X. LI (1), J. Sun (1), T. Zhou (1), Z. Jin (1), F. Gao (1), G. Cai (1), J. Lu (1);
(1) National Engineering Laboratory for Cereal Fermentation Technology, School of Biotechnology, Jiangnan University, Wuxi, China
The filterability defect of domestic barley malt is a long-term disturbing problem in the Chinese brewing industry. In contrast, beer manufacture using Canadian and Australian barley malts seldom encounters this problem. The direct factor impacting filterability historically was considered to be certain incomplete hydrolyzed macromolecules, such as β-glucan, arabinoxylan (AX), and prolamine, which was attributed to complex reasons, including low expression level or activity of endogenous hydrolases, differential expression of isozymes among barley cultivars, affections of enzyme regulators, and even the excessive contents of macromolecules in barley. To systematically characterize filterability related proteins in Chinese barley malt, a proteomic strategy as applied. The Canadian Metcalfe barley malt, which has superior filterability, and the Chinese Dan’er barley malt, which has filterability problems, were chosen for comparative proteomic by two-dimensional difference gel electrophoresis (2D-DIGE). The Dan’er and Metcalfe barleys contained similar amounts of macromolecules. While the contents of macromolecules in their Congress wort were significantly different, with 226.1 mg/L of β-glucan and 1,072 mg/L of AX in Dan’er wort, and 102 mg/L of β-glucan and 750.1 mg/L of AX in Metcalfe wort, respectively. The proteomic results showed that Metcalfe malt had more hydrolases such as β-amylase and arabinofuranohydrolase (AXAH-I), while Dan’er malt expressed higher levels of pathogen related proteins, especially the serpins and peroxidase (POD). At the same time, the activity of AXAH-I was monitored during the Congress mash, using Dan’er and Metcalfe malts. Throughout the mashing process, the AXAH-I activity of Dan’er malt was always lower than that of Metcalfe. The highest AXAH-I activity shown by Dan’er wort was 9 mU/g malt, which was only 60% of the activity of Metcalfe (13 mU/g malt). To verify its role in malt filterability, a certain amount of AXAH-I was purified from Dan’er malt and added to the start of the EBC mashing process. It was found that extra AXAH-I added to the mash enhanced the separation rate, decreased viscosity, and also, unexpectedly, increased turbidity. When 6 mU/g malt of AXAH-I was added, the separation rate and turbidity were increased by 34.2% and 32.5%, and viscosity was reduced by 5.5%. Meanwhile, the contents of macromolecules, including high molecular weight AX and β-glucan, were reduced. Dan’er malt was also found to contain more POD, which could promote the cross-link between AX and ferulaic acid (FA) to form feruloylarabinoxylans. The concentration of monomer AX-FA, the activity of POD, and the separation rate of 10 collected malt samples were detected in their Congress wort, the results showed that the wort separation rate was negatively correlated with these factors. With added ascorbic acid, the POD inhibitor to the EBC mashing process using Dan’er malt, the separation rate of Congress wort was significantly increased. The roles of other different proteins in filterability still need to be researched further.
Xiaomin Li is an associate professor at Jiangnan University. Her research focuses on the molecular genetics of yeast and quality control of raw materials for brewing, such as filterability, toxins, and PYF, etc.