Diastatic Power Research Articles

Genotypic and environmental variation in barley limit dextrinase activity and its relation to malt quality

Variation in the limit dextrinase activity of barley malt, and the relationships between limit dextrinase activity and malt quality parameters were investigated using eight cultivars grown at seven diverse locations in China for two successive years. Limit dextrinase activity varied with genotype and location, with the levels ranging from 0.245 U/g to 0.980 U/g. The results showed that the variation in limit dextrinase activity was more attributable to the environment (location and year) than to the genotype. The response of limit dextrinase activity to the environment differed markedly among cultivars, and was reflected by large difference in coefficient of variation of cultivars across diverse locations. Regression analysis showed that limit dextrinase activity was negatively correlated with malt viscosity (r=-0.52, P<0.01), positively correlated with Kolbach index (r=0.38, P<0.01) and malt extract (r=0.30, P<0.05), but had no significant correlation with malt protein content and diastatic power.

Effects of sulphur on yield and malting quality of barley

Eight field experiments were conducted at four sites in the UK in 2003 and 2004 to investigate the effects of sulphur (S) application on yield and malting quality of barley. Significant yield responses to S additions were obtained in five out of the eight experiments, with yield increases ranging from 0.2 to 1.2 t/ha (4.7–22.5%). At the two most S-deficient sites, S application significantly increased malt diastatic power, alpha-amylase activity, friability and homogeneity, and decreased (1→3,1→4)-β-glucan concentration in the wort, indicating an improved endosperm modification during malting. Sulphur applications also significantly increased the concentration of S-methylmethionine (the precursor of dimethylsulphide) in kilned malt, which could impact on beer flavour. When the supply of N was limiting, S applications decreased grain N concentration due to a dilution effect as a result of increased grain yield. In some cases, S applications resulted in decreased grain size. At sites non-deficient or marginally deficient in S, applications of S had little effect on grain or malting quality parameters. The need to maintain an adequate S supply to barley for both yield and malting quality was demonstrated.

Effect of Dilute Alkaline Steeping on the Microbial Contamination, Toxicity and Diastatic Power of Sorghum Malt

Steeping in dilute alkali was investigated with the primary aim of reducing the moulds, coliforms, and preventing toxicity of sorghum malt. Red tannin-free sorghum cultivars NK 283 and PAN 8546 were steeped in 0.1, 0.2 and 0.3% NaOH and 0.1, 0.3 and 0.5% Ca(OH)2. The effects of these solutions on aerobic plate count, lactic acid bacteria, yeasts, moulds, coliforms, the presence of some mycotoxins (aflatoxins, fumonisins, deoxynivalenol and zearalenone), the cytotoxicity and diastatic power (DP) of sorghum malt was evaluated. Steeping NK 283 in 0.1% Ca(OH)2 increased the DP from 10.5 to 16 Sorghum Diastatic Units (SDU)/g. However, it did not reduce the general levels of malt microflora. Steeping in dilute NaOH reduced the microflora on the malt. Steeping both grain cultivars with 0.2% NaOH resulted in malts with the moulds and coliforms reduced to approximately 3.5 and 2.0 log colony forming units/g, respectively and the percentages of some mould species to very low or undetectable levels. The DP of the 0.2% NaOH steeped malts increased to 16.2 and 26.9 SDU/g in the NK 283 and PAN 8546, respectively. There were no detectable amounts of mycotoxins and no indication of cytotoxicity in the 0.2% NaOH steeped samples. Steeping sorghum grains in food-grade 0.2% NaOH is recommended as a method for the control of bacterial and fungal contamination during sorghum malting.

Expression genetics and haplotype analysis reveal cis regulation of serine carboxypeptidase I (Cxp1), a candidate gene for malting quality in barley (Hordeum vulgare L.)

Using a cDNA array-based functional genomics approach in barley, several candidate genes for malting quality including serine carboxypeptidase I (Cxp1) were previously identified (Potokina et al. in Mol Breed 14:153, 2004). The gene was mapped as a single nucleotide polymorphism (SNP) marker on chromosome 3H using the Steptoe (feeding grade)xMorex (malting grade) mapping population. Subsequently, the relative level of Cxp1 expression was determined by real-time RT-PCR for each of the 134 progeny lines and mapped as a quantitative trait. Only one quantitative trait locus (QTL) could be identified that significantly influenced the level of the Cxp1 expression. The expressed QTL maps to the same region on chromosome 3H as does the structural gene and corresponds to a QTL for "diastatic power," one among several traits measured to assess malting quality. An analysis of 90 barley cultivars sampled from a worldwide collection revealed six SNPs at the Cxp1 locus, three of which display complete linkage disequilibrium and define two haplotypes. The Cxp1 expression level in a set of barley accessions showing haplotype I was significantly higher than that of accessions displaying haplotype II. The data provide evidence that (1) the expression of Cxp1 is regulated in cis and that (2) the level of diastatic power in the barley seed is influenced by the level of Cxp1 expression.

