Suspected microplastic counts and characteristics in fish muscle and gastrointestinal tissue

Microplastic screening of fish tissues followed methods used in Lusher et al., (2017) and Hart et al., (2022). Briefly, muscle and gastrointestinal (i.e., stomach and intestine) tissue from each fish were placed into a glass beaker, and organic (non-plastic) material in the samples was digested by adding a 10% potassium hydroxide (KOH) solution and incubated at 60°C for 24–72 hours (Karami et al., 2017). Following digestion, samples were vacuum filtered onto GF/A 1.6 µm glass fiber filters in a fume hood and left to dry in covered glass petri dishes. Particles of at least 35 µm were characterized visually using a dissection microscope (Leica EZ4, magnification 8-35x) according to physical attributes including shape (e.g., fiber, film, fragment, foam), surface texture (e.g., smooth, rough, rubber), and color (e.g., transparent, blue, black; Shim et al., 2017). Various parameters were used to identify potential plastic material. For example, suspected plastic fibers were indicated by a smooth, uniform surface with a length that exceeded the width (Lusher et al., 2020). Suspected plastic fragments were characterized by smooth or angular edges that appeared to be broken from a larger piece of debris (Lusher et al., 2020). Fragments were further identified as tire wear particles (TWP) if they were black, cylindrical, had a rubbery surface texture, and maintained their shape when manipulated with forceps (Leads and Weinstein, 2019). Suspected foam particles were characterized by a round shape and honeycomb-like porosity (Manikanda Bharath et al., 2023). Suspected fiber bundles were characterized by 20 or more fibers tangled together in a way that prevents them from being separated (Unsworth et al. 2022; Rochman et al. 2019). All particles at least 100 µm in size and with characteristics previously described were tested with a hot needle (250°C) and suspected to be of plastic origin if the needle left a mark on or melted the particle surface (De Witte et al., 2014; Devriese et al., 2015; Leads and Weinstein, 2019). Fourier Transform Infrared (FTIR) spectroscopy was available for polymer identification of particles ranging from 500 µm to 5 mm.