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Microplastic pollution is one of the most critical environmental issues in the world right now. Many studies about human exposure to these micro-and nanoplastics (MNP) and the toxic effects they can cause are being conducted at this moment. Quantification and identification of these MNP lie at the bottom of solving this global problem. The most popular identification methods are Fourier-transform infrared spectroscopy and Raman spectroscopy. These techniques are lacking in non-destructive analysis with high sensitivity and low background noise. This thesis aims to develop an optical spectroscopy method to detect and identify microplastics after staining with Nile Red. The latter is a lipophilic solvatochromic dye. Staining will cause the particle to fluoresce, with its emission spectrum depending on its polarity. More polar plastics will have a more red-shifted emission maximum. Via hyperspectral analysis, the MNP can be identified based on this spectrum. Fluorescence lifetime imaging microscopy is also implemented. The fluorescence lifetime is the time that a fluorophore spends in the excited state, after absorbing a photon, before returning to the ground state. The lifetime depends on the fluorophore's micro-environment and therefore is also different for each MNP type. Differentiation is possible based on this lifetime value, but a graphical, fit-free phasor approach is also used. This allows for the mapping of the lifetime distribution in an image. When two different MNP types are present in one sample, they will each have a separate distribution cloud on the phasor plot, allowing for the graphical distinguishment of MNP. The plastics used are PP, PVC, HDPE, PET, and PS, these samples contain a range of shapes and sizes. A bought suspension of pure PS polymer particles of a fixed shape and size of 0,6 μm was also examined. The spectral data were processed in two ways. By calculating a ‘weighted average’ with the intensity values at each wavelength. This resulted in each MNP being attributed a weighted average value. After recording spectra of different particles, a statistical analysis, ANOVA and t-test, was done. This showed that the weighted averages of PP & HDPE and PS (self-made) & PVC do not differ significantly. The second option is through comparing the spectra themselves, using a relative similarity percentage. Because of the low similarity MNP gave when compared with themselves, only MNP with a significantly large similarity percentage could be distinguished. The same statistical tests were done on lifetime data. Measurements done on different particles of the same plastic showed that only HDPE & PET lifetimes did not differ significantly. Multiple measurements done on the same particle showed slightly different lifetimes. It could be concluded that HDPE & PET and PS (self-made and 0,6 μm) & PET cannot be differentiated. Finally, phasor plot analysis of samples containing two different plastics showed a possibility of recognizing MNP types based on the location of their phasor cloud on a phasor plot. When combining these two techniques, making use of the ’weighted average’ value and lifetime analysis, it could be possible to identify each plastic. The combination with similarly weighted averages, PP & HDPE give significantly different fluorescent lifetimes values and are possible to separate on a phasor plot. Still, both techniques have some flaws and improvement is possible.
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As society becomes more reliant on the use of plastics, consumption grows every year and waste increases proportionally. Plastics are released in nature, polluting the environment, biological life, and ecosystems. Once discarded, plastics degrade to microplastics (<5 mm) and nanoplastics (<1 µm) due to weathering processes and photodegradation. Micro- and nanoplastics (MNP’s) can enter the human body via several pathways, and induce toxicological effects on human health. Very little is known regarding these adverse effects due to a lack of appropriate detection techniques. Also, localization of particles in biological samples is required to investigate local effects, meaning that the preservation of bio-context is essential to achieve this. Staining of MNP’s with Nile Red (NR), a solvatochromic dye, can serve as a fluorescent labeling technique for MNP’s, enabling high-resolution particle detection and polymer identification. However, since NR has a high affinity for lipophilic carbon structures, organic materials like lipids get co-stained, inducing false-positive results. Furthermore, biological samples often are opaque, limiting laser transmission due to light scattering. Thirdly, autofluorescence causes a lot of background signal, interfering with the signal obtained from small plastics and making detection of MNP’s more challenging. For this reason, optical clearing protocols are needed to resolve these issues by extracting these structures from biological samples. 3D spheroids (co-cultured with MNP’s) and tissue samples were prepared and anchored in an acrylamide gel. This enabled the preservation of bio-context, allowing particle localization in a subsequent stage. Next, optical clearing steps were added and different protocols were compared, starting from a protein digestion step. This resulted in a more effective removal of lipids, which enhanced laser transmission (decrease in scattering) and reduced autofluorescence. This facilitates future research towards MNP-detection with NR staining, as false positive results get ruled out more effectively.
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The dependency on fossil fuels and associated greenhouse gas emissions force us to move towards renewable energy sources. A part of the solution lies in the use of hydrogen as an energy carrier. Water can be split, by electrolysis from renewable sources, into hydrogen and oxygen. The production of renewable energy is time and area dependent. Moreover, vast amounts of hydrogen will be needed to drive the steel industry, automotive, ammonia,…. The major challenge in storing and transporting hydrogen is overcoming the high energy needed. Therefore alternatives are needed. Hydrogen-carrying chemical molecules possess hydrogen in their structure. Out of the different storage methods analyzed in this thesis, hydrogen-carrying chemical molecules have the highest hydrogen storage capacity and can be stored and be transported in less energy-demanding conditions. One of the most promising hydrogen carriers is ammonia. Being well familiar with handling and shipping vast amounts of ammonia globally, the infrastructure for ammonia transport is already established. Ammonia is produced from hydrogen and nitrogen in the Haber-Bosch process and can easily be converted back to hydrogen and nitrogen by various routes. A main advantage is the fact that nitrogen is the dehydrogenated carrier which is endlessly supplied and can vent into the atmosphere without any problem, contrary to other (carbon-based) carriers such as methanol, where carbon dioxide is produced beside hydrogen. This thesis investigates the cracking of ammonia to hydrogen and nitrogen by electrocatalysis, which is the most cost-effective option in a decentralized Ammoniato-Hydrogen scenario. Three cell designs are discussed, tested and optimized. A solvent screening shows that DMSO is a suitable solvent due to the high ammonia and electrolyte solubility, conductivity and safety of use. A drawback is the limited oxidation potential of DMSO, which may not be exceeded since it yields hydrogen at the cathode according to our control experiments. To reduce the overall resistance caused by the solution, a proton exchange membrane is used, allowing the electrodes to be in close proximity. For the oxidation of ammonia to nitrogen in solution, Pt electrodes delivered the highest yield, followed by Pd wire and Au in addition to investigating the cracking of ammonia in the liquid. Gaseous, preliminary experiments were performed using Gas Diffusion Electrodes. Here it was shown again that Pt-based electrodes are superior to Au-based electrodes. Further investigation is needed to have more insights into the mechanism and to validate all the data.
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