Effect of nanoplastics in the marine organism Tisbe battagliai

Plastic is a material made out of polymer chains derived from oil and gas, and together with additives these polymers have numerous applications. Plastic is used in daily life as wrapping around food, in cosmetics and in clothes, and is inevitable nowadays. However, of all the 322 million metric tons of plastic that are produced annually, between 4.8 and 12.7 million metric tons are estimated to enter the oceans every year. In the oceans, plastic debris can be degraded by physical and chemical processes to micro- and nanoplastics (MP and NP). These small particles (<5 mm) may be a threat to organisms in the marine environment, as they can affect their development, reproduction and survival. Microcrustaceans are widespread in the oceans with both benthic and pelagic species, which may be affected by plastic particles in the oceans. Many of the microcrustacean species are filter feeders and do not select their food intake. These animals have a high risk of ingesting NP particles (<100 nm), and can thus be suitable species to test and evaluate for ecotoxicological effects of NPs. In this thesis, the toxicity of the NP particles polystyrene (PS) and poly (methyl methacrylate) (PMMA), both non-functionalised and with functional carboxylic (PS-COOH and PMMA-COOH) and aminated (PS-NH2) groups were studied in the copepod Tisbe battagliai to assess potential adverse effects of these particles. The NP particles were characterised by dynamic light scattering (DLS), where size (Z-average), surface charge and polydispersity were measured in Milli-Q water (MQW) and natural seawater (NSW), to describe their properties. Acute mortality and oxidative stress were determined for T. battagliai when exposed to the same NP particles. The NPs PS-NH2 and PMMA-COOH were found to not aggregate in NSW (Z-average < 140nm), while the other particles formed agglomerates (Z-average > 1800nm). Whereas most particles displayed a negative surface charge, PS-NH2 was positively charged in both medias. To test acute mortality, copepods were exposed to different concentrations of NPs (0, 0.5, 1, 5, 10, 25, 50 and 100 g/mL) in NSW for 48 hours. PS-NH2 was shown to be the most toxic particle, with an EC50 value of 7.8 g/mL. This particle was therefore chosen for further studies to determine reactive oxygen species (ROS) formation and lipid peroxidation as markers for potential oxidative stress and damage. To detect ROS formation and lipid peroxidation within the copepods, in vivo tests using fluorescent probes were conducted. Copepods were exposed to PS-NH2 in several concentrations and three different fluorescent detecting probes (DHR123, H2DCFDA and C11-BODIPY) were added. All three probes showed a significant rise in fluorescence compared to the control for the highest concentration tested (100 g/mL), but the methods were not successfully optimised and need to be further developed to get more precise results. Suggestions for improvements in the method and future research are proposed to get an enhanced ecotoxicologial assessment of micro- and nanoplastics. Overall, results suggest that the NPs PMMA-COOH and PS-NH2, which do not agglomerate, have the greatest potential of entering the copepods through ingestion because of their small size. Results also suggest that the positive surface charge of PS-NH2 increased the toxicity of the polymer. Given the results from the acute test where PS-NH2 was the most lethal particle tested, and from the significant level of fluorescence measured in the probe test, it is possible that the mortality of copepods exposed to PS-NH2 is caused by oxidative stress formed inside the animals.