Ecotoxicological investigation of the effect of accumulation of PAH and possible impact of dispersant in resting high arctic copepod Calanus hyperboreus

Highlights

• Effects of PAH exposure on the high Arctic copepod Calanus hyperboreus.

• Effects were examined on depuration, egg production and mortality on resting female copepods.

• Accumulation of PAHs were lower than expected, leading to low bioconcentration factors.

• Depuration and metabolism of PAHs were observed, although the copepods still contained PAHs after 77 days.

• Bioaccumulation of PAHs in Calanus hyperboreus can constitute a risk in the Arctic food chains.

Abstract

Due to high lipid content and a slow metabolism, there is a higher risk of bioaccumulation of oil compounds in Arctic than in temperate copepods. There is also a concern that the bioavailability of oil compounds is higher when oil is dispersed with dispersants. The purpose of this project was to increase the knowledge on how the use of dispersants on an oil spill may affect the passive uptake of PAHs in resting high arctic copepods using Calanus hyperboreus as a model organism. To evaluate this, resting high arctic C. hyperboreus were caught in Disko Bay at > 250 meters depth, November 2013, and subsequent experimental work was initiated immediately after, at nearby Arctic Station at Disko Island Western Greenland. C. hyperboreus females were incubated in phenanthrene (111, 50 and 10 nM), pyrene (57, 28 and 6 nM) and benzo(a) pyrene (10, 5 and 1 nM) for three days in treatments with and without oil (corn oil) and dispersant (AGMA DR372). After exposure, the highest measured concentrations of respectively phenanthrene, pyrene and benzo(a) pyrene in the copepods were 129, 30 and 6 nmol PAH g female⁻¹. Results showed that with addition of oil and dispersant to the water, the accumulation of PAH was significantly reduced, due to the deposition of the PAHs in the oil phase, decreasing the available PAHs for copepod uptake. While PAH metabolites and a depuration of the PAHs were observed, the copepods still contained PAHs after 77 days of incubation in clean seawater. Differences of treatments with and without oil and dispersant on the egg production were not statistically conclusive, although it is the most likely an effect of the highly variable day-to-day egg production between individual copepods. Equally, although there was an indication that the addition of dispersant and oil increased the mortality rate, there was no statistical difference.

Introduction

The use of dispersants is an oil spill response technique which increases the natural potential for removing spilled oil from the sea surface by dispersion of the oil in the water column. Because dispersant can be deployed from aircraft it has a promising potential in remote and icy high Arctic areas where mechanical recovery of spilled oil is almost impossible. However, pelagic organisms in the water column will be exposed to the components in the dispersed oil and the dispersant. It is therefore, very important to study the potential toxic effect of realistic concentrations of dispersed oil and dispersant on pelagic organisms to be able to minimize the ecological impact of an oil spill.

Calanus spp. are ideal high Arctic zooplankton model organisms due to their slow metabolism, high lipid contents and general adaptations to the highly variable Arctic seasons. Calanus spp. are also the predominant copepods in the Arctic areas—in the Disko Bay, the sampling site for this study, Calanus spp. constitute >90% of the zooplankton biomass in the upper 50 meters of the water column from May to July (Nielsen and Hansen, 1995, Auel and Hagen, 2002, Hopcroft et al., 2005, Thor et al., 2005, Madsen et al., 2001, Madsen et al., 2008). Arctic Calanus spp. copepods occupy a key role in high arctic seas as food for fish, sea birds and marine mammals (Hirche, 1991, Nielsen and Hansen, 1995, Hirche and Niehoff, 1996, Falk-Petersen et al., 2009). Calanus spp. spend the winter at deep water hibernating, but ascend prior to the phytoplankton spring bloom, during which they feed to re-fill their energy reserves for the next winter (Scott et al., 2000, Lee et al., 2006, Swalethorp et al., 2011). Calanus hyperboreus produce most of their eggs while still in the deep waters utilizing the lipid stores they build up during the previous summer (Hirche and Niehoff, 1996, Henriksen et al., 2012). Oil exposure during winter may therefore, affect the egg production and the offspring directly through surface exposure while exposure later in the year also may be through the food in-take.

