Claudia Lorenz

Focal plane array detector-based micro-Fourier-transform infrared imaging for the analysis of microplastics in environmental samples

The pollution of the environment with microplastics (plastic pieces ,5 mm) is a problem of increasing concern. However, although this has been generally recognised by scientists and authorities, the analysis of microplastics is often done by visual inspection alone with potentially high error rates, especially for smaller particles. Methods that allow for a fast and reliable analysis of microplastics enriched on filters are lacking. Our study is the first to fill this gap by using focal plane array detector-based micro-Fourier-transform infrared imaging for analysis of microplastics from environmental samples. As a result of ouriteratively optimised analytical approach (concerningfilter material, measuring mode, measurement parameters and identification protocol), we were able to successfully measure the whole surface (>10-mm diameter) of filters with microplastics from marine plankton and sediment samples. The measurement with a highlateralresolutionallowedforthedetectionofparticlesdowntoasizeof20mminonlyafractionalpartoftimeneeded for chemical mapping. The integration of three band regions facilitated the pre-selection of potential microplastics of the ten most important polymers. Subsequent to the imaging the review of the infrared spectra of the pre-selected potential microplastics wasnecessaryforaverificationofplastic polymerorigin.Theapproachwepresenthereishighlysuitableto be implemented as a standard procedure for the analysis of small microplastics from environmental samples. However, a further automatisation with respect to measurement and subsequent particle identification would facilitate the even faster and fully automated analysis of microplastic samples. Additional keywords: microplastic analysis, microplastic detection, microplastic identification.

High Quantities of Microplastic in Arctic Deep-Sea Sediments from the HAUSGARTEN Observatory

Although mounting evidence suggests the ubiquity of microplastic in aquatic ecosystems worldwide, our knowledge of its distribution in remote environments such as Polar Regions and the deep sea is scarce. Here, we analyzed nine sediment samples taken at the HAUSGARTEN observatory in the Arctic at 2340-5570 m depth. Density separation by MicroPlastic Sediment Separator and treatment with Fentons reagent enabled analysis via Attenuated Total Reflection FTIR and μFTIR spectroscopy. Our analyses indicate the wide spread of high numbers of microplastics (42-6595 microplastics kg-1). The northernmost stations harbored the highest quantities, indicating sea ice as a possible transport vehicle. A positive correlation between microplastic abundance and chlorophyll a content suggests vertical export via incorporation in sinking (ice-) algal aggregates. Overall, 18 different polymers were detected. Chlorinated polyethylene accounted for the largest proportion (38%), followed by polyamide (22%) and polypropylene (16%). Almost 80% of the microplastics were ≤25 μm. The microplastic quantities are among the highest recorded from benthic sediments. This corroborates the deep sea as a major sink for microplastics and the presence of accumulation areas in this remote part of the world, fed by plastics transported to the North via the Thermohaline Circulation.

An automated approach for microplastics analysis using focal plane array (FPA) FTIR microscopy and image analysis

The analysis of imaging data derived from micro-Fourier transform infrared (μFTIR) microscopy is a powerful tool allowing the analysis of microplastics enriched on membrane filters. In this study we present an automated approach to reduce the time demand currently needed for data analyses. We developed a novel analysis pipeline, based on the OPUS© Software by Bruker, followed by image analysis with Python and Simple ITK image processing modules. By using this newly developed pipeline it was possible to analyse datasets from focal plane array (FPA) μFTIR mapping of samples containing up to 1.8 million single spectra. All spectra were compared against a database of different synthetic and natural polymers by various routines followed by benchmark tests with focus on accuracy and quality. The spectral correlation was optimized for high quality data generation, which allowed image analysis. Based on these results an image analysis approach was developed, providing information on particle numbers and sizes for each polymer detected. It was possible to collect all data with relative ease even for complex sample matrices. This approach significantly decreases the time demand for the interpretation of complex FTIR-imaging data and significantly increases the data quality.

Microplastics in Aquatic Systems – Monitoring Methods and Biological Consequences

Microplastic research started at the turn of the millennium and is of growing interest, as microplastics have the potential to affect a whole range of organisms, from the base of the food web to top predators, including humans. To date, most studies are initial assessments of microplastic abundances for a certain area, thereby generally distinguishing three different sampling matrices: water, sediment and biota samples. Those descriptive studies are important to get a first impression of the extent of the problem, but for a proper risk assessment of ecosystems and their inhabitants, analytical studies of microplastic fluxes, sources, sinks, and transportation pathways are of utmost importance. Moreover, to gain insight into the effects microplastics might have on biota, it is crucial to identify realistic environmental concentrations of microplastics. Thus, profound knowledge about the effects of microplastics on biota is still scarce. Effects can vary regarding habitat, functional group of the organism, and polymer type for example, making it difficult to find quick answers to the many open questions. In addition, microplastic research is accompanied by many methodological challenges that need to be overcome first to assess the impact of microplastics on aquatic systems. Thereby, a development of standardized operational protocols (SOPs) is a pre-requisite for comparability among studies. Since SOPs are still lacking and new methods are developed or optimized very frequently, the aim of this chapter is to point out the most crucial challenges in microplastic research and to gather the most recent promising methods used to quantify environmental concentrations of microplastics and effect studies.

