Peroxidase-Mediated Oxidative Cross-Linking and Its Potential To Modify Mechanical Properties in Water-Soluble Polysaccharide Extracts and Cereal Grain Residues JAMES A. ROBERTSON,* CRAIG B. FAULDS,ANDREW C. SMITH, AND KEITH W. WALDRON Sustainability of the Food Chain Exploitation Platform, Institute of Food Research, Norwich Research Park, Colney, Norwich NR4 7UA, United Kingdom Analysis of wheat bran and spent grain shows that concentrations of ferulate and diferulates offer considerable scope to modify the cross-linking of feruloylated polysaccharides and hence the mechanical properties of these residues. In solution ferulic acid (FA) can be readily polymerized by horseradish peroxidase, but when esterified to a polysaccharide, the profile of diferulates becomes restricted. This situation also exists in muro and suggests structural constraints may limit the availability of FA for cross-linking. At relatively low polysaccharide concentration, (∼3%), steric restrictions were apparent in gels prepared using isolated polysaccharides. Mechanical properties such as swelling also appear to be fixed at these relatively low polysaccharide concentrations. This limits the potential to modify mechanical properties in muro through oxidoreductase activity. To modify mechanical properties such treatments will need to focus on increasing the “flexibility” of the cell wall matrix and the accessibility of enzymes to the cell wall matrix. KEYWORDS: Peroxidase; ferulate; cross-linking; swelling; wheat bran; spent grain INTRODUCTION Cross-linking of water-extractable polysaccharides through peroxidase treatment is a recognized method to develop products with designed functional properties (1–5). Chemical structure and polymer conformation and the propensity for molecular interaction are important considerations for designing functional properties (6). As well as isolated polysaccharides, such as pectins and arabinoxylans (AX), oxidative cross-linking is also applicable to proteins and protein-polysaccharide mixtures (7). This greatly adds to the diversity of potential gelling agents that might become available for biomedical use (8) and to control texture and stability properties in foods (6, 9, 10). Feruloylated polysaccharides have a recognized ability to be oxidatively cross-linked by oxidoreductases, such as peroxidases and laccases (1–4, 11). Although there has been much activity centered on the use of such polysaccharides when extracted from the cell wall, little information is available on the potential to modify cross-linking in muro and hence to manipulate the properties of cell wall-rich food processing residues, such as wheat bran (WB) and brewers’ spent grain (BSG). These cereal-derived residues have a large proportion of ferulate (FA) as well as diferulates (diFA) in the cell wall (12). There may be a structural or functional requirement to maintain FA and diFA in the cell wall, but equally the persistence of FA may indicate these moieties are unavailable for cross-linking. Unavailability may be related to an inaccessibility of enzyme to the cell wall matrix or the spatial distribution of FA and diferulates being sterically unfavorable for cross-linking. The chemical profiling within a semiflexible polysaccharide matrix of ferulates gives little information on their spatial location either within the cell wall or in different tissue types. It is known that different tissues within the cereal grain can have distinct profiles of phenolic acids (13, 14) and that a manganese-dependent peroxidase can induce modification to mechanical properties, but also this can be tissue specific (10). The ratio of FA to sugars is important for gelation, with an ara/xyl/FA (600:2000:1) ratio considered to be limiting for gelation in AX (6). It is presumed that the important ratio is the degree of substitution of arabinose, although other substitu- tions and esterification to a polysaccharide may also influence the steric accessibility of FA for cross-linking. For example, in ester substitution of pectic polysaccharides O-2 linked arabinose predominates (∼60% O-2-linked arabinose and ∼40% O-6- linked galactose), whereas in AX it is O-5-linked arabinose (15, 16). The overall ratio of FA to AX sugars in WB and BSG (12) was apparently in excess of the ratio reported to be required for oxidative cross-linked gelation (14). However, when ratios are considered, the total spectrum of AX present in each residue and relative distribution of FA and diFA has to be taken into * Corresponding author (telephone +44 1603 255000; fax +44 1603 507723; e-mail jim.robertson@bbsrc.ac.uk). 1720 J. Agric. Food Chem. 2008, 56, 1720–1726 10.1021/jf072445d CCC: $40.75  2008 American Chemical Society Published on Web 02/06/2008