Preliminary results indicate that there is a very high risk for accumulation of oil components in arctic copepods, probably due a high lipid content (>60% of body mass at the end of summer) and a slow metabolism (Scott et al., 2000, Lee et al., 2006, Swalethorp et al., 2011, Nørregaard et al., 2014). There is also an indication, i.e., minor or no significant differences, that exposure to oil compounds can affect hatching of Calanus spp. nauplii (Jensen et al., 2008, Hjorth and Nielsen, 2011, Grenvald et al., 2012, Nørregaard et al., 2014). A high accumulation rate increases the risk of toxic effects on the copepods and offspring as well as the risk for exposure of fish, bird and marine mammals that feed on the copepods.

PAHs are hydrophobic contaminants with mutagenic and carcinogenic characteristics. Because of the additive nature of PAH toxicity, i.e., PAHs have the same mode of action, even small concentrations can potentially impair survival of copepods (Barata et al., 2005). Sublethal effects of PAHs include reduced reproduction and feeding rates and disturbance of cell membrane fluidity resulting in non-polar narcosis leading to decreased activity and ability to react to stimuli (van Wezel and Opperhuizen, 1995, Lotufo, 1997, Jensen et al., 2008).

Marine animals can take up PAHs both passively, i.e., through diffusion, and actively, e.g., through feeding. The speed of the diffusive uptake is primarily governed by the octanol-water partitioning coefficient (log K_ow), which is a measure of the hydrophobicity of the compound, which consequently affects its bioavailability. The importance of active uptake though feeding or other routes is an on going discussion and conflicting results have been reported, although it is known that crude oil droplets, when suspended in the water or attached to plankton, can be ingested by zooplankton species (Almeda et al., 2013 and references therein). Several studies have shown results suggesting that passive partitioning is the dominating process for accumulation of hydrophobic compounds in C. hyperboreus (Fisk et al., 2001; Hoekestra et al., 2002). Other studies, however, have reported results showing that feeding can increase the effects of exposure. In Nørregaard et al. (2014), results indicate an increased accumulation of pyrene in C. hyperboreus when fed and in Jensen et al. (2008) pyrene-exposed Calanus glacialis showed a decrease in egg production when fed but not when starved.

Pyrene has been shown to affect both the reproduction and grazing of female C. finmarchicus, C. glacialis and C. hyperboreus at concentrations above 100 nM (Jensen et al., 2008, Hjorth and Nielsen, 2011, Nørregaard et al., 2014). It is generally believed that PAHs do not biomagnify via the food-chain because both vertebrates and many invertebrates, e.g., crustaceans, can metabolize the parent compound via the cytochrome P-450 (CYP450) enzymatic system (Varanasi et al., 1989, Hylland, 2006). While the majority of marine invertebrates is believed to possess subsystems inducible by PAHs (Rewitz et al., 2006), they generally possess lower capacity for phase I metabolism than vertebrates, and there are large variations in CYP450 activity both between and within taxonomic groups (Hylland, 2006). Nørregaard et al. (2014) showed that at least trace amounts of the primary metabolite of pyrene, hydroxypyrene (OH-pyrene) were present in C. hyperboreus females exposed to pyrene, indicating that C. hyperboreus, at least to some degree, is able to metabolize pyrene.

The common concerns regarding the use of dispersants are the dispersant-increased bioavailability of oil in the water column and the potentially added toxicity caused by the dispersant (Hansen et al., 2012). Dispersants generally function by lowering the interfacial tension between oil and water, causing the formation of small oil droplets (10–50 m) even at low turbulence conditions (Lewis and Daling, 2001, NRC, 2005). These small oil droplets are within the size range for Calanus spp. ingestion, making the oil and dispersant bioavailable through the digestive system of the copepods (Conover, 1971, Gyllenberg, 1981, Hansen et al., 2009).

The purpose of this project was to investigate how the use of dispersants on oil spill may affect the passive uptake of PAHs in resting high arctic copepods using C. hyperboreus as a model organism. This was done by exposing resting female C. hyperboreus from the Disko Bay, Greenland, to PAHs in treatments with and without oil and dispersants, followed by quantification of the PAH concentrations in the copepods using fluorescent high pressure liquid chromatography (HPLC/F).