University of Groningen Polymerization of hyperbrached polysaccharides by combined biocatalysis van der Vlist, Jeroen IMPORTANT NOTE: You are advised to consult the publisher's version (publisher's PDF) if you wish to cite from it. Please check the document version below. Document Version Publisher's PDF, also known as Version of record Publication date: 2011 Link to publication in University of Groningen/UMCG research database Citation for published version (APA): van der Vlist, J. (2011). Polymerization of hyperbrached polysaccharides by combined biocatalysis. s.n. Copyright Other than for strictly personal use, it is not permitted to download or to forward/distribute the text or part of it without the consent of the author(s) and/or copyright holder(s), unless the work is under an open content license (like Creative Commons). The publication may also be distributed here under the terms of Article 25fa of the Dutch Copyright Act, indicated by the “Taverne” license. 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Enschede,The Netherlands Cover photo: branched vein system of a leaf (Bert van 't Hul / stock.xchng) ISSN 1570‐1530 ISBN 978‐90‐367‐4729‐5 (print) ISBN 978‐90‐367‐4730‐1 (electronic) Zernike Institute PhD thesis series 2011‐05 RIJKSUNIVERSITEIT GRONINGEN Polymerization of hyperbranched polysaccharides by combined biocatalysis Proefschrift ter verkrijging van het doctoraat in de Wiskunde en Natuurwetenschappen aan de Rijksuniversiteit Groningen op gezag van de Rector Magnificus, dr. F. Zwarts, in het openbaar te verdedigen op vrijdag 28 januari 2011 om 16.15 uur door Jeroen van der Vlist geboren op 25 oktober 1978 te Gouda Promotor : Prof. dr. K. Loos Beoordelingscommissie : Prof. dr. L. Dijkhuizen Prof. dr. P. Mischnick Prof. dr. A. Gandini TABLE OF CONTENTS CHAPTER 1 7 Introduction CHAPTER 2 35 Synthesis of hyperbranched polysaccharides CHAPTER 3 65 Hyperbranched polyglucan brushes CHAPTER 4 87 Hyperbranched polysaccharide sugar balls CHAPTER 5 111 Hyperbranched polyglucan diblock copolymers SUMMARY 126 SAMENVATTING 129 DANKWOORD 133 CHAPTER 1 General introduction CHAPTER 1 1.1 CARBOHYDRATE CHEMISTRY 1.1.1 Glucose based carbohydrates Carbohydrates or saccharides are one of the four major classes of biomolecules. Proteins, nucleic acids and lipids are the other three. Carbohydrates consist of an aldehyde group (aldoses) or ketone group (ketoses) with multiple hydroxy groups. The simplest carbohydrates are the monosaccharides that exist, due to a stereo center, in two configurations (D and L). Aldoses with six carbon atoms (hexoses) have even four stereo centers resulting in 16 stereoisomers, D‐glucose being one of them. FIGURE 1.1: Fischer projections of 8 of the 16 hexoses (only the D stereoisomers are shown). Although a linear structure of glucose occurs, the ring structure is the predominant configuration in solution (>99%). The aldehyde group can react intramolecular with the C5 hydroxy group to form a pyranose ring. An additional stereo center is created at the C1 atom with this cyclization and as a result, there are two different ring structures. In the configuration, the hydroxyl group attached to C1 is axial positioned while in the configuration the hydroxyl group is equatorially positioned. The configuration is energetically favoured and the ratio of : anomers is 36:63 in solution1. 8 GENERAL INTRODUCTION FIGURE 1.2: The open‐chain form of D‐glucose and the cyclic and configurations. The carbon atoms are numbered from 1 to 6, starting from the aldehyde group. 1.1.2 Polysaccharides Polysaccharides are monosaccharides linked together by glycosidic bonds. The notation used to distinguish different glycosidic bonds consists of the notation or (the 2 possible anomers of cyclic glucose) followed by the carbon numbers which actually join the monosaccharides between brackets. For example, the linkage that connects the two D‐glucose residues of the disaccharide maltose, which are joined at the C1 and C4 position, is written as: (1→4). Besides the multiple linking positions and linking types, monosaccharides can have different ring sizes, various stereoisomers and can carry different substituents. Hence, it is not difficult to imagine that there is a huge variety of polysaccharides. Some abundant homopolysaccharides constructed from D‐glucose are summarized below. Dextran consists of (1→6) linked D‐glucose residues and is the energy storage polysaccharide of yeasts and bacteria. Branching can occur at (1→2), (1→3) and (1→4) depending on the source2. Dextran is highly soluble in water, lacks nonspecific cell binding and resists protein adsorption, which makes dextran an interesting biomaterial for implantable purposes3. 