Vol. 87, No. 4, 2010 283 Wheat Bran AX Properties and Choice of Xylanase Affect Enzymic Production of Wheat Bran-Derived Arabinoxylan-Oligosaccharides Valerie Van Craeyveld, 1 Emmie Dornez, 1 Ulla Holopainen, 2 Emilia Selinheimo, 2 Kaisa Poutanen, 2 Jan A. Delcour, 1 and Christophe M. Courtin 1,3 ABSTRACT Cereal Chem. 87(4):283–291 Wheat bran-derived arabinoxylan-oligosaccharides (AXOS) recently have been shown to potentially exert prebiotic effects. In this study, 15 bran samples obtained by milling different wheat cultivars were treated with xylanases from Hypocrea jecorina (XHJ), Aspergillus aculeatus (XAA), and Pseudoalteromonas haloplanktis (XPH) to assess the effect of bran source and xylanase properties on the AXOS yield and structure. The total arabinoxylan (AX) extraction yield was higher with XHJ (8.2– 10.7%) and XAA (8.2–10.8%) than with XPH (6.9–9.5%). Irrespective of the enzyme, a significant negative correlation was observed between ex- traction yield and arabinose to xylose (A/X) ratio of bran AX (r = –0.7), but not between yield and bran AX level. The A/X ratio of the extracted material was 0.27–0.34 for all bran samples and all enzymes, which com- bined with yield data and microscopic analysis, indicated primary hy- drolysis of aleurone and nucellar epidermis AX. The average degree of polymerization (avDP) of the extracted AX was very low for all enzymes (2–3), owing to the release of high levels of monomeric arabinose and xylose. The release of these monosaccharides could be ascribed to 1) the activity of wheat bran-associated enzymes (arabinofuranosidases and xylosidases); 2) the hydrolytic properties of the xylanases themselves; and 3) the presence of xylosidases as contaminations in enzyme prepa- ration, in that order of importance. Heat treatment of bran before xylanase treatment significantly decreased the levels of monomeric arabinose and xylose in the extract, without affecting the extraction yield, resulting in a higher avDP of 3–7, thus yielding true AXOS. Overall, for AXOS pro- duction, wheat cultivars with a low bran A/X ratio of the AX are prefer- able as starting materials, and inactivation of bran-associated enzymes before incubation is desirable. The XHJ xylanase was the best enzyme for wheat bran-derived AXOS production. Millers’ wheat bran is a multilayered composite comprising a range of tissues including, from outside to inside, the pericarp (epidermis, hypodermis, cross, and tube cells), the seed coat, the nucellar epidermis, the aleurone layer, and some starchy endo- sperm. It is a largely available by-product of the wheat milling process and it contains a high level of arabinoxylans (AX) (16– 25%) (Bataillon et al 1998; Maes and Delcour 2001; Beaugrand et al 2004b; Swennen et al 2006). Wheat bran AX are polysaccha- rides, built up of a backbone of β-(1→4)-linked D-xylopyranosyl residues (xylose), with some L-arabinofuranosyl residues (arabi- nose) on the C(O)-2 or C(O)-3 positions of xylose and some uronic acids, mostly glucuronic acid, on the C(O)-2 position of xylose (Izydorczyk and Biliaderis 1995; Schooneveld-Bergmans et al 1999). However, the fine structure of the AX varies largely within different tissues. AX originating from the aleurone layer and the nucellar epidermis are characterized by low arabinose to xylose (A/X) ratios (0.3–0.5 and 0.1, respectively), while the A/X ratio of outer pericarp (epidermis and hypodermis) AX is ≥1.0 (Antoine et al 2003; Barron et al 2007). The outer pericarp con- tains 1.7% glucuronic acid, whereas no uronic acid residues were detected in the aleurone layer (Benamrouche et al 2002). Wheat bran-derived arabinoxylan-oligosaccharides (AXOS), pro- duced by enzymic or chemical cleavage of AX have selectively stimulated the growth and activity of beneficial colon bacteria, and are therefore considered as potential prebiotics. Several studies reported on the in vitro bifidogenic effects for this type of compo- nent (Yamada et al 1993; Van Laere et al 2000; Grootaert et al 2007) and although bifidobacteria do not produce xylanases to degrade AXOS, several Bifidobacterium species have produced arabinofuranosidases and xylosidases (Zeng et al 2007). Also, they can cooperate with xylanase-producing bacteria (Grootaert et al 2007). A wheat bran-derived AXOS preparation could be fermented completely by Bifidobacterium adolescentis and B. longum, whereas several Bacteroides and Clostridium species showed no degrada- tion of the substrate (Yamada et al 1993; Van Laere et al 2000). The growth of B. bifidum is further stimulated by the presence of ferulic acid residues on AXOS (Yuan et al 2005). In an in vitro, continuous fermentation system mimicking the human gastrointestinal tract, partially hydrolyzed wheat flour AX increased the bifidobacteria concentration and decreased the con- centrations of bacteroides and clostridia (Vardakou et al 2007). In a similar system, an AXOS preparation with an avDP of 29 only slightly modified the total microbial community (Sanchez et al 2009) but significantly increased the amount of health-promoting lactobacilli as well as of Bacteroides–Prevotella and Clostridium coccoides–Eubacterium rectale groups. It also decreased levels of the toxic proteolytic markers phenol and p-cresol in the two distal colon compartments and increased concentrations of beneficial short-chain fatty acids (SCFA) in all colon vessels (Sanchez et al 2009). A similar experiment with AXOS components with a smaller avDP of 15 suggested that AXOS primarily affects microbial me- tabolism by switching on AXOS-degrading enzymes (xylanase, arabinofuranosidase and xylosidase) without significantly affect- * The e -Xtra logo stands for “electronic extra” and indicates that Fig. 2 appears in color online. 1 Laboratory of Food Chemistry and Biochemistry and Leuven Food Science and Nutrition Research Centre (LFoRCe), Katholieke Universiteit Leuven, Kasteel- park Arenberg 20, 3001 Leuven, Belgium. 2 VTT Technical Research Centre of Finland, P.O. Box 1000, FI-02044 VTT, Finland. 3 Corresponding author. Phone: + 32 16 321917. Fax: + 32 16 321997. E-mail: christophe.courtin@biw.kuleuven.be doi:10.1094/ CCHEM-87-4-0283 © 2010 AACC International, Inc. e - Xt ra * SPECIAL SECTION: Molecular Diversity and Health Benefits of Carbohydrates from Cereals and Pulses