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Enzyme Catalyzed Oxidative Cross-Linking of Feruloylated Pectic Polysaccharides from Sugar Beet Kinetics and Rheology

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Enzyme Catalyzed Oxidative Cross-Linking of Feruloylated Pectic Polysaccharides from Sugar Beet Kinetics and Rheology Downloaded from orbit.dtu.dk on: Oct 06, 2021 Enzyme catalyzed oxidative cross-linking of feruloylated pectic polysaccharides from sugar beet Kinetics and rheology Abang Zaidel, Dayang Norulfairuz Publication date: 2012 Document Version Publisher's PDF, also known as Version of record Link back to DTU Orbit Citation (APA): Abang Zaidel, D. N. (2012). Enzyme catalyzed oxidative cross-linking of feruloylated pectic polysaccharides from sugar beet: Kinetics and rheology. Technical University of Denmark. General rights Copyright and moral rights for the publications made accessible in the public portal are retained by the authors and/or other copyright owners and it is a condition of accessing publications that users recognise and abide by the legal requirements associated with these rights. Users may download and print one copy of any publication from the public portal for the purpose of private study or research. You may not further distribute the material or use it for any profit-making activity or commercial gain You may freely distribute the URL identifying the publication in the public portal If you believe that this document breaches copyright please contact us providing details, and we will remove access to the work immediately and investigate your claim. Enzyme catalyzed oxidative cross‐linking of feruloylated pectic polysaccharides from sugar beet – Kinetics and rheology PhD Thesis Dayang Norulfairuz Abang Zaidel Enzyme catalyzed oxidative cross‐linking of feruloylated pectic polysaccharides from sugar beet – Kinetics and rheology This thesis was prepared by Dayang Norulfairuz Abang Zaidel Supervised by Professor Anne S. Meyer January 2012 Center for Bioprocess Engineering Department of Chemical and Biochemical Engineering Technical University of Denmark Enzyme catalyzed oxidative cross‐linking of feruloylated pectic polysaccharides from sugar beet – Kinetics and rheology PREFACE This thesis is submitted in fulfillment of the requirements for the Doctorate (PhD) degree at the Technical University of Denmark (DTU). The work presents in this thesis was conducted during my PhD study at the Department of Chemical and Biochemical Engineering, DTU from 1st August 2008 until 31st January 2012. The work was accomplished under the supervision of Professor Anne S. Meyer. The PhD study was financed by a scholarship from the Ministry of Higher Education Malaysia (MOHE) and Universiti Teknologi Malaysia (UTM). Most analytical work was performed at the Center for Bioprocess Engineering (BioEng) whereas macromolecular investigation was carried out at the DTU Food. We acknowledge Danisco A/S for supplying the materials for this study and Southern Danish University for performing the MALDI- TOF analysis. For completing this study, I am most grateful to Anne Meyer, for the thorough supervision, valuable comments and continuous support throughout this study. I have enjoyed the “boot camps” session the most. I would like to express my appreciation to Ioannis Chronakis who has assisted me a lot in the rheological part of the work. It has been an amazing three-and-a-half-year journey of my life, working and living in Denmark. I would like to thank all the people who have helped me during this study, for the analytical work, sharing of ideas, and inspirational discussion, especially my present and former colleagues in BioEng. My gratitude also goes to all my friends for sharing the happy moments, supporting me during my ups and downs, and for being who you are. Last but not least, thanks to my parents, for your love and encouragement. “Adat periuk berkerak, adat lesung berdedak” Dayang Norulfairuz Abang Zaidel Lyngby, January 2012 i Dayang N. Abang Zaidel 2012 ABSTRACT Sugar beet pulp is a byproduct from sugar production consisting mainly of cellulose and pectic polysaccharide. Its utilization has been mostly as feedstock due to its high content of energy and fiber. This study emphasizes on the utilization of the pectin and arabinan fractions extracted from sugar beet pulp as a potential starting material for production of pectin derived products which could help maintain the competitiveness of the sugar beet based industry. The overall objective of this study has been focusing on understanding the kinetics of enzyme catalyzed oxidative cross- linking of feruloylated polysaccharide from sugar beet and relating the kinetics of this cross- linking to the properties of the cross-linked products. Several hypotheses have been formulated in order to accomplish our objective. The first part of the study utilized arabinan-oligosaccharide fraction from sugar beet pulp byproduct. In this study we