In vitro synthesis and macromolecular analysis of dextrans from Weissella confusa dextransucrase

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http://urn.fi/URN:NBN:fi:hulib-201709125375
Title: In vitro synthesis and macromolecular analysis of dextrans from Weissella confusa dextransucrase
Author: Wang, Yu
Contributor: University of Helsinki, Faculty of Agriculture and Forestry, Department of Food and Environmental Sciences
Publisher: Helsingin yliopisto
Date: 2017
Language: eng
URI: http://urn.fi/URN:NBN:fi:hulib-201709125375
http://hdl.handle.net/10138/222931
Thesis level: master's thesis
Discipline: Elintarvikekemia
Food Chemistry
Livsmedelskemi
Abstract: Dextrans are polysaccharides and mainly produced from lactic acid bacteria, which have great values in industrial applications. Dextrans with different molar mass and branches have different applications. In this thesis, the effects of different reaction factors on the molar mass and composites of dextrans had been studied. Weissella confusa VTT E-90392 dextransucrase was used to synthesize dextran with sucrose as substrates. The effects of sucrose concentration, enzyme concentration, reaction time, temperature and pH on dextran macromolecular properties were detected. Sucrose concentration, enzyme concentration and reaction time were selected as the most influential factors for the subsequent response surface modeling to analyze how they affect dextran properties, according to experimental design. 18 reaction conditions were conducted and the product mixtures containing dextran were analyzed by HPSEC. The effects of the three reaction factors on dextran production were also studied. The predictive contour plot showed that dextran conversion was positively proportional to the enzyme concentration and increased in the early stage, but decreased in the late stage. In the preliminary study of the effects of various factors on dextran profile, two or three main dextran size populations eluting at different volumes were found in HPSEC analysis. The 1st dextran size population (eluted around 10.5 ml) had the highest Mw ranging from 7 × 107 to 2 × 108 g/mol. The 2nd dextran size population (eluted at around 12.5 ml) had Mw ranging from 2 × 107 to 4×107 g/mol. Besides, there seemed to be a 3rd dextran size populations (eluted at 16.2 ml to 17.5 ml) and the Mw was within the widest range (5 × 106 g/mol to 1.3 × 107 g/mol).We kept the samples' 4 factors constant and one factor changed and compared the effect of each factor. Under conditions: low sucrose, dextransucrase concentration, low temperature, low pH or short reaction time, the amount of the 2nd dextran size population was in equal or higher amount than the 1st one and 3rd one hardly appeared. On the contrary, the 1st size population became predominant over 2nd one, and the 3rd one started to form. In the modeling experiments, the overall effects of reaction time, sucrose concentration and enzyme concentration on the molar mass and amounts of different dextran size populations were studied. In our study, as the sucrose and enzyme concentration increased, the 2nd dextran size population decreased. The amount of 3rd one was positively proportional to the sucrose and enzyme concentrations. At the early stage of reaction, the amount of 3rd one was positively proportional to the reaction time, but the amount of 3rd one was negatively proportional to the reaction time. We hypothesize that as the reaction proceeds, 2nd dextran size population can be elongated into the 1st population. It was not clear whether the 2nd dextran size population could combine with themselves to form 1st one (internal insertion) or the 2nd one could only be inserted with D-glucosyl units one by one (external insertion). At the end of synthesis, dextran can be degraded possibly by hydrolyzing D-glucosyl units externally, because there should be more than 3 dextran populations if the dextran was internally broken. Based on the above results, it might give some new clue about the process of dextran synthesis and the way the dextran was hydrolyzed.
Subject: Weissella confusa
dextran
dextran size population
SEC (Size-Exclusion Chromatography)


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