Development of bioactive paper based on Trametes versicolor laccase: Master's thesis

The aim of this thesis was to develop bioactive paper based on Trametes versicolor laccase enzyme. Bioactive paper is a paper or paper-like product that includes functionalities based on selective reactions of bioactive compounds, such as enzymes. Laccases catalyze the oxidation of many phenolic substrates with simultaneous reduction of oxygen to water. Many of laccase substrates, such as ABTS (2,2'-azino-bis(3-ethylbenzthiazoline-6- sulphonic acid)) also change colour while oxidizing. Therefore, laccase based bioactive paper could be used for oxygen scavenging and oxygen indicating applications. These applications are further explored by reviewing previous studies and inventions. In this study, the bioactive ink composition was first developed. The stability and activity of T. versicolor laccase was determined on various solutions. Next, T. versicolor laccase was coated on different substrates with various polymers with and without additives. Various ink compositions and coating substrates were compared by evaluating the laccase activity of paper samples containing the different bioactive coatings. Besides, the surface profile of the paper samples was investigated by measuring roughness with profilometer. Finally, the indicator performance was examined. The colour changing reaction of ABTS was observed visually and instrumentally by applying ABTS solution to the papers containing laccase based ink. Based on the determination of the optimum pH for T. versicolor laccase activity (pH < 4.5 with ABTS substrate) and stability (pH 7 - 9), pH 6 was chosen for further use. Sulfo polyester resin, HZ1100D was a favourable coating component. The tests proved that laccase cannot retain its activity long time on papers without any ink. When laccase was coated with HZ1100D, laccase activity typically did not significantly decrease during eight weeks of storage. Additives enhanced the stability of laccase in solutions, but not on the paper with the coating. EUCA paper proved to be a good substrate for laccase based coating. The colour changing reaction of ABTS on EUCA paper was rapid and excellent colour change from colourless to dark green was observed on the paper samples containing laccase. The findings of the study indicate that T. versicolor laccase can be used together with HZ1100D polymer dispersion to prepare functional bioactive paper products. These kinds of bioactive paper products were found to be particularly useful for indicator applications. However, when used in oxygen scavenging applications, calculations suggest that more optimization is needed.