been determined by at least two independent analytical methods with measurements performed by NIST and at least one collaborating laboratory. The methods utilized different extractions, chromatographic separations, modes of detection, and approaches to quantitation. The SRMs are primarily intended for method validation and for use as control materials to support the analysis of dietary supplements and related botanical materials.

Standard reference materials (SRMs) for measurement of inorganic environmental contaminants.

NIST has developed an extensive collection of environmental SRMs, starting with fuel and biologically related materials in the late 1960s and now encompassing all sectors of environmental research. Advances in analytical methodology, including multi-element isotope-dilution mass spectrometry (IDMS) and expanded instrumental neutron-activation analysis (INAA) capabilities, enable value assignment based on fewer but better-characterized independent analytical techniques. The special advantages of IDMS for determination of S and Hg and for multi-element characterization of small-sample air particulate matter (SRM 2783) by IDMS and INAA are emphasized. Developments in materials production include the issuance of fresh-frozen biological materials and of jet-milled natural-matrix materials with improved homogeneity, including highly homogeneous air particulate matter and sediment SRMs for small-sample analytical techniques.

New NIST sediment SRM for inorganic analysis.

NIST maintains a portfolio of more than 1300 standard reference materials (SRM), more than a third of these relating to measurements in the biological and environmental fields. As part of the continuous renewal and replacement efforts, a set of new marine sediments has been recently developed covering organic and inorganic determinations. This paper describes the steps taken in sample preparation, homogeneity assay, and analytical characterization and certification with specific emphasis on SRM 2702 inorganics in marine sediment. Neutron activation analysis showed the SRM to be highly homogeneous, opening the possibility for use with solid sampling techniques. The certificate provides certified mass fraction values for 25 elements, reference values for eight elements, and information values for 11 elements, covering most of the priority pollutants with small uncertainties of only several percent relative. The values were obtained by combining results from different laboratories and techniques using a Bayesian statistical model. An intercomparison carried out in field laboratories with the material before certification illustrates a high commutability of this SRM.

The Sampling and Analysis of Human Livers.

A comprehensive approach to the analysis of human livers has been developed in a pilot program for a National Environmental Specimen Bank (NESB). Since 1980, the pilot NESB program has examined the collection, processing, storage and analysis of human livers. Sampling protocols, handling procedures and analytical methods have been developed and implemented considering the requirements for valid analytical results. Sampling and handling included the use of cleanroom technology, specific clean implements and packing materials made from titanium and Teflon and flash-freezing and preservation at liquid nitrogen temperature. Neutron activation analysis played a major role in the implemented analytical scheme. The scheme combined up to four analytical techniques to determine the distribution of 29 trace elements in 66 human livers.