20.500.12556/DKUM-90245 Proizvodnja ADH iz Saccharomyces cerevisiae gojene na kompleksnem gojišču diplomsko delo univerzitetnega študijskega programa I. stopnje Production of ADH from Saccharomyces cerevisiae grom on complex culture medium Namen diplomskega dela je bil proučiti vpliv različnih parametrov, kot so temperatura, pH in sestava rastnega medija na rast Saccharomyces cerevisiae, gojene na kompleksnem gojišču in vpliv parametrov na izražanje alkohol dehidrogenaze (ADH), celokupnih proteinov in njihovega antioksidativnega potenciala. V prvem delu smo optimirali temperaturo rasti kvasovke (28 °C in 30 °C) z namenom pridobitve ADH z visoko aktivnostjo. Ugotovili smo, da je bila aktivnost ADH višja v primeru, ko smo kvasovko gojili pri 28 °C. Nadalje smo optimirali sestavo rastnega medija ter sestavo medija (rastni medij ali različni pufri) v katerem smo izvedli razbitje celičnih sten kvasovke za pridobitev ADH. Primerjali smo rezultate gojenja v klasičnem rastnem mediju za kvasovke, ki vsebuje 10 g/L glukoze, z rezultati gojenja v rastnem mediju, kjer je bila glukoza zamenjana z različnimi količinami trsnega sladkorja v koncentracijah 5 g/L, 10 g/L ter 15 g/L. Prav tako smo primerjali učinkovitost homogenizacije v rastnem mediju, v 0,05 M pirofosfatnem pufru s pH 8,8 ter v 0,05 M natrijevem citratnem pufru s pH 4,8. Optimalne pogoje smo dosegli pri gojenju S. cerevisiae v rastnem mediju z 10 g/L trsnega sladkorja ter homogenizaciji kvasnih celic v 0,05 M natrijevem citratnem pufru pri pH 4,8. Izvedli smo tudi študijo vpliva pH rastnega medija na celokupno pridobljeno koncentracijo proteinov in ADH z visoko aktivnostjo iz celic kvasovk . Naprej smo kvasovko gojili v rastnem mediju pri pH = 6,6, nato pa smo s pomočjo kislin oz. baz uravnali pH rastnega medija na pH 6,0 ter pH 7,0. Ugotovili smo, da sta tako aktivnost ADH kot tudi celokupna vsebnost proteinov najvišji, kadar je bila fermentacija izvedena v mediju pri pH = 6,61, medtem ko je bila aktivnost ADH najnižja pri gojenju v mediju s pH 7,0. The aim of the thesis was to study the influence of various parameters, such as temperature, pH and the composition of the growth medium on the growth of Saccharomyces cerevisiae grown in complex medium, as well as studying the influence on alcohol dehydrogenase (ADH) expression and the production of total proteins and their antioxidant potential. In the first part, we optimized the temperature by comparing the growth of yeast at 30 °C and 28 °C with the aim of obtaining ADH with high activity. We found that ADH activity was higher when our yeast was grown at 28 °C. Furthermore, we optimized the composition of the growth medium and the composition of the medium (growth medium or various buffers) in which we broke down the yeast cell walls to obtain ADH. We compared the results of cultivation in the classical growth medium for yeasts, which contains 10 g/L glucose, with the results of cultivation in the growth medium, where we replaced glucose with different amounts of cane sugar in concentrations of 5 g/L, 10 g/L and 15 g/L. We also compared homogenization efficiency in growth media, in 0.05 M pyrophosphate buffer with pH 8.8 and in 0.05 M sodium citrate buffer with pH 4.8. Optimal conditions were achieved when growing S. cerevisiae in a growth medium with 10 g/L cane sugar and homogenizing the cells in 0.05 M sodium citrate buffer at pH 4.8. We also carried out a study of the influence of the growth medium´s pH on the total concentration of proteins and on the production of ADH with high activity obtained from yeast cells. Firstly, our yeast was cultivated in growth medium without adjusting the pH (pH 6.6), and then with the help of acids or bases the pH of the growth medium was adjusted pH 6.0 and pH 7.0. We found that both ADH activity and total protein content were highest when cultivation was carried out in growth medium at pH = 6.6, while ADH activity was lowest when cultivated in growth medium at pH 7.0. S. cerevisiae ADH encimi ekstrakcija homogenizacija optimizacija antioksidativnost S. cerevisiae ADH enzymes extraction homogenization optimization antioxidative properties true true false [A. Pajtler] Slovenski jezik Angleški jezik Diplomsko delo/naloga 2024-08-27 12:42:36 2024-09-20 09:38:00 2025-01-09 03:08:46 0000-00-00 00:00:00 2024 0 Maribor 0 Maribor 1 spletni vir (1 datoteka PDF (VIII, 36 f.)) 0000-00-00 NiDoloceno NiDoloceno NiDoloceno 0000-00-00 0000-00-00 0000-00-00 577.15(043.2) 221290499 UN_Pajtler_Andrej_2024.pdf UN_Pajtler_Andrej_2024.pdf 1 CC278BA3578BA985CD99FD65056165BD 2feab8b0057a9b25067fd159d057fc7cbc41467a4eaa955eb4805daf457c65b9 a2c53ce9-70ff-11ef-93e7-00155d000105 https://dk.um.si/Dokument.php?lang=slv&id=185033 Fakulteta za kemijo in kemijsko tehnologijo 0 0 0