Journal of Foraminiferal Research, v. 55, no. 3, p. 276–295, July 2025
A COMPARISON OF SEQUENTIAL EXTRACTION METHODS USED FOR BIOAVAILABILITY ASSESSMENT OF POTENTIAL TOXIC ELEMENT MICHAEL MARTÍNEZ-COLÓN1,*, BENJAMIN ROSS1,2, JEREMY D. OWENS3, GWENDOLINE DUONG4, OLUGBENGA T. FAJEMILA5, VINCENT M. P. BOUCHET4 AND MARÍA VIRGINIA ALVES MARTINS6,7 ABSTRACT
increased in recent years in response to environmental health assessments (Abd Malek & Frontalini, 2024). At present, the methods applied to assess potentially toxic element (PTE; e.g., As, Se) concentrations in sediments have generally been inappropriate because what is bioavailable to the foraminifers has been severely overlooked by most authors. Discrepancies exist between researcher methods when applying benthic foraminifera as bioindicators of PTE pollution and scaling to the pollutant’s bioavailability. No consensus exists to determine the bioavailability for foraminiferal bioaccumulation and, most importantly, which chemical extraction method is most appropriate. Studies have relied on using either very strong acid (Hydrofluoric and Perchloric), relatively weaker acid (three to one ratio of Hydrochloric and Nitric 5 Aqua regia), or x-ray fluorescence to determine “total” PTE concentrations (e.g., Yanko et al., 1998; Sulkowski & Hirner, 2006; EPA, 2007; Schintu et al., 2016; Oron et al., 2021; El-Kahawy & Mabrouk, 2023; Ganugapenta et al., 2025) from sediments. These approaches report both bioavailable and non-bioavailable PTEs indiscriminately and simultaneously combine them into a single concentration value (Fig. 1; Method C). According to Tessier et al. (1979), using total PTE extraction is not an accurate approach because it does not reflect the physicochemical behavior of PTEs. In addition, they speculated that when using total extraction (p. 844), “. . .all forms of a given metal have an equal impact on the environment; such an assumption is clearly untenable.” We agree with this since PTEs bound to organic matter, for example, should be readily bioavailable to the foraminifers since most are detritivores. Thus, this approach has been shown to not consider the bioavailability of PTEs (Quevauviller et al., 1993; Morillo et al., 2004; Arain et al., 2008; Martínez-Colón et al., 2009, 2017, 2018; Martins et al., 2020) in sediments and to inadvertently overestimate the impact of the pollutants on ecological responses of benthic foraminifera as seen in numerous studies (e.g., Bergin et al., 2006; Ferraro et al., 2006; Frontalini & Coccioni, 2008; Ayadi et al., 2016; Hess et al., 2020; Cong et al., 2022; Schmitz et al., 2024; Ganugapenta et al., 2025). Bioaccumulated PTEs in an organism are not related to the total concentration in the sediment but to what is biologically extracted (Tessier & Campbell, 1987), which is the case with benthic foraminifera.
Understanding potentially toxic element (PTE) bioavailability has eluded the research community for decades. Here we compare three PTE extraction methods that are widely used in foraminiferal pollution studies. Evident discrepancies between methods were observed when diversity indices and relative abundances were influenced differently by the same and, in some cases, by different PTEs depending on the method. This was reflected in the canonical correspondence analysis (CCA), where Cr-Ni (Method B) and Zn-Pb-Cu (Method C) had a respective positive and negative influence over the foraminiferal relative abundances while no positive correlations were observed for Method A. These discrepancies stem from each method extracting different concentration levels of the same PTE from the same sediment samples. We recommend bioavailable PTEs to be extracted from the following fractions: (1) mud-bound from Method A (F1- exchangeable); (2) organicbound from Method B (F3- oxidizable); and (3) completely avoid using Method C. INTRODUCTION In environmental micropaleontology, benthic foraminifera are bioindicators used in marine environmental assessments and monitoring. Their rapid ecological response to changes in abiotic stressors (e.g., salinity, pollutants) is a key trait that has been exploited when conducting anthropogenic pollution studies (e.g., Alve, 1995; Nigam et al., 2006; Frontalini et al., 2016; El-Kahawy et al., 2018; Bergamin et al., 2019; Barik et al., 2022). In addition, the use of benthic foraminifera in describing and assessing spatial/ temporal ecological conditions of coastal (e.g., Jorissen et al., 2018; Castelo et al., 2021; Fajemila et al., 2022a; Ghandour et al., 2025) and deep-water settings (e.g., Schwing et al., 2017; O’Malley et al., 2021) have steadily 1
School of the Environment, Florida A&M University, 1515 S Martin Luther King Jr. Blvd, FSH Science Research Center, Tallahassee, FL 32307 2 Rosenstiel School of Marine, Atmospheric, and Earth Science, University of Miami, 4600 Rickenbacker Causeway, Miami, FL 33149 3 Department of Earth, Ocean, and Atmospheric Science, Florida State University, 1011 Academic Way, Tallahassee, FL 32306 4 Université de Lille, CNRS, IRD, Univ. Littoral Côte d’Opale, UMR8187, LOG, Laboratoire d’Océanologie et de Géosciences, Station Marine de Wimereux, 59000, Lille, France 5 Department of Geological Sciences, Osun State University, 230261, Osogbo, Nigeria 6 Faculdade de Geologia, Universidade do Estado do Rio de Janeiro, UERJ, Av. São Francisco Xavier, 24, sala 2024A, Maracanã, Rio de Janeiro, RJ 20550-013, Brazil 7 Universidade de Aveiro, GeoBioTec, Departamento de Geociências, Campus de Santiago, 3810-193 Aveiro, Portugal * Correspondence author. E-mail: michael.martinez@famu.edu
OBJECTIVES This paper comprises two sections that address two major objectives. The first objective is to guide the researchers on the general background aspects of bioavailability. In addition, this section provides a brief explanation of PTE chemical fractionation in sediments and three methods of extraction widely used in pollution studies involving benthic foraminifera.
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