Adaptation of the Daphnia sp. acute toxicity test: miniaturization and prolongation for the testing of nanomaterials

Abstract

Manufacturing of nanomaterials (NMs) is often complex and expensive, and their environmental risks are poorly understood or even unknown. An economization of testing NMs is therefore desirable, which can be achieved by miniaturizing test systems. However, the downsizing of test vessels and volumes can enlarge the surface/volume ratio (SVR) which in turn can affect the bioavailable concentration of adsorbing substances like NMs. The present study focused on the miniaturization of the acute toxicity test with Daphnia magna. The adaptations were verified with three reference substances, the non-adsorbing potassium dichromate (K2Cr2O7) and as potentially highly-adsorbing substances silver nanoparticles (AgNPs) and silver nitrate (AgNO3). The miniaturized test was conducted in 24-well microtiter plates (MT) and simultaneously compared to the OECD standard test (ST). Furthermore, the test duration was prolonged from 48 to 96 h since NMs tend to show effects only after extended exposure. The toxicity of K2Cr2O7 and AgNPs continued to increase within the prolonged test span. The test comparisons with K2Cr2O7 did not reveal any significant differences between ST and MT. AgNO3 toxicity was significantly decreased in MT compared to ST due to the enlarged SVR. The toxicity of AgNPs in MT after 24 h was equal to ST. Contrary to our expectations an exposure longer than 24 h resulted in an increase of AgNP toxicity in MT, possibly due to enhanced dissolution of silver. Microtiter plates are appropriate alternative test vessels for the Daphnia sp. acute toxicity test; thus, its miniaturization is feasible. The enlarged SVR has to be taken into account since it can affect the toxicity of potentially adsorbing substances. Furthermore, the standard test duration of 48 h might underestimate the toxicity of many substances, especially of NMs.

Short-term exposure with high concentrations of pristine microplastic particles leads to immobilisation of Daphnia magna

“…In agreement with a study on nanoparticle toxicity (Dabrunz et al, 2011), immobilsation exceeding 10% was not observed within the recommended exposure time of 48 h (OECD guideline 202), but after 72 and 96 h. The pronounced increase in toxicity if exposure time is prolonged to 96 h is similar to another study (Baumann et al, 2014) and underlines the need for adaptations and different testing procedures on impacts of particles in general and microplastics in particular.…”

“…Like suggested in literature for testing of particulate material (e.g. Baumann et al, 2014), exposure length was prolonged from 48 to 96 h in all treatments. Pre-tests confirmed that neonates from our culture younger than 24 h survive for 96 h in the absence of food without showing immobilisation or abnormal behaviour like being trapped at the water surface.…”

Acute sensitivity of three Cladoceran species to different types of microplastics in combination with thermal stress

“…Comparison of 48 h and 96 h LC 50 values indicated a strong time dependence of toxicity, as has been previously suggested in a study assessing the acute toxicity of polyethylene microspheres to D. magna (Rehse et al, 2016). A similar observation was also made in a study investigating the acute exposure effects of nanomaterials to D. magna (Baumann et al, 2014). The marked increase in toxicity when the exposure time is prolonged to 96 h highlights the need for modifications of existing testing standards, which normally stipulate 48 h of exposure for acute toxicity assays (Rehse et al, 2016).…”

Behavior and chronic toxicity of two differently stabilized silver nanoparticles to Daphnia magna

“…For detAgNP in the same medium, colloidal instability and increasing agglomerate size as well as relatively stable small aggregates of about 60 nm size have been reported (Baumann et al, 2014b;Cupi et al, 2015;unpublished data). In the present study, agglomerate sizes were constant, but a bit larger than reported for the stable dispersions.…”