The influence of complex matrices on method performance in extracting and monitoring for microplastics.

Previous studies have evaluated method performance for quantifying and characterizing microplastics in clean water, but little is known about the efficacy of procedures used to extract microplastics from complex matrices. Here we provided 15 laboratories with samples representing four matrices (i.e., drinking water, fish tissue, sediment, and surface water) each spiked with a known number of microplastic particles spanning a variety of polymers, morphologies, colors, and sizes. Percent recovery (i.e., accuracy) in complex matrices was particle size dependent, with ∼60-70% recovery for particles >212 µm, but as little as 2% recovery for particles <20 µm. Extraction from sediment was most problematic, with recoveries reduced by at least one-third relative to drinking water. Though accuracy was low, the extraction procedures had no observed effect on precision or chemical identification using spectroscopy. Extraction procedures greatly increased sample processing times for all matrices with the extraction of sediment, tissue, and surface water taking approximately 16, 9, and 4 times longer than drinking water, respectively. Overall, our findings indicate that increasing accuracy and reducing sample processing times present the greatest opportunities for method improvement rather than particle identification and characterization.

A new method for microplastic extraction from fish guts assisted by chemical dissolution.

A new protocol for the extraction of microplastic is proposed and demonstrated which combines dissection, ultrasonication, and filtration with chemical dissolution in order to estimate microplastic contamination in fish or other samples with significant biomass. This protocol enables initial characterization of the sample through dissection followed by chemical dissolution to isolate polymer debris while minimizing analytical uncertainties and maintaining microplastic particle integrity. The extraction method begins with dissection and inspection of the stomach contents, followed by pulsed ultrasonic extraction to remove the majority of biomass and surface contaminants. Subsequent chemical dissolution of the extracted contents using KOH and HCl removes any remaining biomass and inorganic interferences. Incorporating chemical dissolution post-extraction minimizes the overall biomass subjected to dissolution, thereby enabling faster processing and subsequently a cleaner sample compared to methods involving digestion of the entire organism. Furthermore, the chemical dissolution step enables direct filter analysis for microplastics, thereby minimizing the potential loss of microplastic particles associated with manual particle transfer. Hence, the microplastic extraction method presented here is suitable for the extraction and identification of small (>20 µm) and potentially brittle microplastic.

Microplastic fragment and fiber contamination of beach sediments from selected sites in Virginia and North Carolina, USA.

Microplastic particles (<5 mm) constitute a growing pollution problem within coastal environments. This study investigated the microplastic presence of estuarine and barrier island beaches in the states of Virginia and North Carolina, USA. Seventeen sediment cores were collected at four study sites and initially tested for microplastic presence by pyrolysis-gas chromatography-mass spectrometry. For the extraction, microplastic particles were first separated from the sediment using a high-density cesium chloride solution (1.88 g/mL). In a second step, an oil extraction collected the remaining microplastic particles of higher densities. Under the light microscope, the extracted microplastic particles were classified based on their morphologies into fragments and fibers. Raman microspectroscopy chemically identified a subset of microplastic particles as polypropylene, polyethylene terephthalate, poly(4-vinylbiphenyl), polystyrene, polyethylene, and nylon. The results show a concentration of microplastic particles (1410 ± 810 per kg of dry sediment) even in protected and ostensibly unpolluted estuarine and beach sediments of Virginia and North Carolina.

Nondestructive Extraction and Identification of Microplastics from Freshwater Sport Fish Stomachs.

Microplastics were extracted from freshwater sport fish stomachs containing substantial biomass and identified using optical microscopy, scanning electron microscopy plus energy-dispersive X-ray spectroscopy (SEM/EDS), and Fourier transform infrared (FTIR) micro-spectroscopy with automated spectral mapping. An extraction method is presented that uses a negatively pressurized sieve stack and purified water to preserve plastic surface characteristics and any adsorbed persistent organic pollutants (POPs). This nondestructive extraction method for large predators' stomachs enables multiple trophic-level studies from one fish sampling event and provides other dietary and behavioral data. FTIR-identified microplastics 50-1500 µm, including polyethylene (two with plastic additive POPs), styrene acrylonitrile, polystyrene, and nylon and polyethylene terephthalate fibers 10-50 µm wide. SEM/EDS revealed characteristic surface weathering on the plastic surfaces. The nylon fibers appear to be from human fishing activities, suggesting options for management. Some particles visually identified as potential plastics were revealed by micro-spectroscopy to be mineralized, natural polyamide proteins, or nonplastic shell pieces. A low-cost, reflective sample preparation method with stable particle mounting was developed to enable automated mapping, improved FTIR throughput, and lower detection size limit. This study yielded 37 intact prey items set aside for future analyses.

