Malto-oligosaccharide alcohols (MOSA) are one of the most important sugar alcohols widely used as sweetener in food, cosmetic, and pharmaceutical industries in recent years, of which maltotriitol-rich MOSA is much more recognized. With the aim of preparing maltotriitol-rich MOSA from starch, a novel process was developed and optimized. Starch was first liquefied with thermostable Bacillus licheniformis alpha-amylase. The liquefied starch was then saccharified to yield maltotriose-rich malto-oligosaccharides under the cooperative actions of Bacillus naganoensis pullulanase, Bacillus amyloliquefaciens alpha-amylase, and barley bran beta-amylase. The maltotriitol-rich MOSA was finally prepared by chemical hydrogenation. Under the optimized conditions, maltotriitol-rich MOSA containing 42.18% maltotriitol was obtained with a conversion rate of 104.57% from starch. The process can be employed for large-scale preparation of maltotriitol-rich MOSA, and a further modification of the process can lead to the formulation of various types of MOSA with different percentages of components of sugar alcohols. (C) 2016 Elsevier Ltd. All rights reserved.