9 CHAPTER 1 FIGURE 1.3: Linear chain of dextran, (1→6) linkage. Amylose is an (1→4) linked glucan and is the linear component of starch. Although the composition of starch from each plant is unique, most starches contain 20 to 25 % amylose. The degree of polymerization of amylose varies also with the origin. Amylose from potato or tapioca starch has a DP of 1000 to 6000 while amylose from maize or wheat amylose has a degree of polymerization varying between 200 and 12004. Due to intramolecular hydrogen bonding of the hydroxyl groups, amylose tends to wind up in a left‐handed double stranded helix5. This helical conformation, with a relatively hydrophobic inner part, can be filled with water molecules but also with more hydrophobic compounds, such as fatty acids6. FIGURE 1.4: Structure of amylose chain, (1→4) linkage. Cellulose is one of the most abundant biopolymers in the biosphere7. Almost half of the cell wall material of wood constitutes of cellulose but it is also produced by algae, bacteria and prokaryotes8. Cellulose is built up from (1→4) linked D‐glucose residues and is the isomer of amylose. In nature, most cellulose is synthesized as crystalline microfibrils. Within these microfibrils the cellulose chains are parallel aligned and forms intermolecular hydrogen bonding between neighbouring chains. The degree of polymerization varies by origin. Cotton and other plant fibers have for example DP values in the 800 – 10000 range while wood cellulose has a DP in the range of 300 to 17009. 10 GENERAL INTRODUCTION FIGURE 1.5: Structure of cellulose, (1→4) linkage. Chitin like cellulose is a (1→4) linked polyglucan but has an acetamido attached to C2 instead of a hydroxy group. Exoskeletons of insects are constructed from chitin and it is present in the cell walls of most fungi and many algae. Chitin is the second most abundant polysaccharide. Chitin itself is a hydrophobic polymer and insoluble in aqueous solutions at neutral pH. However, the (often incomplete) N‐deactetylation of chitin increases the water solubility and provides primary amines for further chemical modification10. The (partly) N‐deacetylated analogue of chitin is known as chitosan and has been investigated for different biomedical applications11. FIGURE 1.6: Structure of chitin, (1→4) linkage. Amylopectin and glycogen are branched (1→4) linked D‐glucose polysaccharides with a DP in the range of 60 000 and 6 000 00012,13. Branching occurs at the (1→6) position. Amylopectin is the branched energy storage polymer of plants while glycogen is the energy storage polymer of mammals. Glycogen has typically branch lengths of 10 D‐glucose units while amylopectin has branch lengths between 24 and 30 D‐glucose residues. Another major difference is the branching pattern. While the branching pattern of glycogen is random, the branching points of amylopectin are clustered in regions14. A stable opalescent solution is obtained when glycogen or amylopectin are dissolved in water. 11 CHAPTER 1 FIGURE 1.7: Part of the branched structure of glycogen. Amylopectin is branched in a similar way to glycogen but with the difference that the branch points are clustered. 1.1.3 Polysaccharides in industry Polysaccharides are an abundant source of raw materials that are interesting due to their biodegradable, biocompatible and renewable character. Saccharides are expected to play an increasingly relevant role as raw material in the future and, in particular, as a potential candidate to replace petrol‐based materials. Already, polysaccharides find their way in many different fields of industry. A short overview is given in TABLE 1.1. TABLE 1.1: Polysaccharide processing industries and used polysaccharides. Industry Polysaccharide Main function Paper Cellulose, starch Structural material, coating Food Starch Rheological control, texturizer Biomedical Dextran, chitosan, hyaluronan Drug carrier, wound dressing Package Starch/cellulose derivatives Reduction of synthetic polymers Coating Starch Rheological control Adhesive Starch Tackifier Textile Cellulose (cotton) Woven fabric