Identifying regional soil as the potential source of PM2.5 particulate matter on air filters collected in Imperial Valley, California - A Raman micro-spectroscopy study.

This work explores the use of Raman micro-spectroscopy to determine sources of airborne particulate matter collected on PM2.5 air filters in Imperial Valley, California. The goal is to examine if nearby soil is a potential source of particles sampled on air filters deployed in an urbanized desert area during events of unusually high PM2.5 excursions. Particle specific composition information can be an indicator of potential origin. This can provide insights into the source of unexpectedly high proportion of large particles sampled on PM2.5 filters in the vicinity of Imperial Valley. The measured spectral correspondence between the filter and soil particles, in the size range of 2.5-10 µm, is consistent with windblown dust being a likely source of the larger (>2.5 µm) particles collected on the PM2.5 filters. Additionally, these particles were identified as components of commonly occurring crustal minerals in the vicinity of the sampling site, such as iron oxides, hydroxides, sulfides, titanium dioxides and aluminosilicates. A substantial portion of the analyzed filter particles displayed a strong broadband fluorescence signal, which is consistent with the presence of organic matter and has been recognized as a marker for soil related origin of the filter particles. Elemental carbon (soot) was found to be prevalent among the particles as well, suggesting the existence of combustion related sources. Comparison between a heavily loaded filter sample and a filter with a more typical, lower loading did not show any obvious difference in chemical compositions. In both cases the particles appeared to be of crustal origin with the prevalence of elemental carbon. The primary difference between these two filter samples appear to be their particle size distribution - the heavily loaded filter sample contained greater proportion of large particles (>2.5 µm), and was more consistent with spectral signature of soils analyzed from the region.

Molecular identification of polymers and anthropogenic particles extracted from oceanic water and fish stomach - A Raman micro-spectroscopy study.

Pacific Ocean trawl samples, stomach contents of laboratory-raised fish as well as fish from the subtropical gyres were analyzed by Raman micro-spectroscopy (RMS) to identify polymer residues and any detectable persistent organic pollutants (POP). The goal was to access specific molecular information at the individual particle level in order to identify polymer debris in the natural environment. The identification process was aided by a laboratory generated automated fluorescence removal algorithm. Pacific Ocean trawl samples of plastic debris associated with fish collection sites were analyzed to determine the types of polymers commonly present. Subsequently, stomach contents of fish from these locations were analyzed for ingested polymer debris. Extraction of polymer debris from fish stomach using KOH versus ultrapure water were evaluated to determine the optimal method of extraction. Pulsed ultrasonic extraction in ultrapure water was determined to be the method of choice for extraction with minimal chemical intrusion. The Pacific Ocean trawl samples yielded primarily polyethylene (PE) and polypropylene (PP) particles >1 mm, PE being the most prevalent type. Additional microplastic residues (1 mm - 10 µm) extracted by filtration, included a polystyrene (PS) particle in addition to PE and PP. Flame retardant, deca-BDE was tentatively identified on some of the PP trawl particles. Polymer residues were also extracted from the stomachs of Atlantic and Pacific Ocean fish. Two types of polymer related debris were identified in the Atlantic Ocean fish: (1) polymer fragments and (2) fragments with combined polymer and fatty acid signatures. In terms of polymer fragments, only PE and PP were detected in the fish stomachs from both locations. A variety of particles were extracted from oceanic fish as potential plastic pieces based on optical examination. However, subsequent RMS examination identified them as various non-plastic fragments, highlighting the importance of chemical analysis in distinguishing between polymer and non-polymer residues.

SEM/EDS and optical microscopy analyses of microplastics in ocean trawl and fish guts.