Comparison of Raman and Fourier Transform Infrared Spectroscopy for the Quantification of Microplastics in the Aquatic Environment

Microplastics (MPs, <5 mm) have been reported as emerging environmental contaminants, but reliable data are still lacking. We compared the two most promising techniques for MP analysis, namely, Raman and Fourier transform infrared (FTIR) spectroscopy, by analyzing MPs extracted from North Sea surface waters. Microplastics >500 μm were visually sorted and manually analyzed by μ-Raman and attenuated total reflection (ATR)-FTIR spectroscopy. Microplastics ≤500 μm were concentrated on gold-coated filters and analyzed by automated single-particle exploration coupled to μ-Raman (ASPEx-μ-Raman) and FTIR imaging (reflection mode). The number of identified MPs >500 μm was slightly higher for μ-Raman (+23%) than ATR-FTIR analysis. Concerning MPs ≤500 μm, ASPEx-μ-Raman quantified two-times higher MP numbers but required a four-times higher analysis time compared to FTIR imaging. Because ASPEx-μ-Raman revealed far higher MP concentrations (38-2621 particles m-3) compared to the results of previous water studies (0-559 particles m-3), the environmental concentration of MPs ≤500 μm may have been underestimated until now. This may be attributed to the exceptional increase in concentration with decreasing MP size found in this work. Our results demonstrate the need for further research to enable time-efficient routine application of ASPEx-μ-Raman for reliable MP counting down to 1 μm.

Using the FlowCam to Validate an Enzymatic Digestion Protocol Applied to Assess the Occurrence of Microplastics in the Southern North Sea

As the plastic production has been rising since the last five decades, so does the concern for the occurrence of microplastic particles (< 5 mm) in the marine environment during the past years. But still by now the extent of this microplastic pollution of coastal waters and the open ocean remains unclear. Since monitoring the abundance of microplastics in the marine environment is requested by the Marine Strategy Framework Directive (MSFD) standardized and reliable methods for the detection of microplastics are urgently needed. Studies differ mainly in their purification methods, aiming to reduce biogenic material in environmental samples without altering the plastic polymers to facilitate a clear assignment of the microplastics. In the present and ongoing study the purification method consists of a treatment with technical enzymes and detergents to reduce the use of oxidants and avoid the use of strong acids as well as the subsequent identification and quantification of microplastics applying Focal Plane Array (FPA)-based µ-Fourier-Transform Infrared (µFT-IR) spectroscopy. A new approach involving the FlowCam (Fluid Imaging Technologies) was used to validate the efficacy of the applied digestion protocol. Hereby untreated plankton samples were run through the FlowCam and compared to the samples after treatment with the various technical enzymes (protease, cellulase and chitinase), sodium dodecyl sulfate and hydrogen peroxide according to protocol regarding changes to particle count, particle sizes and appearance. Furthermore a metabolic fingerprint of every digestion step was drawn up applying the same µFT-IR spectroscopic analysis used for the identification of microplastics. Exemplary results derived by applying the validated protocol show the occurrence of microplastics in sediments of the German coastal waters and highlight the need for a further assessment of the microplastic pollution in various compartments i.e. sediments and surface waters and investigating the link between these two.

Automated Analysis of µFTIR Imaging Data for Microplastic Samples

The pollution of the oceans with plastic particles smaller than 5 mm, called microplastics is moving into the focus of science and governments. To determine the amount of microplastics several steps are necessary, starting with the sampling, work up and finally analysis. Each step has its own challenges due to small size of the particles.[1] For analysis the imaging with µFTIR microscopy is a powerful tool allowing the analysis of complete filters. Systematic screening for optimal conditions and filter materials have already been performed.[2] While the measurement is performed mostly by the spectrometer, the data interpretation process is highly time consuming as it has to be made by hand on the basis of false color images. To overcome the manual part we developed a novel approach based on the Bruker OPUS© Software to decrease the high time demand for the analysis of microplastics. With this approach it was possible to analyze measurement files from focal plane array (FPA) FTIR mapping containing up to 1.8 million single spectra. These spectra were compared with a database of different synthetic and natural polymers by various methods. By benchmark tests their performance was monitored with the focus on accuracy and data quality. After optimization high quality data was generated, which allowed image analysis. Based on these results an approach for image analysis was developed, giving information for the particle size distribution for each polymer type as well as their distribution on the filter. It was possible to collect all data with relative ease even for complex sample matrices. This approach has significantly decreased the time demand for the interpretation of FTIR-imaging data and increased the generated data quality. [1] M. Bergmann, L. Gutow, M. Klages, Marine Anthropogenic Litter, Springer, Cham Heidelberg New York Dordrecht London, 2015. [2] M. G. J. Loder, M. Kuczera, S. Mintenig, C. Lorenz, G. Gerdts, Environ. Chem. 2015, 12, 563-581.