Plastic pollution is ubiquitous throughout the marine environment, with microplastic (i.e. <5 mm) contamination a global issue of emerging concern. The lack of universally accepted methods for quantifying microplastic contamination, including consistent application of microscopy, photography, an spectroscopy and photography, may result in unrealistic contamination estimates. Here, we present and apply an analysis workflow tailored to quantifying microplastic contamination in marine waters, incorporating stereomicroscopic visual sorting, microscopic photography and attenuated total reflectance (ATR) Fourier transform infrared (FTIR) spectroscopy. The workflow outlines step-by-step processing and associated decision making, thereby reducing bias in plastic identification and improving confidence in contamination estimates. Specific processing steps include (i) the use of a commercial algorithm-based comparison of particle spectra against an extensive commercially curated spectral library, followed by spectral interpretation to establish the chemical composition, (ii) a comparison against a customised contaminant spectral library to eliminate procedural contaminants, and (iii) final assignment of particles as either natural- or anthropogenic-derived materials, based on chemical type, a compare analysis of each particle against other particle spectra, and physical characteristics of particles. Applying this workflow to 54 tow samples collected in marine waters of North-Western Australia visually identified 248 potential anthropogenic particles. Subsequent ATR-FTIR spectroscopy, chemical assignment and visual re-inspection of photographs established 144 (58%) particles to be of anthropogenic origin. Of the original 248 particles, 97 (39%) were ultimately confirmed to be plastics, with 85 of these (34%) classified as microplastics, demonstrating that over 60% of particles may be misidentified as plastics if visual identification is not complemented by spectroscopy. Combined, this tailored analysis workflow outlines a consistent and sequential process to quantify contamination by microplastics and other anthropogenic microparticles in marine waters. Importantly, its application will contribute to more realistic estimates of microplastic contamination in marine waters, informing both ecological risk assessments and experimental concentrations in effect studies.

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