Available online at www.sciencedirect.com Journal of Molecular Catalysis B: Enzymatic 51 (2008) 110–117 Lecitase ® ultra as regioselective biocatalyst in the hydrolysis of fully protected carbohydrates Strong modulation by using different immobilization protocols Gloria Fernandez-Lorente a , Marco Filice b , Marco Terreni b , Jose M. Guisan a , Roberto Fernandez-Lafuente a,∗ , Jose M. Palomo a,∗ a Departamento de Biocat ´ alisis, Instituto de Cat ´ alisis (CSIC), Campus UAM, Cantoblanco, 28049 Madrid, Spain b Dipartimento di Chimica Farmaceutica, Universit` a di Pavia, via Taramelli 12, 27100 Pavia, Italy Received 6 August 2007; received in revised form 14 November 2007; accepted 20 November 2007 Available online 26 November 2007 Abstract This paper shows that Lecitase Ultra is an enzyme preparation with a great interest as regioselective biocatalyst in the deprotection of 4 different peracetylated sugars: 1,2,3,4,6-penta-O-acetyl--d-galactopyranose (1), 2-acetamido-2-deoxy-1,3,4,6-tetra-O-acetyl--d-glucopyranose (4), 1,2,3,4,6-penta-O-acetyl--d-mannopyranose (7) and 2,3,4,6-tetra-O-acetyl--d-galacto pyranosyl-(1 → 4)-1,2,3,6-tetra-O-acetyl--d- glucopyranoside (9). The enzyme properties (specificity, preference for the per-acetylated sugar and regio-selectivity) were strongly modulated by the immobilization conditions, for example the octyl-LECI preparation was 10 fold more active than the PEI-LECI preparation, while it was more than 40 fold less active against some other substrates. Very interestingly, these changes also affected the regioselectivity, depending on the preparation used it was possible to get free OH groups in anomeric position, position 6 or the mixture of both. Finally, the octyl-LECI preparation did not recognize the -sugars, favouring the -isomers (in opposition to most commercial lipases or the other LECI preparations). This is potentially useful to obtain pure -peracetylated monosaccharides from a mixture of anomers. © 2007 Elsevier B.V. All rights reserved. Keywords: Regioselective reaction; Enzyme modulation; Enzyme specificity; Phospholipase; Peracetylated carbohydrates 1. Introduction Phospholipases A 1 are of particular interest for industrial applications, to produce 2-acyl-lysophospholipids (good emul- sifiers) with interesting fatty acid composition (eicosapentaenoic acid, conjugated linoleic acid and docosahexaenoic acid) and degumming process of oils [1]. The interest on finding new phospholipases is shown by the high number of sources for this enzyme discovered in the recent years and the trials to improve their performance by diverse techniques [2]. Recently, a new enzyme preparation with phospholipase A 1 activity, namely Lecitase ® Ultra (LECI), was patented [3] and made commer- cially available. Following the supplier information, this is a preparation obtained from the fusion of the genes of the lipase ∗ Corresponding authors. Tel.: +34 91 585 48 09; fax: +34 91 585 47 60. E-mail addresses: rfl@icp.csic.es (R. Fernandez-Lafuente), josempalomo@icp.csic.es (J.M. Palomo). from Thermomyces lanuginosa and the phospholipase from Fusarium oxysporum developed mainly for degumming pro- cesses [3]. This new enzyme presented the stability of the lipase from T. lanuginosa and the activity of the enzyme from F. oxys- porum [3]. However, it also has biocatalytic properties that are useful in the synthesis of tailored phospholipids, although due to the relative novelty of this product only few manuscripts may be found in literature on its use [4]. Phospholipases have been described to suffer interfacial acti- vation [5], similarly to lipases [6]. Lipases present a broad substrate specificity combined with a high regio and enantios- electivity, and may be used in many different reaction media [7]. However, to our knowledge, LECI as biocatalyst for fine chemistry – using substrates far from phospholipids – has been only recently utilized to resolve racemic mixtures of chiral com- pounds [8]. Here, we present a first study on the use of this new com- mercial enzyme in the regioselective hydrolysis of peracetylated carbohydrates. Pure regioisomers of O-acetyl-glycopyranosides 1381-1177/$ – see front matter © 2007 Elsevier B.V. All rights reserved. doi:10.1016/j.molcatb.2007.11.017