Microplastic particles from Atlantic and Pacific Ocean trawls, lab-fed fish guts and ocean fish guts have been characterized using optical microscopy and SEM/EDS in terms of size, morphology, and chemistry. We assessed whether these measurements could serve as a rapid screening process for subsequent identification of the likely microplastic candidates by micro-spectroscopy. Optical microscopy enabled morphological classification of the types of particles or fibers present in the sample, as well as the quantification of particle size ranges and fiber lengths. SEM/EDS analysis was used to rule out non-plastic particles and screen the prepared samples for potential microplastic, based on their element signatures and surface characteristics. Chlorinated plastics such as polyvinyl chloride (PVC) could be easily identified with SEM/EDS due to their unique elemental signatures including chlorine, as could mineral species that are falsely identified as plastics by optical microscopy. Particle morphology determined by optical microscopy and SEM suggests the fish ingested particles contained both degradation fragments from larger plastic pieces and also manufactured microplastics. SEM images of microplastic particle surfaces revealed characteristic cracks consistent with environmental exposure, as well as pigment particles consistent with manufactured materials. Most of the microplastic surfaces in the fish guts and ocean trawls were covered with biofilms, radiolarians, and crustaceans. Many of the fish stomachs contained micro-shell pieces which visually resembled microplastics.

Raman spectroscopy based identification of flame retardants in consumer products using an acquired reference spectral library.

Flame retardants (FRs), a class of commonly used chemical additives in consumer products such as polyurethane foams, are well known for their persistence in the environment, bioaccumulation and potential toxicity [1]. In order to address the potential health concerns and environmental impacts associated with the wide-spread use these chemicals, it is essential to identify them efficiently in the environment and consumer products. Raman spectroscopy (RS) offers an attractive option for the non-invasive, in-situ identification of flame retardants in a variety of sample formats [2-4]. RS based chemical identification relies on the availability of spectral libraries for identification through spectral matching with reference chemicals. Here we present the application of Raman spectroscopy for identifying FR additives in select consumer products using an acquired spectral library of commonly used FRs. The RS based method described here enables simultaneous identification of multiple components within a sample, which can offer important insights into the sources of FR contamination, in addition to identification of the FR component itself. The availability of Raman spectral library of commercially used FRs, such as the one presented here, will facilitate the identification of these chemicals in consumer products.

Microplastics in four estuarine rivers in the Chesapeake Bay, U.S.A.

Once believed to degrade into simple compounds, increasing evidence suggests plastics entering the environment are mechanically, photochemically, and/or biologically degraded to the extent that they become imperceptible to the naked eye yet are not significantly reduced in total mass. Thus, more and smaller plastics particles, termed microplastics, reside in the environment and are now a contaminant category of concern. The current study tested the hypotheses that microplastics concentration would be higher in proximity to urban sources, and vary temporally in response to weather phenomena such as storm events. Triplicate surface water samples were collected approximately monthly between July and December 2011 from four estuarine tributaries within the Chesapeake Bay, U.S.A. using a manta net to capture appropriately sized microplastics (operationally defined as 0.3-5.0 mm). Selected sites have watersheds with broadly divergent land use characteristics (e.g., proportion urban/suburban, agricultural and/or forested) and wide ranging population densities. Microplastics were found in all but one of 60 samples, with concentrations ranging over 3 orders of magnitude (<1.0 to >560 g/km(2)). Concentrations demonstrated statistically significant positive correlations with population density and proportion of urban/suburban development within watersheds. The greatest microplastics concentrations also occurred at three of four sites shortly after major rain events.

Morphology, spatial distribution, and concentration of flame retardants in consumer products and environmental dusts using scanning electron microscopy and Raman micro-spectroscopy.