investigated the effect of arabinans backbone length on the kinetics of horseradish peroxidase (EC 1.11.1.7) (HRP) catalyzed oxidative cross-linking of ferulic acid (FA) moieties esterified to -(1,5)-linked arabinans; taking into account that FA can be oxidatively cross-linked by HRP catalysis in the presence of hydrogen peroxide (H2O2) to form ferulic acid dehydrodimers (diFAs). The composition of the substrate was analyzed by HPAEC, HPLC and MALDI-TOF, confirming the structural make up of the arabinan-oligosaccharide (Arabinose: 2.9- 3.4 mmolg-1 DM; FA: 2.5-7.0 mgg-1 DM) and verifying the formation of diFAs as a result of the enzyme catalyzed cross-linking reaction. The result demonstrates the influence of arabinans backbone length on the rates of FA cross-linking; longer arabinans exhibit a slower cross-linking rate than shorter, all other things being equal. It has been our intention to study the rheological properties of cross-linked feruloylated arabinan- oligosaccharide, however the attempt has not been fully achieved. It might be due to small molecular weight of the arabinan (1.3 kDa) which prevented the measurement of the rheological properties since the change in viscosity resulting from the cross-linking was insignificant. Therefore, the next part of the work presented in this thesis utilized sugar beet pectin (SBP) solid fraction extracted from sugar beet pulp which has molecular weight >100 kDa. The compositional analysis of the substrate shows abundant amount of FA (7.3 mgg-1 DM) in SBP which can be oxidatively cross-linked via enzyme catalyzed reaction by oxidoreductase enzymes. We hypothesized that different mechanisms of two oxidoreductase enzymes, i.e. HRP and laccase (EC 1.10.3.2), might influence the kinetics of the oxidative cross-linking and consequently the ii Enzyme catalyzed oxidative cross‐linking of feruloylated pectic polysaccharides from sugar beet – Kinetics and rheology properties of the gels formed. The kinetics of oxidative gelation of SBP, taking place via enzyme catalyzed cross-linking of FA, was evaluated by small angle oscillatory measurements. The result indicates a significant difference between the SBP gels produced from the catalysis of HRP and laccase, that is, laccase catalysis produced stronger SBP gels albeit slower rates of gelation than the HRP catalysis. Statistically design experiment has been constructed to investigate the effect of several reaction factors which might influence the rates of gelation of SBP catalyzed by HRP or laccase, particularly the pectin level, temperature, enzyme dosage, pH and, for HRP, the H2O2 concentration. The result reveals that these reaction factors could be tuned in order to adjust the enzyme catalyzed gelation and the properties of the gels produced. Moreover, positive correlation between the rates of gelation and gel strengths was obtained for laccase catalyzed gels, but no such correlation exists for HRP catalyzed gels. Chemical analysis confirmed the formation of diFAs in the cross-linked products by both enzymes catalysis supporting that the gelation was a result of oxidative cross-linking of FA. It is uncertain how the kinetics of enzyme catalyzed oxidative cross-linking of SBP and the gels properties are affected in emulsion systems. Thus, investigation on the enzyme catalyzed oxidative gelation of SBP was further performed on the SBP in emulsion systems. In this study, we have formulated two separate, identically composed, oil-in-water emulsion systems to study the effect of different methods of emulsion preparation on the emulsion stability in the presence of SBP and the kinetics of enzyme catalyzed oxidative gelation of SBP. The result shows that the different methods of emulsion preparation affect the emulsion stability and the rates of gelation of SBP in emulsion systems, and stronger gels were produced in the SBP containing emulsions as compared to the SBP without emulsions. From this study, we have shown that arabinan-oligosaccharide and SBP solid fractions extracted from sugar beet pulp byproduct could undergo oxidative cross-linking of the feruloyl group, which abundantly esterified to the arabinan side-chains, through enzyme catalyzed reaction. Our study provides the insight into the relationship between the kinetics of the oxidative cross-linking of FA with the structural characteristic of the oligosaccharide, and the correlation between the rates of enzyme catalyzed oxidative gelation of feruloylated polysaccharide and the rheological properties of the gels produced. This knowledge could be useful for designing application of sugar beet pectin in food technology or similar application. iii Dayang N. Abang Zaidel 2012 DANSK SAMMENFATNING Sukkerroepulp produceres som et biprodukt ved fremstilling af sukker fra sukkerroer og består hovedsagelig af cellulose og pektin, og indeholder herunder forskellige pektiske polysakkarider. Sukkerroepulp har hidtil hovedsagelig
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