Photon Upconverting Nanophosphors: Unique Reporters for Optical Biosensing

Upconversion photoluminescence is a unique property of mostly certain inorganic materials, which are capable of converting low-energy infrared radiation into a higher-energy emission at visible wavelengths. This anti-Stokes shift enables luminescence detection without autofluorescence, which makes the upconverting materials a highly suitable reporter technology for optical biosensing applications. Furthermore, they exhibit long luminescence lifetime with narrow bandwidths also at the optical window of biomaterials enabling luminescence measurements in challenging sample matrices, such as whole blood. The aim of this thesis was to study the unique properties and the applicability of nano-sized upconverting phosphors (UCNPs) as reporters in biosensing applications. To render the inorganic nanophosphors water-dispersible and biocompatible, they were subjected to a series of surface modifications starting with silica-encapsulation and ending with a bioconjugation step with an analyte-recognizing biomolecule. The paramagnetism of the lanthanide dopants in the nanophosphors was exploited to develop a highly selective separation method for the UCNP-bioconjugates based on the magnetic selectivity of the high gradient magnetic separation (HGMS) system. The applicability of the nano-sized UCNPs as reporters in challenging sample matrices was demonstrated in two homogeneous sensing applications based on upconversion resonance energy transfer (UC-RET). A chemosensor for intracellular pH was developed exploiting UC-RET between the UCNP and a fluorogenic pH-sensitive dye with strongly increasing fluorescence intensity in decreasing pH. The pH-independent emission of the UCNPs at 550 nm was used for referencing. The applicability of the pH-nanosensor for intracellular pH measurement was tested in HeLa cells, and the acidic pH of endosomes could be detected with a confocal fluorescence microscope. Furthermore, a competitive UC-RET-based assay for red blood cell folic acid was developed for the measurement of folate directly from a whole blood sample. The optically transparent window of biomaterials was used in both the excitation and the measurement of the UC-RET sensitized emission of a near-infrared acceptor dye to minimize sample absorption, and the anti-Stokes detection completely eliminated the Stokes-shifted autofluorescence. The upconversion photoluminescence efficiency is known to be dependent on crystallite size, because the increasing surface-to-volume ratio of nano-sized UCNPs renders them more susceptible to quenching effects of the environment than their bulk counterpart. Water is known to efficiently quench the luminescence of lanthanide dopants. In this thesis, the quenching mechanism of water was studied using luminescence decay measurements. Water was found to quench the luminescence of UCNPs by increasing the non-radiative relaxation of the excited state of Yb3+ sensitizer ion, which had a very strong quenching effect on upconversion luminescence intensity.

Fractionation of wood hydrolysate by ultrafiltration membranes at high temperature

Ultrafiltration (UF) is already used in pulp and paper industry and its demand is growing because of the required reduction of raw water intake and the separation of useful compounds from process waters. In the pulp and paper industry membranes might be exposed to extreme conditions and, therefore, it is important that the membrane can withstand them. In this study, extractives, hemicelluloses and lignin type compounds were separated from wood hydrolysate in order to be able to utilise the hemicelluloses in the production of biofuel. The performance of different polymeric membranes at different temperatures was studied. Samples were analysed for total organic compounds (TOC), lignin compounds (UV absorption at 280 nm) and sugar. Turbidity, conductivity and pH were also measured. The degree of fouling of the membranes was monitored by measuring the pure water flux before and comparing it with the pure water flux after the filtration of hydrolysate. According to the results, the retention of turbidity was observed to be higher at lower temperature compared to when the filtrations were operated at high temperature (70 °C). Permeate flux increased with elevated process temperature. There was no detrimental effect of temperature on most of the membranes used. Microdyn-Nadir regenerated cellulose membranes (RC) and GE-Osmonics thin film membranes seemed to be applicable in the chosen process conditions. The Polyethersulphone (NF-PES-10 and UH004P) and polysulphone (MPS-36) membranes used were highly fouled, but they showed high retentions for different compounds.