Synthesis of fructooligosaccharides and oligolevans by the combined use of levansucrase and endo-inulinase in one-step bi-enzymatic system Feng Tian, Salwa Karboune ⁎, Andrea Hill Department of Food Science and Agricultural Chemistry, McGill University, 21,111 Lakeshore, Ste-Anne de Bellevue, Québec H9X 3V9, Canada abstract article info Article history: Received 29 April 2013 Accepted 4 December 2013 Available online 14 December 2013 Editor Proof Receive Date 6 January 2014 Keywords: Levansucrase Endo-inulinase Fructooligosaccharides Oligolevans Levan Bacillus amyloliquefaciens Levansucrase from Bacillus amyloliquefaciens and endo-inulinase from Aspergillus niger were used in a one-step reaction to produce short chain fructooligosaccharides (scFOSs) and oligolevans from sucrose. Levansucrase catalyzed the synthesis of levan, while endo-inulinase regulated the product molecular size. The bi-enzymatic system showed higher yield and productivity (67% w/w; 96 g/L/h) than the levansucrase enzymatic system (3.0% w/w; 0.8 g/L/h) alone. The contribution of endo-inulinase to the formation of scFOSs and oligolevans through its hydrolytic activity was higher than that of levansucrase through its acceptor reaction; however, the production of intermediate levans with appropriate MW by levansucrase was prerequisite. The maximal concen- tration of scFOSs was higher as compared to that of oligolevans. Among scFOSs, 6-kestose was the most dominant product. The use of immobilized levansucrase resulted in a lower production of scFOSs and higher yield of levan. The current study is the first to highlight the potential of levansucrase/endo-inulinase bi-enzymatic system. Industrial relevance: Functional food products are receiving a substantial amount of interest from consumers. Considering the size of this market, the food industry sector could benefit considerably from improvements in the functional ingredients used to support the development of these health promoting food products. In this con- text, fructooligosaccharides (FOSs), a class of ingredients whose potential health benefits in terms of supporting intestinal health and reducing the risk of cancers are increasingly being recognized. However, commercially available FOSs are exclusively β-(2→1)-inulin-type prebiotics with short chains, which are mainly absorbed in the small intestine. β-(2→6) and neolevan-type-FOSs have shown prebiotic activities that surpass the current β-(2→1)-FOSs generation. The development of innovative bi-enzymatic process for the production of FOSs and β-(2→6)-oligolevans in the present study is of high interest. The developed bienzymatic system showed high yield and productivity for the production of FOSs and oligolevans from sucrose as abundant starting materials. These products being of high- degree of polymerization are expected to exhibit an increased colonic prebiotic persistence and reach the distal intestinal region where most of the chronic diseases are originated. © 2014 Elsevier Ltd. All rights reserved. 1. Introduction Fructooligosaccharides (FOSs) have attracted greater attention due to the increased demand for functional ingredients. In addition to their low caloric value and non-cariogenic properties, FOSs promote the intestinal health by stimulating the growth/activity of beneficial lactic acid bacteria and enhance the immune response (Fanaro et al., 2005). Commercially available FOSs for human consumption are exclusively β-(2→1)-inulin-type prebiotics that are mainly produced from sucrose by the action of fructofuranosidases (Mussatto, Rodrigues, & Teixeira, 2009). The β-(2→6) and neolevan-type-FOSs have shown prebiotic activities that surpass the current FOS genera- tion (Kilian, Kritzinger, Rycroft, Gibson, & du Preez, 2002). In addition, β-(2→6)-levan-type-hetero-FOSs may have new anti-adhesive activity against pathogens and immunomodulatory capacity (Shibata et al., 2009). Levansucrases have been investigated by our group and others as potential biocatalysts for the synthesis of β-(2→6)-levan-type-FOSs (van Hijum, Szalowska, van der Maarel, & Dijkhuizen, 2004; Homann, Biedendieck, Gotze, Jahn, & Seibel, 2007; Beine et al., 2008; Tian & Karboune, 2012). Levansucrase (EC 2.4.1.10), which belongs to the glycoside hydro- lase family 68 (GH68), catalyzes the synthesis of FOSs and high- molecular weight β-(2→6)-levan from sucrose (van Hijum, Bonting, van der Maarel, & Dijkhuizen, 2001; Meng & Fütterer, 2003). Levansucrases from Gram-negative bacteria Gluconacetobacter diazotrophicus (Hernandez et al., 1995) and Zymomonas mobilis (Crittenden & Doelle, 1993) synthesize mostly FOSs and low amounts of levan; while levansucrases from Gram-positive bacteria, Bacillus subtilis (Euzenat, Guibert, & Combes, 1997), Bacillus megaterium (Homann et al., 2007), Lactobacillus reuteri 121 (Ozimek, Kralj, Van der Innovative Food Science and Emerging Technologies 22 (2014) 230–238 ⁎ Corresponding author. Tel.: +1 514 398 8666; fax: +1 514 398 7977. E-mail address: salwa.karboune@mcgill.ca (S. Karboune). 1466-8564/$ – see front matter © 2014 Elsevier Ltd. All rights reserved. http://dx.doi.org/10.1016/j.ifset.2013.12.004 Contents lists available at ScienceDirect Innovative Food Science and Emerging Technologies journal homepage: www.elsevier.com/locate/ifset