Chitinase digestion for the analysis of microplastics in chitinaceous organisms using the terrestrial isopod Oniscus asellus L. as a model organism

Emilie M.F Kallenbach,Rachel R Hurley,Amy Lusher,Nikolai Friberg

doi:10.1016/j.scitotenv.2021.147455

Emilie M.F Kallenbach, Rachel R Hurley + Show 2 more

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https://doi.org/10.1016/j.scitotenv.2021.147455

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Abstract

Chitinaceous organisms have been found to ingest microplastic; however, a standardised, validated, and time- and cost-efficient method for dissolving these organisms without affecting microplastic particles is still required. This study tested four protocols for dissolving organisms with a chitin exoskeleton: 1) potassium hydroxide (KOH) + chitinase, 2) Creon® + chitinase, 3) hydrogen peroxide (H2O2) + chitinase, and, 4) Nitric Acid (HNO3) + hydrogen peroxide (H2O2). The effects on microplastics composed of eight different polymers were also tested. The use of H2O2 followed by chitinase was found to be a highly efficient method. The three other protocols either did not digest the chitin sufficiently or negatively affected the tested polymers. A recovery test using microplastic fibres, beads and tyre particles revealed high recovery rates of 0.85, 0.89 and 1 respectively. This further supported the applicability of the H2O2 and chitinase (protocol 3) for dissolving chitinaceous organisms. Thus, we recommend that future investigations of microplastic (0.05 μm–5000 μm) in chitinaceous organisms (0.3 cm–5 cm) utilise the here presented methodology. This represents an important component of the ongoing validation and harmonization of methodological approaches that are urgently needed for the advancement of microplastic assessments globally.

Highlights

Plastic contamination is firmly established on the global environmental agenda (e.g. Rochman and Hoellein, 2020)
Methods testing B: effect on plastic polymers To test if the selected protocols caused any impacts of plastic particles, eight different plastic polymers: polyethylene terephthalate (PET), polymethyl methacrylate (PMMA), polypropylene (PP), polystyrene (PS), high-density polyethylene (HDPE), low-density polyethylene (LDPE), polycarbonate (PC), and polyamide-6,6 (PA-6,6) were selected and exposed to the four chemical treatments
The woodlice exoskeletons were bleached but remained intact (Fig. 2a). If this protocol were to be applied to environmental samples, microplastic ingested by the organisms may be trapped inside which will hinder subsequent analysis

Summary

Introduction

Plastic contamination is firmly established on the global environmental agenda (e.g. Rochman and Hoellein, 2020). One of the key focus areas of the microplastic research field is the potential ecological impact of microplastics in natural ecosystems This goes beyond the aesthetical issues associated with visible plastic litter accumulations (Li et al, 2018; Wright et al, 2013). We don't have a complete understanding of the risk and effects that microplastic poses to the different environmental matrices This has led to significant research effort aiming at elucidating the potential sources, occurrence, and effects of microplastics in biota (Beer et al, 2018; Bergami et al, 2020; Bour et al, 2018; Bråte et al, 2018; Catarino et al, 2018; De Witte et al, 2014; Digka et al, 2018; Foekema et al, 2013; Hurley and Nizzetto, 2018; Lusher et al, 2013; Lwanga et al, 2017)

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