Additives, plasticizers, small microplastics (<100 μm), and other microlitter components in the gastrointestinal tract of commercial teleost fish: Method of extraction, purification, quantification, and characterization using Micro-FTIR

https://doi.org/10.1016/j.marpolbul.2022.113477Get rights and content

Highlights

  • A pretreatment method for both SMP and microlitter in fish was developed.

  • Concurrent quantification and identification of SMPs and microlitter via MicroFTIR

  • Biopolymers were identified and quantified.

  • Tire particles and vulcanizing agents were identified and quantified.

  • Additive and plasticizers can be proxies of microplastics' occurrence.

Abstract

One of the aims of this study is the development of a pretreatment method for additives, plasticizers and other components of micro-litter (APFs), and small microplastics (SMPs <100 μm) in the gastrointestinal tract (GIT) of five of the most widely distributed and consumed commercial fish species, Engraulis encrasiculos, Sardina pilchardus, Mullus surmuletus, Solea solea, and Sparus aurata. The second aim was to develop a simultaneous quantification and identification method via Micro-FTIR of APFs and SMPs ingested by these commercial fish species. The distribution of SMPs and APFs is characteristically different for each species investigated. E. encrasiculos and S. pilchardus had a higher weight of SMPs than the other species investigated. Regarding APFs, the highest abundance was observed in E. encrasiculos. This study highlights the importance of studying additives and plasticizers that can be used as efficient proxies of microplastics, as shown by the presence of vulcanizing agents such as Vanax®.

Introduction

Plastic pollution has become a relevant and critical issue of concern for aquatic and terrestrial environments; this concern is even worrisome when considering plastics below 100 μm in size (small microplastics, SMPs; Corami et al., 2020b; Iannilli et al., 2019, Iannilli et al., 2020). Microplastics (MPs) contain additives and plasticizers, as the definition given by ECHA (2019), and are part of micro-litter, together with natural and non-plastic synthetic fibers (i.e., rayon). MPs have been reported to be ingested by a variety of organisms: invertebrates, fish, birds, and mammals as well (Corami et al., 2020b; Iannilli et al., 2019; Lopes et al., 2020; Lusher et al., 2013; Nelms et al., 2019; Provencher et al., 2018; Reinold et al., 2021; Savoca et al., 2021). However, it should be emphasized that organisms ingest plastic particles and other microlitter components (i.e., additives, plasticizers, natural fibers, non-plastic synthetic fibers (APFs)) according to their mouthparts' size. Hence, invertebrates, the prey of other organisms in the food web, usually ingest SMPs and other particles less than 100 μm in size; these particles can then enter the food web and may accumulate and pose a threat to organisms of higher trophic levels, including humans (Carbery et al., 2018).

Plastic polymers have the most diverse uses, such as fishing equipment, fishing nets, boat coating, boat hulls, marine ropes, piping, gaskets, pumps, carpeting, paint, and coating. Additives and plasticizers are used to make plastic polymers achieve specific properties. Plastic polymers can be directly released into the environment as microscopic particles, and their primary sources are washing machine exhausts, tire wear, and cosmetic products (Corami et al., 2020a; Knight et al., 2020; Napper et al., 2015). Exposure to ultraviolet rays from the sunlight, erosion, and abrasion from wind and waves can further fragment large plastic particles (Andrady, 2017; Auta et al., 2017; Jiang, 2018); this fragmentation into secondary MPs may promote their uptake by organisms and their entrance into food webs (Beiras et al., 2021; Corami et al., 2020b, Corami et al., 2021).

Potential adverse effects of MPs on the organisms can be a) physical (i.e., ingestion and consequent pseudo-satiation), b) chemical, due to additives and plasticizers blended during plastic manufacturing and contaminants adsorbed during the permanence in the environment, and c) biological, due to the possible presence of pathogens in the biofilm formed during the permanence in the environment (Rodrigues et al., 2019; Tang et al., 2020). The leaching of additives and plasticizers can significantly contribute to toxic effects on different organisms (Beiras et al., 2021).

However, the presence of SMPs and other components of microlitter fish's gastrointestinal tract (GIT) has not been investigated yet. There is evidence of the occurrence of MPs in GIT of several commercial fish species (Avio et al., 2015; Baalkhuyur et al., 2020; Bimalli Koongolla et al., 2020; Hanachi et al., 2019; Karbalei et al., 2019; Neves et al., 2015; Sbrana et al., 2020; Thiele et al., 2021; Zakeri et al., 2020; Mistri et al., 2022). GITs are not usually eaten, but they are employed to produce fish meals and fish oil employed as food supplements (Castelvetro et al., 2021; Karbalei et al., 2020; Thiele et al., 2021). It should be noted that laboratory studies showed translocation of SMPs into other organs (von Moos et al., 2012; Avio et al., 2015). Hence, fish consumption can be an indirect yet hazardous, relevant route of exposure to MPs and SMPs in humans.

