Industrial bioprocesses and biocatalysis Lectures L1.1 Enzymatic acylation of starch

Starch is an interesting, natural, abundant and renewable raw material. For many applications, its properties can be improved by chemical modification, and acylation has been commonly used. Traditionally this makes use of powerful acylating agents (anhydrides and acid chlorides) and/or harsh conditions. Enzymatic acylation would be attractive because of the use of mild conditions, specific reaction, and the avoidance of by-products, particularly if fatty acids can be used as acyl donors. It is difficult to find a reaction system in which the three species can be brought into mutual contact: the starch; an enzyme; and water insoluble fatty acids or derivatives. However, there are some literature reports of successful acylation in both organic and aqueous gel media. This talk will present our recent studies of acylation with medium chain fatty acids, mainly with the liquid acid dispersed as droplets in an aqueous starch gel containing a lipase (1). It is possible to obtain substitution up to about 1 acyl chain per 50 glucose residues, which is the level desired for most intended applications. Acylation does not proceed further, but it is not clear whether this reflects reaction equilibrium or kinetic issues. There is a sharp optimum in water content at just under 2 g per g starch – below this the starch does not gelatinize properly, while above this the excess water may oppose the desired reverse hydrolysis reaction. Product analysis is an important issue in these systems, with several commonly used methods not reliable for the type of products obtained by enzymatic acylation. Alkaline methanolysis followed by GC seems to be a sound approach [1]. Bifunctional fatty acids may usefully be esterified to the starch in this system. Alkynoic acids with a terminal triple bond provide a site at which various azidecontaining reagents can be attached under mild conditions using click chemistry [2]. This was exemplified by attaching a fluorescent reporter, and also biotin, giving a starch preparation able to bind the protein streptavidin. References 1. Alissandratos A, Baudendistel N, Flitsch SL, Hauer B, Halling PJ (2010) Lipase-catalysed acylation of starch and determination of the degree of substitution by methanolysis and GC. BMC Biotechnology 10: 82. 2. Alissandratos A, Baudendistel N, Hauer B, Baldenius K, Flitsch SL, Halling PJ (2011). Biocompatible functionalisation of starch. Chemical Communications 47: 683-685. L1.2