Xylitol fatty acid esters of increasing chain length were synthesised from xylitol and free fatty acids in binary mixtures of tert-butanol (t-BuOH) and pyridine using Candida antarctica lipase B as biocatalyst. NMR and GC analyses revealed that monoesters of all acyl donors were primarily substituted at the C-1 position (93-97%, mol/mol) and to a minor extent at the C-2 position introducing asymmetry to the structure; while acylation at C-3 was not found. Diesters were esterified mainly at the C-1 and C-5 positions (87-100%, mol/mol) and to a minor extent 1,2-di-O-acyl- and 1,4-di-O-acyl xylitol esters were identified. Acylation of the secondary alcohol groups was interpreted as a result of acyl migration, because it occurred at a later stage of the process at a significantly lower rate than acylation of the primary alcohol groups and because it decreased with increasing chain length of the acyl donor. For acyl donors of chain lengths higher than four, the degree of esterification was controlled by the pyridine content of the reaction medium with the highest yield of monoesters obtained in 55% (v/v) pyridine. With butanoic acid the highest yield of monoester was obtained in pure t-BuOH. Under these conditions a degree of conversion of 26-41% of xylitol - and a yield of 22-29% monoester was obtained already after 4.5 It of incubation at 45 degrees C. The initial reaction rate for monoester synthesis was 5-19 times higher than for diester synthesis correlating inversely with the chain length of the acyl donor. At an acyl acceptor to donor ratio of 1:5 (mol/mol) the highest initial reaction rate of monoester synthesis was observed with hexanoic acid. (c) 2007 Elsevier Inc. All rights reserved.