We characterized flame retardant (FR) morphologies and spatial distributions in 7 consumer products and 7 environmental dusts to determine their implications for transfer mechanisms, human exposure, and the reproducibility of gas chromatography-mass spectrometry (GC-MS) dust measurements. We characterized individual particles using scanning electron microscopy/energy dispersive x-ray spectroscopy (SEM/EDS) and Raman micro-spectroscopy (RMS). Samples were screened for the presence of 3 FR constituents (bromine, phosphorous, non-salt chlorine) and 2 metal synergists (antimony and bismuth). Subsequent analyses of select samples by RMS enabled molecular identification of the FR compounds and matrix materials. The consumer products and dust samples possessed FR elemental weight percents of up to 36% and 31%, respectively. We identified 24 FR-containing particles in the dust samples and classified them into 9 types based on morphology and composition. We observed a broad range of morphologies for these FR-containing particles, suggesting FR transfer to dust via multiple mechanisms. We developed an equation to describe the heterogeneity of FR-containing particles in environmental dust samples. The number of individual FR-containing particles expected in a 1-mg dust sample with a FR concentration of 100ppm ranged from <1 to >1000 particles. The presence of rare, high-concentration bromine particles was correlated with decabromodiphenyl ether concentrations obtained via GC-MS. When FRs are distributed heterogeneously in highly concentrated dust particles, human exposure to FRs may be characterized by high transient exposures interspersed by periods of low exposure, and GC-MS FR concentrations may exhibit large variability in replicate subsamples. Current limitations of this SEM/EDS technique include potential false negatives for volatile and chlorinated FRs and greater quantitation uncertainty for brominated FR in aluminum-rich matrices.

Correlated Raman micro-spectroscopy and scanning electron microscopy analyses of flame retardants in environmental samples: a micro-analytical tool for probing chemical composition, origin and spatial distribution.

We present correlated application of two micro-analytical techniques: scanning electron microscopy/energy dispersive X-ray spectroscopy (SEM/EDS) and Raman micro-spectroscopy (RMS) for the non-invasive characterization and molecular identification of flame retardants (FRs) in environmental dusts and consumer products. The SEM/EDS-RMS technique offers correlated, morphological, molecular, spatial distribution and semi-quantitative elemental concentration information at the individual particle level with micrometer spatial resolution and minimal sample preparation. The presented methodology uses SEM/EDS analyses for rapid detection of particles containing FR specific elements as potential indicators of FR presence in a sample followed by correlated RMS analyses of the same particles for characterization of the FR sub-regions and surrounding matrices. The spatially resolved characterization enabled by this approach provides insights into the distributional heterogeneity as well as potential transfer and exposure mechanisms for FRs in the environment that is typically not available through traditional FR analysis. We have used this methodology to reveal a heterogeneous distribution of highly concentrated deca-BDE particles in environmental dust, sometimes in association with identifiable consumer materials. The observed coexistence of deca-BDE with consumer material in dust is strongly indicative of its release into the environment via weathering/abrasion of consumer products. Ingestion of such enriched FR particles in dust represents a potential for instantaneous exposure to high FR concentrations. Therefore, correlated SEM/RMS analysis offers a novel investigative tool for addressing an area of important environmental concern.

Raman microspectroscopy-based identification of individual fungal spores as potential indicators of indoor contamination and moisture-related building damage.

We present an application of Raman microspectroscopy (RMS) for the rapid characterization and identification of individual spores from several species of microfungi. The RMS-based methodology requires minimal sample preparation and small sample volumes for analyses. Hence, it is suitable for preserving sample integrity while providing micrometer-scale spatial resolution required for the characterization of individual cells. We present the acquisition of unique Raman spectral signatures from intact fungal spores dispersed on commercially available aluminum foil substrate. The RMS-based method has been used to compile a reference library of Raman spectra from several species of microfungi typically associated with damp indoor environments. The acquired reference spectral library has subsequently been used to identify individual microfungal spores through direct comparison of the spore Raman spectra with the reference spectral signatures in the library. Moreover, the distinct peak structures of Raman spectra provide detailed insight into the overall chemical composition of spores. We anticipate potential application of this methodology in the fields of public health, forensic sciences, and environmental microbiology.

Spatially resolved characterization of water and ion incorporation in Bacillus spores.

