Comparisons of barley malt amylolytic enzyme thermostabilities to wort osmolyte concentrations, malt extract, ASBC measures of malt quality, and initial enzyme activities

Authors: DUKE, STANLEY - University Of Wisconsin; Henson, Cynthia; Vinje, Marcus

Submitted to: Journal of the American Society of Brewing Chemists
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 4/22/2014
Publication Date: 11/20/2014
Citation: Duke, S.H., Henson, C.A., Vinje, M.A. 2014. Comparisons of barley malt amylolytic enzyme thermostabilities to wort osmolyte concentrations, malt extract, ASBC measures of malt quality, and initial enzyme activities. Journal of American Society of Brewing Chemists. 72(4):271-284.

Interpretive Summary: The industrial process that converts seed starch to fermentable sugars, known as mashing, takes place at high temperatures. Barley seed is typically germinated for 4–6 days during the malting process during which time enzymes that convert starch to sugars are either synthetized and/or activated. These enzymes subsequently function under the high temperature mashing conditions of the brewer’s choice. Exposure to high temperature results in the enzymes, to varying extents, become thermally inactivated thus limiting the actual yield of fermentable sugars to less than what can be produced from the starch stored in the seed. This work was conducted to determine the utility of a simple, inexpensive and quantitative measure of the amount of material extracted from germinated barley seeds, osmolyte concentration (OC), during industrial processing for brewing as an indicator of the ability of enzymes to tolerate high temperatures. This work demonstrates that OC successfully predicts the thermostability of the components of the system most limited by high temperatures, which are beta-amylase and limit dextrinase. The impact of this work is the development of a simple and inexpensive method suitable for high throughput application in screening large numbers of barley genotypes for increased efficiency in starch conversion.

Technical Abstract: In this study the hypothesis that wort osmolyte concentration (OC) would correlate much better than malt extract (ME) with barley amylolytic enzyme thermostabilities of malts produced over several days of germination was tested. Seeds of 4 two-row and 4 six-row North American elite barley cultivars were steeped and germinated in a micromalter. At 24 hour intervals throughout 6 days of germination, green malt was removed and kilned. Malts were assayed for a- and ß-amylase and limit dextrinase before and after mashing at 70°C for 30 min to determine thermostabilities. Wort OC, ME, and ASBC measures of malt quality were determined for all days of germination. For all cultivars combined over all days of germination, wort ß-amylase thermostabilities correlated negatively and highly significantly with wort OC and much better than for wort ME. Correlations for limit dextrinase thermostabilities with wort OC were also much better than with ME. Two-row and six-row cultivar ß-amylase and limit dextrinase thermostabilities correlated much better with OC than ME. a-Amylase thermostability was either unaffected or higher after mashing at 70°C. These data suggest that ß-amylase and limit dextrinase thermostabilities become more limiting to starch degradation as reflected by OC than as reflected by ME as germination proceeds. In general, wort OC values peaked with malts from 5 days of germination, whereas ME peaked with malts from 4 days of germination. This difference in pattern of OC and ME development may have contributed to the better correlations of OC with thermostabilities. ß-Amylase intron III allelic variation had no effect on OC or ME on the days where LSD analysis was significant, indicating that, as in past studies, North American barley germplasm, ß-amylase thermostability and activity are not influenced by intron III allelic variation. Except for correlations of initial activities of a-amylase with thermostabilities of initial a-amylase, high initial activities of barley malt a- and ß-amylases and limit dextrinases correlated negatively and significantly with high thermostabilities of ß-amylases and limit dextrinases. For all cultivars combined over six days of germination, the correlations for ß-amylase and limit dextrinase thermostabilities versus the initial activities of a- and ß-amylase and limit dextrinase were significant and negative. These data suggest that selection for high initial activity of any of these amylolytic enzymes would also select for high thermostability of ß-amylase and limit dextrinase.