Optimisation of method for extraction and quantification of microplastics from freshwater sediments and biogenic structures

Mikroplastika predstavlja prijetnju ekosustavima jer dolazi u interakciju s organizmima, a dugoročne posljedice njenog utjecaja nisu poznate. Slatkovodni ekosustavi smatraju se značajnim u transportu mikroplastike, ali su nedovoljno istraženi. Tulari (Trichoptera, Insecta) su potencijalni bioindikatori za mikroplastiku u slatkovodnim ekosustavima jer grade kućice od svile kojom lijepe čestice sedimenta, ostatke biljaka, pa i mikroplastiku. Cilj ovog diplomskog rada je optimizacija metode za ekstrakciju i kvantifikaciju čestica mikroplastike u sedimentu i biogenim tvorbama slatkovodnih ekosustava. Testirano je više metoda i utvrđen je optimalni protokoli za: razgradnju organske sastavnice korištenjem otopine H2O2, izdvajanje mikroplastike iz smjese mikroplastika-mineralni ostatak otopinom NaBr, detekciju čestica mikroplastike uz bojanje i UV svjetlo. Korištenjem navedenog protokola testirane su kućice tulara i sediment iz pokusa u kojem su isti izlagani utjecaju mikroplastike te smjesi mikroplastike i farmaceutika. Unutar istih tretmana, nema razlike u broju i veličini čestica mikroplastike u sedimentu ni ugradnji u kućice različitih stadija tulara tijekom vremena. Nasuprot tome, nađena je razlika u veličini i broju ugrađenih čestica između različitih tretmana na kraju pokusa, koja je najvjerojatnije povezana s razlikama u adsorpciji, formiranju obraštaja, te sedimentacijom čestica mikroplastike. Navedeni rezultati ukazuju na nužnost provođenja dodatnih pokusa koji će utvrditi ponašanje čestica mikroplastike u uvjetima višestrukih stresora.Microplastic poses a threat for ecosystems because it interacts with organisms, however the long-term consequences are unknown. Freshwater ecosystems are considered as important in microplastics transport but are insufficiently researched. Caddisflies (Trichoptera, Insecta) are potential bioindicators for microplastics in freshwater ecosystems because they build protective cases using larval silk to glue sediment particles, plant parts, and even microplastics. The goal of this thesis is to optimise the method for extraction and quantification of microplastic particles in sediment and biogenic formations of freshwater ecosystems. I tested different methods and determined the optimal protocol for: digestion of the organic component (H2O2 treatment), separation of microplastics from the microplastic-mineral residue mixture (NaBr solution); detection of microplastic particles (staining and UV light). The protocol was then used to test incorporation of microplastics into the caddisfly protective cases and sediment from a previously conducted microcosm experiment with two treatments: microplastics and microplastics+pharmaceuticals. There were no differences in number and size of microplastics found in the sediment of different treatments. Moreover, there were no differences in microplastics incorporated into the caddisfly cases over the course of the experiment within each treatment. However, differences were found in both number and size of microplastics incorporated into the pupal cases at the end of experiment between treatments. These are most likely related to differences in adsorption, biofilm growth and sedimentation of microplastic particles. Thus, further research is needed to determine whether microplastics behave differently in the environment if found in combination with multiple stressors