Low micrometer microplastics (LMMPs), typically 1 – 10 μm, may make a significant portion of the total microplastics in aquatic systems. However, it remains elusive to quantify LMMPs reliably and cost-efficiently in complex water matrices because of the limitations of the current analytical methods. Herein, we leveraged fractionated membrane filtration to improve the reliability of Raman micro-spectroscopy (μ-Raman) for LMMP quantification. Four commercial polymericmembrane layers with pore sizes ranging from 10 to 0.4 μm were utilized in sequence to separate a mixture of polystyrene (PS) and poly(methyl methacrylate) (PMMA) LMMPs (i.e., 1.5, 3, 5, and 8 μm) from water. Over 78.5% of PS and 81.9% of PMMA were collected on membrane surfaces after sequential filtration. Sequential filtration, which separated 1.5 and 3 μm MPs from 5 and 8 μm MPs, improved the Raman image quality by reducing the particle size differences collected on each membrane layer. By direct μ-Raman imaging of mixed PS and PMMA LMMPs on the membranes, we clearly delineated the morphologies of the retained MPs and quantified the number of PS and PMMS particles with a detection limit of 2 ×10⁶ particles L^-1 (3.7 μg L^-1 for PS and 4.2 μg L^-1 PMMA LMMPs). μ-Raman coupled with the fractionated membrane filtration was also successfully applied to PS and PMMA spiked in real lake water with the same detection limit, and further improved with hyperspectral imaging by adding/multiplying Raman images. This method provides a reliable tool to separate and quantify LMMP mixtures in complex water matrices.

Weathering of a single microplastic particle can yield up to billions of nanoplastics and nanoplastic pollution is expected to be ubiquitous in the environment. Nanoplastics are potentially more hazardous than microplastics because they can cross biological membranes; yet, there is little data on the occurrence, fate and impacts of nanoplastics. A key challenge in understanding the environmental burden of nanoplastics is the detection of such small, carbon-based particles in complex natural matrices such as soils or whole organisms. Our group has been working on the development of novel plastic labeling and imaging techniques for detection of nanoplastics and microplastics in complex samples. The first approach relies on stimulated emission depletion microscopy (STED) to detect labeled nanoplastics in whole organisms or other complex samples. The second approach does not require pre-labeled plastic particles. Rather, internalized unlabeled microplastics are stained with a fluorescent dye but only after uptake into the organism. A tissue clearing technique is then used to remove tissue-bound fluorescent dye while also rendering the structurally intact organism transparent. The fluorescent dye remains bound to the internalized plastics that can now be visualized in the cleared tissue. This process yields a sample with fluorescently labeled plastic that can be rapidly imaged with light sheet microscopy. This presentation will describe the new imaging approaches and show examples of nanoplastic and microplastic detection in whole organisms. Our results show the versatility of these advanced imaging techniques for detection of the smallest plastics in complex environmental samples.

Understanding the spatial and temporal penetration patterns of organic pollutants in microplastics (μP) is important for evaluating their environmental and biological impacts, such as the “Trojan Horse” effect. However, there is a lack of an effective method to monitor the penetration processes and patterns in situ. This study aimed to develop a simple and sensitive approach for in situ imaging of organic pollutant penetration into μP. The novel method was developed using surface-enhanced Raman spectroscopy (SERS) coupled with gold nanoparticles as nanoprobes that could sensitively detect organic pollutants in low-density polyethylene (LDPE) μP spatially and temporally. The detection limit of this SERS-based method was 0.36 and 0.02 ng/mm² for ferbam (pesticide) and methylene blue (synthetic dye), respectively. The results showed that both ferbam and methylene blue could penetrate LDPE μP. The penetration depth and amount increased as the interaction time increased. Most of the absorbed organic pollutants accumulated within the top 90 μm layer of the tested μP. Compared to methylene blue, ferbam was more quickly absorbed and achieved higher accumulation in μP with a maximum of 32.57 ng/mm² after 168 h interaction. The results of this study clearly demonstrated that SERS mapping is a sensitive and in situ approach to visualize and quantify the penetration patterns of organic pollutants in μP. The innovative approach developed here can advance our understanding of μP as pollutant carriers and their influence on the environmental fate, behavior, and biological impacts of organic pollutants.

Microplastics (MPs) and nanoplastics (NPs) can serve as a vector for other harmful pollutants. However, current MP research is largely based on experiments with uniform particles. In reality, environmental MPs are more heterogenous in size and shape and are transformed upon entering the environment. There is a critical need to generate environmentally relevant MPs to better investigate MP interactions with organic compounds in laboratory experiments. Further, eco-corona formation, or interaction with Natural Organic Matter (NOM), is a surface specific interaction that can dictate particle behavior and mobility. The objective of this study is to characterize and quantify eco-corona formation and its impact on environmentally relevant MPs. Cryomilling was successfully utilized to transform polystyrene (PS) bulk material into heterogenous fragments. Additionally, an ultraviolet light chamber was employed to simulate sunlight on the fragments. Fourier transform infrared spectroscopy (FTIR) and x-ray photoelectron spectroscopy (XPS) were used to measure changes in surface chemistry due to photooxidation and cryomilling of plastics. Cryomilling didn’t alter the surface chemistry of the plastic. UV aging increased the oxygen content on the particle surfaces. Coulter counter and Nanoparticle Tracking Analysis (NTA) were used to measure the size distributions of the generated MPs. Ongoing work includes eco-corona formation experiments utilizing 3 main components of NOM (proteins, humic substances, and polysaccharides). Adsorption isotherms coupled with Quartz Crystal Microbalance with Dissipation (QCM-D) and XPS surface chemistry measurements will be utilized to better understand organic compound interaction with MP surfaces and the impacts of eco-corona formation on pollutant adsorption and particle stability.

Plastic pollution is a growing concern. To analyze plastics in environmental samples, plastics need to be isolated. We present an acidic/oxidative method optimized to preserve plastics while digesting synthetic cellulose acetate and a range of organics encountered in environmental samples. Cellulose acetate was chosen for optimization as it can be purchased as a reference material, can co-occur with plastics in environmental samples and, if it can be completely digested, is a potential filter material for the collection of nano- and micro-plastics from natural waters. Other forms of particulate organic matter (POM) were chosen to provide a range of chemistries that might alter digestion efficiency and due to the interest in the community of isolating plastics from samples where these organics occur. For instance, microalgal POM occurs in lake and ocean waters, riverine POM in rivers, and inclusion of tuna provides a test for the suitability of the method for isolating plastics from animal tissues. The method is a one-pot overnight (16-18 hours) digestion in 5 M nitric acid with 0.3 M sodium persulfate heated to 80 °C. We will present results of this method, including high retention >99% by mass of C-C bonded polymers polyethylene, polypropylene, and polystyrene, and >96% by mass of polyethylene terephthalate. Discussion will also include recommendations for the application of this method, as well as limitations and areas for future improvement.