A standardized protocol for microplastic and non-synthetic microfiber extraction and µFTIR identification in...

Elasmobranch gastrointestinal tissues are a complex matrix for microplastic work: high in lipids, dense in connective tissue, and prone to losing small particles during handling. In this work, we developed a protocol that targets these problems. Tissue is digested in 20% (w/v) KOH at 40°C until the soft fraction dissolves (24-72 h); when organic residues persist, 30% (v/v) H₂O₂ is added until the solution clears to amber. The digest is then density-separated in 5 M NaCl, filtered through 1.2 µm glass fiber, sorted under a stereomicroscope, and identified by µFTIR-ATR. Validation on eight samples spiked with 100 polyethylene reference particles (50-500 µm) gave recoveries of 90-96% (mean 93.4 ± 2.06%); the procedural blank returned 97%. From the real samples, 172 particles were recovered, mostly fibers, with cellulose as the dominant material, alongside PET, acrylic, nylon, and polypropylene. Detection was reliable down to ∼150 µm by stereomicroscopy and ∼60-70 µm by µFTIR. The method is not designed for particles below 50 µm or polymers denser than the saline brine (∼1.2 g cm⁻³); because both require adapting the density step. Key features of the method:•Sequential KOH-H₂O₂ digestion handles the fat- and protein-rich matrix of shark and ray digestive tissue without altering common polymers.•Mean recovery of 93.4% on polyethylene-spiked samples (90-96%); 97% in procedural blanks.•µFTIR-ATR identification down to ∼60 µm, with a reliable visual sorting threshold of ∼150 µm.