We present the first direct visualization and quantification of water and ion uptake into the core of individual dormant Bacillus thuringiensis subsp. israelensis (B. thuringiensis subsp. israelensis) endospores. Isotopic and elemental gradients in the B. thuringiensis subsp. israelensis spores show the permeation and incorporation of deuterium in deuterated water (D(2)O) and solvated ions throughout individual spores, including the spore core. Under hydrated conditions, incorporation into a spore occurs on a time scale of minutes, with subsequent uptake of the permeating species continuing over a period of days. The distribution of available adsorption sites is shown to vary with the permeating species. Adsorption sites for Li(+), Cs(+), and Cl(-) are more abundant within the spore outer structures (exosporium, coat, and cortex) relative to the core, while F(-) adsorption sites are more abundant in the core. The results presented here demonstrate that elemental abundance and distribution in dormant spores are influenced by the ambient environment. As such, this study highlights the importance of understanding how microbial elemental and isotopic signatures can be altered postproduction, including during sample preparation for analysis, and therefore, this study is immediately relevant to the use of elemental and isotopic markers in environmental microbiology and microbial forensics.

Interfacial behavior of perchlorate versus chloride ions in aqueous solutions.

In recent years, theoretical as well as experimental studies have presented a novel view of the aqueous interface, wherein hard and/or multiply charged ions are excluded from the interface but large polarizable anions show interfacial enhancement relative to the bulk. The observed trend in the propensity of anions to adsorb at the air/water interface appears to follow an inverse order of the Hofmeister series for anions. This study focuses on experimental and theoretical examination of the partitioning behavior of perchlorate (ClO(4)(-)) and chloride (Cl(-)) ions at the air/water interface. We have used ambient pressure X-ray photoelectron spectroscopy to directly probe the interfacial concentrations of ClO(4)(-) and Cl(-) ions in sodium perchlorate and sodium chloride solutions, respectively. In the case of ClO(4)(-) ion, experimental observations are compared with molecular dynamics simulations utilizing both first principles based interaction potentials as well as polarizable classical force fields. Both the experimental and the theoretical results show enhancement of ClO(4)(-) ion at the interface, compared with the absence of such enhancement in the case of the Cl(-) ion. Our observations are in agreement with the expected trend in the interfacial propensity of anions based on the Hofmeister series.

Ion partitioning at the liquid/vapor interface of a multicomponent alkali halide solution: a model for aqueous sea salt aerosols.

The chemistry of Br species associated with sea salt ice and aerosols has been implicated in the episodes of ozone depletion reported at Arctic sunrise. However, Br(-) is only a minor component in sea salt, which has a Br(-)/Cl(-) molar ratio of approximately 0.0015. Sea salt is a complex mixture of many different species, with NaCl as the primary component. In recent years experimental and theoretical studies have reported enhancement of the large, more polarizable halide ion at the liquid/vapor interface of corresponding aqueous alkali halide solutions. The proposed enhancement is likely to influence the availability of sea salt Br(-) for heterogeneous reactions such as those involved in the ozone depletion episodes. We report here ambient pressure X-ray photoelectron spectroscopy studies and molecular dynamics simulations showing direct evidence of Br(-) enhancement at the interface of an aqueous NaCl solution doped with bromide. The experiments were carried out on samples with Br(-)/Cl(-) ratios in the range 0.1% to 10%, the latter being also the ratio for which simulations were carried out. This is the first direct measurement of interfacial enhancement of Br(-) in a multicomponent solution with particular relevance to sea salt chemistry.

Imaging and 3D elemental characterization of intact bacterial spores by high-resolution secondary ion mass spectrometry.

We present a quantitative, imaging technique based on nanometer-scale secondary ion mass spectrometry for mapping the 3D elemental distribution present in an individual micrometer-sized Bacillus spore. We use depth profile analysis to access the 3D compositional information of an intact spore without the additional sample preparation steps (fixation, embedding, and sectioning) typically used to access substructural information in biological samples. The method is designed to ensure sample integrity for forensic characterization of Bacillus spores. The minimal sample preparation/alteration required in this methodology helps to preserve sample integrity. Furthermore, the technique affords elemental distribution information at the individual spore level with nanometer-scale spatial resolution and high (microg/g) analytical sensitivity. We use the technique to map the 3D elemental distribution present within Bacillus thuringiensis israelensis spores.

Electron spectroscopy of aqueous solution interfaces reveals surface enhancement of halides.

