Process Biochemistry 50 (2015) 2168–2176 Contents lists available at ScienceDirect Process Biochemistry jo u r n al homep age: www.elsevier.com/locate/procbio Synthesis of large-ring cyclodextrin from tapioca starch by amylomaltase and complex formation with vitamin E acetate for solubility enhancement K. Kuttiyawong a , S. Saehu b , K. Ito c , P. Pongsawasdi b,∗ a Faculty of Liberal Arts and Science, Kasetsart University KamphaengSaen Campus, Nakhon Pathom 73140, Thailand b Starch and Cyclodextrin Research Unit, Department of Biochemistry, Faculty of Science, Chulalongkorn University, Bangkok 10330, Thailand c Lab Enzyme Chemistry, Graduate School of Science, Osaka City University, Osaka 558-8585, Japan a r t i c l e i n f o Article history: Received 21 April 2015 Received in revised form 6 October 2015 Accepted 21 October 2015 Available online 28 October 2015 Keywords: Large-ring cyclodextrin Amylomaltase Tapioca starch Vitamin E acetate Solubility enhancement a b s t r a c t Recombinant amylomaltase from Corynebacterium glutamicum (CgAM) was used to synthesize large-ring cyclodextrins (LR-CDs) from tapioca starch. A mixture of LR-CDs (CD22–CD54) was obtained from both raw and soluble tapioca starch, but with different ratio of main products (CD25–CD30). Almost three- fold higher amount of products was formed indicating the preference for raw tapioca starch. The optimal condition was to incubate 1.0% (w/v) starch with 0.05 U/ml CgAM in buffer pH 6.0 at 30 ◦ C for 18 h. Total LR-CDs yield was 60.5 mg/g starch. LR-CDs were resistant to hydrolysis by Rhizopus sp. glucoamylase but completely hydrolyzed by CgAM at high enzyme concentration. Solid inclusion complex between LR-CDs mixture and vitamin E acetate was formed, as analyzed by FTIR. For complex preparation, the 1:10 mol ratio of LR-CDs to vitamin E was chosen because maximum amount of the vitamin was bound to LR-CDs and high complex yield was obtained. Aqueous solubility of vitamin E acetate in complex form increased 800-fold while the antioxidant activity decreased about 30%. This is the first work to demonstrate the use of tapioca starch in LR-CDs synthesis by CgAM and the complex formation between LR-CDs and vitamin E acetate as solubility enhancement process of vitamin. © 2015 Elsevier Ltd. All rights reserved. 1. Introduction Amylomaltase (EC 2.4.1.25, AM), a member of 4- glucanotransferase (4-GTase) group in the -amylase family, catalyzes the intramolecular and intermolecular transglucosyla- tion of starch substrate to yield large-ring cyclodextrin (LR–CD) and linear maltooligosaccharides, respectively. AM displays four dif- ferent types of reactions: cyclization, disproportionation, coupling and hydrolysis. It is classified in the glycoside hydrolase GH77, an efficient 4-GTases with remarkably low hydrolytic activity compared to GH13 enzymes [1]. AM is an intracellular enzyme, first found in Escherichia coli as a maltose-inducible enzyme that is essential for metabolism of maltose [2]. Several AMs from hyperthermophilic bacteria i.e., Thermus aquaticus [3], Thermus thermophilus [4], Thermus brockianus [5], Aquifex aeolicus [6] and archaea i.e., Pyrobaculum aerophilum [7], Thermococcus litoralis [8] have been reported. A novel AM from the mesophilic bacterium ∗ Corresponding author. Fax: +66 2218 5418. E-mail address: piamsook.p@chula.ac.th (P. Pongsawasdi). Corynebacterium glutamicum (CgAM) has recently been reported and characterized [9,10]. This AM has only about 30% amino acid sequence similarity and shows different LR-CD production profile when compared with the well characterized AM from T. aquaticus [9]. A few applications of AM have been reported. Firstly, they are used for the production of LR-CDs with a degree of polymeriza- tion of 17 onwards from starch: potato and pea starch have been reported [1]. Secondly, they are used in the synthesis of functional oligosaccharides, such as a prebiotic isomaltooligosaccharide (IMO) as neutraceuticals with health benefits [11]. The use of AM for the synthesis of an anticariogenic maltooligosylsucrose has also been reported [12]. Recently, AM has been used in the production of a thermoreversible starch gel to be used as a substitute for gelatin or agar in food products [13]. LR-CDs or cycloamyloses produced from amylomaltase are in the range of CD17–CD60 [1]. However, up to CD31 have been characterized for their structures and physicochemical properties [14]. LR-CDs are highly soluble in water, assumed to form a single helicle V-amylose conformation and a toroidal shape, with an anhy- drophilic channel-like cavity [15]. They are able to form complexes http://dx.doi.org/10.1016/j.procbio.2015.10.014 1359-5113/© 2015 Elsevier Ltd. All rights reserved.