Solving a Sticky Situation: Microplastic Analysis of Lipid-Rich Tissue

Abstract

Given current concerns regarding the extent of microplastic contamination in the environment, routine monitoring for microplastics in biological tissues is becoming increasingly common place. However, complex sample matrices, such as lipid-rich tissues, require multiple pre-treatment steps which may lead to increased sample processing time and costs, and a reduction in microplastic recovery rates thereby hindering monitoring efforts. Lipid-rich (fat) tissues often pose difficulties for traditional potassium hydroxide (KOH) digestion methods due to saponification. This reaction produces a suspension of glycerol and fatty acids (soaps), which may entrap microplastics inhibiting their recovery and clog filters thus reducing the efficiency of the filtration or inhibiting it altogether. In this study, the incorporation of 100% ethanol (EtOH) to existing KOH digestion methods was found to completely redissolve the viscous saponified gel formed in these reactions, with a digestion efficiency greater than 97% for all treated lipid-rich tissue samples. Recovery of spiked polyethylene and polystyrene fragments, and rayon and polyester fibers, ranged from 93% to 100%. The addition of EtOH did not induce physical or chemical degradation on these polymers. The inclusion of an ad hoc decision-making tool within the digestion workflow reduced pre-processing time for samples and allowed for solid saponified samples to be completely redissolved. This validated workflow facilitates high through-put sampling of biota, by enabling lipidrich tissues to be filtered with a high degree of efficiency thereby successfully separating microplastics from their gelatinous matrix.

Ingestion and Depuration of Microplastics by a Planktivorous Coral Reef Fish, Pomacentrus amboinensis

“…1310-58) at 40 • C for 48 h in a ratio of 1:20 of GIT wet weight (g) to volume of KOH (mL). Digested GIT solutions were filtered through 77 and 26 µm stainless steel mesh filters (19 mm diameter) (Schlawinsky, 2020), and rinsed with 70% EtOH to remove fat vestiges (Dawson et al, 2020). Microplastics retained on mesh filters were visually identified, counted and photographed using stereomicroscopy (Leica MZ16A, Leica DFC 500, Leica Application Suite LAS 4.4.0).…”

Improved microplastic processing from complex biological samples using a customized vacuum filtration apparatus

“…Similar high recovery rates of spiked MPs have been reported from other marine biota. For example, Dawson et al (2020) achieved a 93–100% recovery rate of PE and PS fragments, and rayon and PET fibers from spiked barramundi GIT tissue after digestion with 10% KOH. In addition, a > 95% recovery rate of eight spiked polymer types (PE, polyamide, PP, PC, polyvinylchloride, acrylonitrile‐butadiene‐styrene, PS, and PET) has been reported after 17 d of 10% KOH digestion of the GIT spiral valve of Porbeagle shark ( Lamna nasus ) (Maes et al 2020).…”

“…This is likely to be reflective of the incomplete digestion of the shark tissue (Fig. S2B) and exacerbated by the discoloration of rayon rather than the effectiveness of the filtration unit and highlights the need to optimize digestion protocols to the matrix and polymer type being investigated (Dawson et al 2020; Santana et al 2022).…”

Digesting the Indigestible: Microplastic Extraction From Prawn Digestive Tracts

“…Digestive efficiency alone has recently been suggested to be an inadequate metric to evaluate complete digestion (Dawson et al, 2020). Thus in Phase 1, image analysis was applied to examine the retention of chitinous tissue after treatment as visualized by lactophenol cotton blue stain: 1) overall digestion level, 2) normalized undigested residue coverage, and 3) average blue value.…”

“…Since chitin is resistant to degradation by many chemicals including water, organic solvents, mild acidic or basic solutions (Roy et al, 2017), its presence within the digestive tract of decapods has previously been suggested to inhibit the extraction of MP from organic tissue (Cole et al, 2014). For example, potassium hydroxide (KOH) is a commonly used solution which effectively dissolves the organic tissue of most taxa for the extraction of MP (Dawson et al, 2020). However, when applied to decapod intestinal tracts, KOH has been shown to be ineffective, with intact gut linings visible amongst the filter retentate (Figure 1B) (Hara et al, 2020).…”