Protein and hordein fraction content in barley seeds as affected by sowing date and their relations to malting quality

The effect of sowing date on grain protein, hordein fraction content and malting quality of two-rowed spring barley was investigated by using ten commercial cultivars with different grain protein content and the relationships among these traits were examined. The results showed that grain protein content and B hordein content increased as the sowing date postponed and were significantly affected by sowing date, while C and D hordein contents were less influenced by sowing date. There were significant differences in grain protein and hordein fraction content among the ten cultivars. The coefficient of variation of D hordein content was much larger than that of B and C hordein contents, suggesting its greater variation caused by different sowing dates. Beta-amylase activity and diastatic power were also significantly affected by sowing date, with malt extract being less affected. Significant differences in measured malt quality were found among the ten cultivars. Grain protein was significantly correlated with B hordein and malt extract positively and negatively, respectively. There was no significant correlation between beta-amylase activity or diastatic power and grain protein content. B hordein was negatively and significantly correlated with malt extract, but no significant correlations between C hordein, D hordein and malting quality traits.

Assessing the Impact of the Level of Diastatic Power Enzymes and Their Thermostability on the Hydrolysis of Starch during Wort Production to Predict Malt Fermentability

In this study, commercially produced malts were used for small-scale simulated mashing trials to investigate the impact of differences in the level and thermostability of malt diastatic power (DP) enzymes on the resultant wort fermentability. A modified European Brewery Convention/American Society Brewing Chemists mashing protocol was used with mash-in temperatures ranging between 45 and 76°C for full-malt and 30% rice adjunct mashes. Malt extract yield varied little with mashing temperature for most varieties in this temperature range. However, the fermentability, maltose content, and free amino nitrogen of that extract was considerably affected by mashing temperature with 65°C achieving the highest fermentability for all malt varieties. Multilinear regression analysis of full-malt and rice adjunct mashing trials at 65°C using 43 commercial malts showed that the level of α-amylase and total limit dextrinase activity, Kolbach Index, and the total β-amylase activity level and thermostability were the most important malt quality predictors of wort fermentability. These conclusions suggest that the conventional DP assessment could be replaced with the measurement of its component enzymes outlined above so that maltsters could better satisfy brewers malt quality expectations by blending and defining their malt quality in terms of these fermentability predicting factors. This information would be particularly useful to brewers who brew with multiple varieties and blends from different suppliers. The focus on individual enzyme characteristics by barley breeders is likely to provide selection targets that are more accurate and achievable.

The Microbial Contamination, Toxicity and Quality of Turned and Unturned Outdoor Floor Malted Sorghum

ABSTRACT J. Inst. Brew. 111(2), 190–196, 2005 Turned and unturned outdoor floor malted sorghum were studied for their total microbial contamination, nature and extent of con-tamination by moulds, cytotoxicity (IC 50 ) and quality in terms of diastatic power (DP). The presence of aflatoxins, fumonisins, deoxynivalenol and zearalenone were also investigated. Total microbial counts were high (10 7 –10 8 cfu/g) in both turned and unturned samples. All samples showed contamination by differ-ent moulds, with the dominant being Mucor species, Rhizopus oryzae, Fusarium moniliforme and Phoma sorghina as well as Aspergillus flavus and Alternaria alternata. The latter four are known for producing mycotoxins. Malt samples had very low cytotoxicity (IC 50 from 62.5 to >1000 kg/kg), though all con-tained fumonisins, deoxynivalenol and zearalenone at levels of <0.25–2 µg/g, 15–20 and 10–15 µg/kg, respectively. Malt DP was generally lower in turned samples compared to unturned samples probably because the heat conserved in the latter en-sured better germination conditions. Overall, turning during ger-mination did not affect the microbial load, mould population and levels of deoxynivalenol and zearalenone in sorghum malt but decreased sorghum malt DP. Thus, alternative methods of con-trolling the sorghum malt microbial load should be sought.

Hordein and Malting Quality in Northern Barleys

The objective of this study was to evaluate the effect of D, C, and B hordein polypeptides on malting quality of 19 spring barley (Hordeum vulgare L.) cultivars. Results from the study indicated that B hordein polypeptide combinations had some effect on malting quality through regulating diastatic power. Comparison of the relative amounts of hordein extracted in the presence and absence of reducing agent was made to determine the effect of disulphide bonds on malting quality. In the absence of reducing agent in the extraction solvent, a high concentration of B hordein was correlated with an increase in extract yield. Conversely, in the presence of reducing agent, an increase in B hordein concentration increased the Kolbach Index. A higher D hordein to B hordein ratio, when the former contained a high concentration of disulphide bonds, decreased malting quality to a greater extent than when fewer disulphide bonds were present. When environmental conditions favoured high N uptake efficiency, a larger proportion of D hordein disulphide bonds was synthesized, leading to decreased malting quality. It is speculated that cultivars with lower grain N yield and a lower D:B ratio yield good quality malt.