It has been suggested that enhanced anion concentrations at the liquid/vapor interface of airborne saline droplets are important to aerosol reactions in the atmosphere. We report ionic concentrations in the surface of such solutions. Using x-ray photoelectron spectroscopy operating at near ambient pressure, we have measured the composition of the liquid/vapor interface for deliquesced samples of potassium bromide and potassium iodide. In both cases, the surface composition of the saturated solution is enhanced in the halide anion compared with the bulk of the solution. The enhancement of anion concentration is more dramatic for the larger, more polarizable iodide anion. By varying photoelectron kinetic energies, we have obtained depth profiles of the liquid/vapor interface. Our results are in good qualitative agreement with classical molecular dynamics simulations. Quantitative comparison between the experiments and the simulations indicates that the experimental results exhibit more interface enhancement than predicted theoretically.

In situ study of water-induced segregation of bromide in bromide-doped sodium chloride by scanning polarization force microscopy.

The adsorption of water on Br-doped NaCl crystals has been studied in situ using scanning polarization force microscopy, a noncontact electrostatic atomic force microscopy operation mode. Both topography and contact potential images were acquired as a function of relative humidity at room temperature, from 0% to more than 55%. It was found that the surface of the freshly cleaved crystal has an inhomogeneous electrical surface potential distribution with the steps more negative than the terraces below 40% relative humidity. This difference disappears when the humidity reaches 40% and higher. Below 40% the step morphology experiences only small changes due to water adsorption; however, above 40% major changes take place due to solvation, segregation, and redistribution of lattice ions. Bromide-rich islands and crystallites segregate to the surface above 40% relative humidity followed by drying. These islands and crystallites have a negative surface potential relative to the rest of the surface. These effects are attributed to the preferential solvation and segregation of Br- ions.

Rigid Molecular Rods with Cumulene-Bridged Polyphosphines: Synthesis, Electronic Communication, Molecular Photophysics, Mixed-Valence State, and X-ray Photoelectron Spectroscopic Study.

The synthesis, molecular photophysics, redox characteristics, and electronic interactions, as well as an X-ray photoelectron spectroscopic (XPS) study of a series of Ru(II) and Os(II) complexes with a polyphosphine/cumulene spacer, namely, 1,1',4,4'-tetrakis(diphenylphosphino)cumulene (C(4)P(4)), are studied and compared with the corresponding systems containing spacers with shorter sp carbon chain (C(n)()) lengths. Characterizations of all mono-, homo-, and heterobimetallic complexes with PF(6)(-) counteranions are accomplished using (1)H, (13)C, and (31)P{(1)H} NMR, fast atom bombardment (FAB/MS), and matrix-assisted laser desorption ionization time-of-flight (MALDI-TOF/MS) mass spectroscopy and elemental analysis. From the electrochemical study it is observed that the length of the C(n)() bridges has a profound influence on redox potentials and the electronic interaction between the two metal-based termini. XPS studies reveal that a simple change in carbon chain length affects the electron donation of the phosphine spacer to the metal-based termini. As a result, the redox potential of the Ru(II) or Os(II) center is shifted significantly. The comproportionation constant, K(c), is calculated as 1.3 x 10(7) (M = Ru(II)) or 4.5 x 10(10) (M = Os(II)) for homobimetallic [(bpy)(2)M(C(4)P(4))M(bpy)(2)](4+), suggesting a strong electronic communication across the C(4)P(4) spacer. However, the K(c) value is estimated to be ca. 4 for the corresponding complexes [(bpy)(2)M(C(3)P(4))M(bpy)(2)](4+) (M = Ru, Os; C(3)P(4) = 1,1',3,3'-tetrakis(diphenylphosphino)allene), indicative of a system with electronic isolation between the two termini. In heterobimetallic [(bpy)(2)Ru(C(n)()P(4))Os(bpy)(2)](4+) (n = 3, 4), the energy transfer from Ru(II) to Os(II) is found to be very efficient, with rate constants k(en) of ca. 3 x 10(9) s(-)(1) (n = 3) and 1 x 10(11) s(-)(1) (n = 4). The increased value of k(en) upon the change from C(3) to C(4) can be explained by the increase in the electronic communication across spacers. Detailed studies and calculations have revealed a Dexter-type of mechanism for the triplet energy transfer in the system.