This study aims to develop a suitable pretreatment method (i.e., extraction and purification) to simultaneously collect ingested SMPs, additives, plasticizers, non-plastic synthetic and natural fibers, and other components of micro-litter (APFs) in commercial teleost fish species. The fish species investigated in this study are Engraulis encrasicolus (Linnaeus, 1758; European anchovy); Sardina pilchardus (Walbaum, 1792; European pilchard); Solea solea (Linnaeus, 1758; Common sole) Mullus surmuletus (Linnaeus, 1758; surmullet); and Sparus aurata (Linnaeus, 1758; Gilthead seabream).

The fish species under study are extensively distributed, from the Northeastern Atlantic to Southeastern Atlantic, including the Mediterranean sea, Black Sea, and Sea of Marmara. They are fished or reared in the eastern Mediterranean (FAO sub-areas 37.2.1 and 37.3.1). They are appreciated and commonly consumed unprocessed in all Mediterranean countries at least, and as processed products, their consumption may be widely expanded (EUMOFA, 2019; FAO, 2020, FAO, 2020). Besides, they can be bioindicators of MP pollution (Bray et al., 2019; Fossi et al., 2018, Fossi et al., 2020; Palazzo et al., 2021), and they may be bioindicators of pollution by APFs and SMPs.

The second goal of this research is to develop a suitable analytical method for the quantification and concurrent characterization (i.e., polymer identification) of ingested APFs and SMPs.

In developing a suitable pretreatment method, high temperatures were avoided since they can deeply influence the characteristics of a polymer according to the polymer's glass transition temperature (Tg). Tg is the critical temperature at which a material changes its features from “glassy material” to “rubbery material” (Corami et al., 2020b). For instance, Tg of polyamide (PA) is 56 °C (according to Broudin et al., 2015, this is the most widely accepted Tg of dry nylon 6); hence, heating at higher temperature severely contributes to polymer's denaturation, preventing a valid and appropriate polymer identification when using FTIR or Micro-FTIR. For the same reason, strong acids, alkali, or other strong oxidant agents were not employed to avoid further degradation/denaturation of polymers and APFs.

The third focus of this research is to investigate potential variability and differences in SMPs and APFs ingested by the five commercial teleost fish species chosen.

Section snippets

Sample collection

The five commercial teleost fish species under exam were bought at the fish market. E. encrasicolous, S. pilchardus, and M. surmuletus were acquired directly from fishers working along the North Adriatic Sea coasts (Fig. 1). S. solea and S. aurata were purchased in a local fish market in North-Eastern Italy; S. solea was fished in the North-Eastern Adriatic Sea, while S. aurata came from an aquaculture facility in Turkey and was acquired at a fish market in the Venetian mainland (Fig. 1). Other

SMPs in the GIT of commercial fish species

All the polymers identified in the five commercial species and their acronyms are shown in Table 1. SMPs, MPs, and APFs were not detected in reagent and procedural blanks, and contamination was hence minimized.

The abundance of SMPs (SMPs/g GIT ww, with Poisson's confidence interval) in the five commercial species studied are shown, together with their weight (mg/g GIT ww), in Fig. 2(a and b). The highest abundance of SMPs was observed in M. surmuletus, while the lowest was observed in S. aurata

Conclusions

As far as we are aware, it is the first study where APFs, i.e., additives and plasticizers, in the GIT of fish were identified and quantified.

To the best of our knowledge, this is the first study that allows the concurrent identification and quantification of SMPs (MPs < 100 μm) and APFs (particles < 100 μm), employing the same pretreatment and the same analytical method.

Correct quantification of particles less than 100 μm without under- or overestimation was possible thanks to their optimal

Funding

This research was partially supported by the Research Incentive Fund – FIR 2018 of the University of Ferrara attributed to C. Munari.

CRediT authorship contribution statement

Fabiana Corami: Conceptualization, Investigation, Methodology, Visualization, Data curation, Validation, Formal analysis, Writing – original draft, Writing – review & editing. Beatrice Rosso: Conceptualization, Investigation, Methodology, Data curation, Validation, Writing – review & editing. Andrea Augusto Sfriso: Conceptualization, Methodology, Resources, Writing – review & editing. Andrea Gambaro: Supervision. Michele Mistri: Conceptualization, Supervision. Cristina Munari:

Declaration of competing interest

The authors declare that they have no known competing financial interests or personal relationships that could have influenced the work reported in this paper.

Acknowledgments

Elga Lab water, High Wycombe UK, supplied the pure water system used in this study.

The authors thank the fishers for their support in the project activity.

The authors would like to thank two English-speaking reviewers for carefully editing the proper English language, grammar, punctuation, spelling, and style.

The authors would like to thank an anonymous reviewer and Chief editor Dr. François Galgani for their insightful comments that helped improve this manuscript.

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