Biokatalys medför ett alternativt tillvägagångsätt för att kunna utforska och utveckla kemiskt syntetiserade vägar för produktionen av eftertraktade kemikalier, där hållbarhet och miljön tas till beaktan. I denna studie undersöktes potentialen av en enzymatisk kaskad för produktion av 5-hydroximetylfurfurylamin (HMFA). HMFA är en förening med tillämpningar inom flera industrier som till exempel jordbruks- och läkemedelsindustrin. Den enzymatiska kaskaden består av två reaktioner, varav den första involverar dekarboxylering av lysin med användning av lysindekarboxylas för att producera en så kallad ”smart amindonator” kadaverin. Den andra reaktionen i kaskaden består utav ett transaminas från Silicibacter pomeroyi (SpTA) som konverterarar 5-hydroximetylfurfuryl (HMF) till HMFA med hjälp av det framkallade kadaverinet från den första reaktionen i kaskaden. En enzymatisk kaskad tillåter mildare reaktionsbetingelser, mindre avfall och energisnål användning som därmed minskar miljöpåverkan samtidigt som det beaktar några av dem 12 principerna av grön kemi. Det uppstod utmaningar som hindrade slutförandet av den enzymatiska kaskaden, men trots detta erhölls värdefulla insikter. Denna studie belyser de invecklade reaktionsmekanismerna och några av de svårigheterna med immobilisering av enzym på EziG bärare. Trots att den avsedda kaskaden inte slutfördes, gav lärdomarna nya perspektiv samt potentiella områden att fortsätta undersöka för framtida framsteg inom biokatalys. / Biocatalysis is a promising alternative to chemical synthesis routes for high value chemicals which considers the sustainability and environmental aspect. In this study the feasibility of utilizing an enzymatic cascade for the production of 5-hydroxymethylfurfurylamine (HMFA) was explored. HMFA is a compound with diverse applications in industries such as agriculture and pharmaceuticals. The cascade consists of two main reactions, the first of which involves the decarboxylation of lysine using a lysine decarboxylase to produce cadaverine. The cadaverine produced will then be utilized as an amine donor in the second reaction, which involves the use of a transaminase derived from Silicibacter pomeroyi (SpTA) together with 5-hydroxymethylfurfural (HMF). This cascade considers the principals of green chemistry such as milder reaction conditions and less waste, hence aiming to reduce the environmental impact. Although there were challenges preventing the completion of the enzymatic cascade, valuable insights were gained. The contribution of this study sheds light on the intricate reaction mechanisms and some of the key difficulties with enzyme immobilisation. While the intended cascade was not finalized, the lessons learned will provide for new perspectives and potential future advancements in biocatalysis.

The aim of this thesis was to determine the content of biogenic amines (BA)and polyamines (PA)in selected types of cheese and the influence of storage time on the content of biogenic amines.

The thesis deals with the use and effectiveness of some less common methods of conservation of fish meat. The formation of biogenic amines in meat is observed in connection with the non-traditional preservative methods. Amines can serve as indicators of protein degradation. The quality of fish was considered in connection with the increasing content of selected biogenic amines (putrescine, cadaverine, spermidine, spermine, 2-fenylathylamine, histamine, tyramine and tryptamine). Ultra performance liquid chromatography (UPLC) was used as the method for determination of biogenic amines. Amines were derivatized by dansylchloride before their UPLC separation. The fish flesh was vacuum-packed. Samples were stored for several weeks in a thermostat at the selected storage temperature after the application of selected preservative technique. Beta-irradiation and high hydrostatic pressure were used for the preservation of fish flesh. Control samples were not exposed to the â-irradiation and high pressure. The organoleptic properties were studied for all samples (smell/odor, insight and shape). Beta-irradiation was applied to fish meat of common carp (Cyprinus carpio) and rainbow trout (Oncorhynchus mykiss). Both these species of freshwater fish are economically significant. Carp and trout are the species being mostly consumed in the Czech Republic. Fish meat is considered to be provided the flesh is fresh. A testing series of samples was created with common carp to determine the appropriate dose of â-irradiation. The maximum permissible dose of irradiation for fish meat is 3 kGy. Fish samples were exposed this dose in the first experiment. The dose of irradiation was reduced in following experiments based on the experience from the initial experiment. The doses of 0.25, 0.5, 0.75, 1.0 and 2.0 kGy were applied to rainbow trout. The value of 0.75 kGy of â-irradiation or higher (1.0, 2.0 and 3.0 kGy) prolonged the shelf life of fish meat, which was stored for three months (98 days). Prolonging of the shelf life of fish meat to approximately 98 days at 3.5 °C is redundant from technical point of view. For that reason lower doses 0.25, 0.5 and 0.75 kGy were tested in more detail in the repeated experiment with carp meat. Lower doses of â-irradiation are considered to be more acceptable and-at the same time-sufficiently effective for delaying the beginning of degradation processes. 6 High hydrostatic pressure was applied to meat of common carp (Cyprinus carpio), rainbow trout (Oncorhynchus mykiss) and pike (Exos lucius). Pike is another very popular kind of freshwater fish. Pike flesh is very tasty, but in spite of this, pike is not so much popular among consumers compared to carp and trout. The cause is its high price. Samples of pike were stored at standard temperature 3.5 °C and also at higher temperature 12 °C (unlike experiments with â-irradiation). Lower temperature of storage (3.5 °C) followed the conditions of storing of fish meat in industrial refrigeration facilities and households. The high pressure might not be sufficient for preservation at higher storage temperatures. This assumption was based on available information. Samples were treated by high pressure and stored at both 3.5 °C and 12 °C to verify this assumption. Higher temperature simulated either failure of refrigeration equipment or unsuitable store temperature of meat. In all species selected freshwater fish two levels of high pressure were applied ? 300 and 500 MPa. Both levels had significantly reduced the formation of biogenic amines, especially in samples stored at 3.5 °C. At this temperature, the effect of 300 and 500 MPa delayed start of degradation processes in fish meat by 3?4 weeks. At 12 °C and 500 MPa, high pressure extended the sustainability of meat by no more than one week. 500 MPa is effective treatment at the lower temperature of 3.5 °C. High pressure is not reliable preservative techniques at higher temperature.