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.

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.

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.

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.

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.

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.

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.