Submitted to: Journal of American Society of Brewing Chemists
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: September 29, 2011
Publication Date: March 2, 2012
Citation: Duke, S.H., Vinje, M.A., Henson, C.A. 2012. Tracking amylolytic enzyme activities during congress mashing with North American barley cultivars: Comparisons of patterns of activity and ß-amylases with differing Bmy1 Intron III alleles and correlations of amylolytic enzyme activities. Journal of American Society of Brewing Chemists. 70(1):10-28. Interpretive Summary: This research was conducted to determine if the three most important barley enzymes responsible for converting stored starch to fermentable sugars had the same patterns of activity during the industrial process of starch conversion and to determine if these activity patterns vary among some of the most important germplasm used in commerce in North America and/or used in the development of new cultivars for commercial use in malting or as feed. Germplasm was selected to represent a diversity of malts, including barleys with different head types that are adapted for different growing regions of the country and have different malting quality and different gene sequences in one enzyme that is frequently subjected to genetic selection by barley breeders around the world. We demonstrated that differences in the gene sequence frequently targeted for selection are not useful in selection of U.S. germplasm with enhanced malting quality and that two of the three enzyme activity patterns are sufficiently static across genotypes that a single measure of activity conducted early in the process is sufficient to predict the true contribution of these enzymes to starch conversion. This eliminates the need to monitor these enzyme activities continuously throughout the process, thereby increasing the efficiency of operations in quality control laboratories, and eliminates the need for U.S. malting barley breeders to invest in the molecular marker studied.
Technical Abstract: This study was conducted to test three hypotheses: 1) that a-amylase will have less consistent patterns of activity during mashing than ß-amylase and limit dextrinase 2) that differing ß-amylase 1 intron III alleles (Bmy1.a and Bmy1.b) would not be useful in predicting high or low activities or thermostabilities amongst cultivars, and 3) that correlations of a-amylase versus ß-amylase activities would be more significant than for either amylase versus limit dextrinase during the initial 55 min of mashing that produces the bulk of wort sugars. Malts of six two-row and six six-row barley cultivars were mashed and amylolytic enzyme were assayed at 6 time points during mashing. a-Amylase activities of four cultivars peaked at 30 min (45ºC), six at 55 min (70ºC), and two at 75 min (70ºC). ß-Amylase activities of all cultivars peaked sharply at 30 min (45ºC) into mashing and declined precipitously thereafter, rapidly approaching near nil by 75 min. Limit dextrinase activities of 11 of 12 cultivars peaked at 55 min (70ºC) and one peaked at 30 min (45ºC). These data indicate no uniform pattern for the development of a-amylase activity as compared to patterns for ß-amylase and limit dextrinase during mashing. Cultivar Steptoe, a feed barley, had much lower peak activities for all three amylolytic enzymes than malting cultivars. The highest ß-amylase activities during mashing of cultivars with either the Bmy1.a (Legacy, Lacey, Morex) or Bmy1.b (Merit, Harrington) intron III alleles were not significantly different as determined by least significant difference (LSD) analysis (P<0.0001). As temperatures increased from 45ºC to 70ºC (55 min), percentages of the highest ß-amylase activities decreased from 11.9 (Rasmusson) to 41.5% (Steptoe). LSD analysis revealed that Steptoe (Bmy1.a allele) had significantly higher ß-amylase thermostability malting cultivars. The highest malting barley thermostabilities for ß-amylases [Lacey (Bmy1.a allele)] and [Harrington (Bmy1.b, allele)] were not significantly different (P<0.0001). Correlations of a-amylase versus ß-amylase activities were positive and significant in 7 of 9 possible correlations from 5 to 55 min of mashing, whereas a-amylase and ß-amylase versus limit dextrinase activity correlations over the same period were significant in 2 of 9 and 1 of 9 correlations, respectively. This study supports all three of the proposed hypotheses.