Enzymatic Solubilization of Brewers’ Spent Grain by Combined Action of Carbohydrases and Peptidases JANNEKE TREIMO,* ,† BJORGE WESTERENG, † SVEIN J. HORN, † PIRKKO FORSSELL, § JAMES A. ROBERTSON, # CRAIG B. FAULDS, # KEITH W. WALDRON, # JOHANNA BUCHERT, § AND VINCENT G. H. EIJSINK † Department of Chemistry, Biotechnology and Food Science, Norwegian University of Life Sciences, N-1432 Aas, Norway; VTT Biotechnology, Espoo, Finland; and Sustainability of the Food Chain Exploitation Platform, Institute of Food Research, Norwich, United Kingdom Brewers’ spent grain (BSG), a high-volume coproduct from the brewing industry, primarily contains proteins, barley cell wall carbohydrates, and lignin. To create new possibilities for the exploitation of this large biomass stream, the solubilization of BSG by the combined action of carbohydrases (Depol 740 and Econase) and peptidase (Alcalase and Promod 439) was explored. Hydrolysis protocols were optimized with respect to temperature (influencing both microbial contamination and rate of enzymatic hydrolysis), pH, enzyme dose, order of enzyme addition, and processing time. On the basis of this approach, one- and two-step protocols are proposed taking 4-8 h and yielding combined or separate fractions of hydrolyzed oligosaccharides and liberated hydrolyzed protein. Optimized procedures resulted in the solubilization of >80% of the proteinaceous material, up to 39% of the total carbohydrates, and up to 42% of total dry matter in BSG. Of the original xylan present in BSG, 36% could be solubilized. Sequential and simultaneous treatments with the two enzyme types gave similar results. In sequential processes, the order of the carbohydrase and peptidase treatments had only minor effects on the outcome. Depol 740 released more pentoses than Econase and gave slightly higher overall dry matter solubilization yields. KEYWORDS: Brewers’ spent grain; enzymatic solubilization; hydrolysis; carbohydrase; peptidase; Depol 740; Econase; Alcalase INTRODUCTION Barley (Hordeum Vulgare L.) is among the major cereal crops in the world. The primary use of barley is in the production of animal feed, whereas smaller quantities are used for food and beer production (1-3). An early step in the production of beer involves the germination of barley to synthesize and activate hydrolytic enzymes, a process called malting. The enzymes hydrolyze primarily the barley starch (but also other compo- nents) during the mashing process, and the resulting sugar-rich liquor (wort) is fermented to beer. This process yields an abundant coproduct called brewers’ spent grain (BSG), which consists of the barley malt residue after separation of the wort. About 200 g of wet BSG is produced per liter of beer [reviewed by Mussatto et al. (2)]. BSG is a lignocellulosic material that consists of hemicellulose (∼28%, mainly arabinoxylans), cel- lulose (∼17%), lignin (∼28%), and protein (∼20%) (2, 3), as well as a few percent of residual starch and starch-derived products (glucose, maltosaccharides). Traditionally, BSG has been utilized directly as animal fodder (2). Because large amounts of BSG are produced continuously worldwide, other exploitation routes are of current interest (2). The recovery of carbohydrates and/or proteins from BSG is potentially attractive because this has the potential create new, higher value applications. Several promising compounds have been extracted from BSG (2, 4), such as arabinoxylan or corresponding oligosaccharides (5, 6), the phenolic compounds ferulic and p-coumaric acid (7, 8), and lignin (9). Novel potential products may include protein-enriched material, prebiotics (10, 11), and hydrolysates to be used, for example, as growth media for (probiotic) bacteria (12-17) or for the production of xylitol (18). Use of enzymes in the barley grain mashing process step or for solubilization of BSG has previously been investigated to some extent, with the main focus on the use of carbohy- drases (6, 8, 19-31). We (17) and others (27) have recently studied how peptidases may be used to hydrolyze and release most of the protein in BSG. Despite these previous studies, there still is limited knowledge concerning the potential of modern commercial enzyme technology to transform BSG within an industrially acceptable processing time frame. For example, integrated processes, combining several types of enzymatic * Corresponding author (telephone +47 64 96 59 00; fax +47 64 96 59 01; e-mail janneke.treimo@umb.no). † Norwegian University of Life Sciences. § VTT Biotechnology. # Institute of Food Research. 3316 J. Agric. Food Chem. 2009, 57, 3316–3324 10.1021/jf803310f CCC: $40.75  2009 American Chemical Society Published on Web 03/13/2009 Downloaded by CSIC CTRO INV BIOLOGICAS on September 30, 2009 | http://pubs.acs.org Publication Date (Web): March 13, 2009 | doi: 10.1021/jf803310f