Fine mapping of a malting-quality QTL complex near the chromosome 4H S telomere in barley.

Malting quality has long been an active objective in barley (Hordeum vulgare L.) breeding programs.However, it is difficult for breeders to manipulate malting-quality traits because of inheritance complexity and difficulty in evaluation of these quantitative traits. Quantitative trait locus (QTL) mapping provides breeders a promising basis with which to manipulate quantitative trait genes. A malting-quality QTL complex, QTL2, was mapped previously to a 30-cM interval in the short-arm telomere region of barley chromosome 4H in a "Step-toe"/"Morex" doubled haploid population by the North American Barley Genome Project, using an interval mapping method with a relatively low-resolution genetic map. The QTL2 complex has moderate effects on several malting-quality traits, including malt extract percentage(ME), a-amylase activity (AA), diastatic power (DP), malt 13-glucan content (BG), and seed dormancy, which makes it a promising candidate gene source in malting barley-cultivar development. Fine mapping QTL2 is desirable for precisely studying barley malting-quality trait inheritance and for efficiently manipulating QTL2 in breeding. A reciprocal-substitution mapping method was employed to fine map QTL2. Molecular marker-assisted backcrossing was used to facilitate the generation of isolines. Fourteen different types of "Steptoe" isolines, including regenerated "Steptoe" and 13 different types of "Morex" isolines,including regenerated "Morex", were made within a 41.5-cM interval between MWG634 and BCD265B on chromosome 4H. Duplicates were identified for 12 "Steptoe" and 12 "Morex" isoline types. The isolines together with "Steptoe" and "Morex" were grown variously at three locations in 2 years for a total of five field environments.Four malting-quality traits were measured: ME, DP, AA,and BG. Few significant differences were found between duplicate isolines for these traits. A total of 15 putative QTLs were mapped; three for ME, four for DP, six for AA,and two for BG. Background genotype seemed to make a difference in expression/detection of QTLs. Of the 15 QTLs identified, ten were from the "Morex" and only five from the "Steptoe" background. By combining the results from different years, field environments, and genetic backgrounds and taking into account overlapping QTLsegments, six QTLs can be conservatively estimated: two each for ME and AA and one each for DP and BG with chromosome segments ranging from 0.7 cM to 27.9 cM. A segment of 15.8 cM from the telomere (MWG634-CDO669) includes all or a portion of all QTLs identified. Further study and marker-assisted breeding should focus on this 15.8-cM chromosome region.

Mapping of QTL for malting quality attributes in barley based on a cross of parents with low grain protein concentration

The barley population used for this research was derived from a cross of parents inherently low in grain protein concentration (GPC). Because of the established inverse relationship between GPC and malt extract, our aim was to characterise quantitative trait loci (QTL) that affect eight malting quality attributes in the hope of identifying novel alleles. The mapping population comprised 180 doubled haploid progeny of a cross between VB9524 with ND11231 ∗12. QTL analysis was based on a 167-point genetic linkage map and malting quality data obtained by micro-malting grains from two trial sites. The number of QTL identified for individual traits ranged from 3 to 5. The distribution across chromosomes showed a paucity of QTL on chromosome 1H and 6H (with one locus each) and a high concentration on chromosome 2H (nine loci). Together, the QTL explained 20–58% of the phenotypic variation observed for each trait. The inclusion of GPC as a covariate increased the number of QTL identified for malt extract and diastatic power, largely due to increased power of detection.

Using unbalanced data from a barley breeding program to estimate gene effects: the Ha2, Ha4, and sdw1 genes

The effects of genes in self-pollinated crops are usually estimated from designed experiments where selection is minimised. In this study, we used a large, but unbalanced, dataset from a barley breeding program to estimate the effects of the Ha2, Ha4, and sdw1 genes on grain yield, grain weight, grain protein, malt extract, and diastatic power. The Ha2 and Ha4 genes for resistance to cereal cyst nematode were under intense selection pressure, whereas the sdw1 gene, which reduces plant height, was under mild selection pressure.From a mixed-model analysis of mainly F5-derived lines over 5 years, resistance due to the Ha2 gene was found to increase grain yields at 2 sites where the nematode was expected to be present, but not at 3 other sites. There was no significant effect of Ha4 on grain yield. Because of selection, data from later stages of evaluation were not useable for Ha2 or Ha4. From analyses of both early stage and later stages of evaluation, the semi-dwarf allele of the sdw1 gene increased grain yields at high-yielding sites, but decreased yields at low-yielding sites. The semi-dwarf allele reduced grain weight. The effects of Ha2 or Ha4 on malt extract and diastatic power were not significant, but the semi-dwarf allele at sdw1 reduced grain protein.We concluded that plant breeding data can be used to successfully estimate the effects of important genes, with bias due to selection minimised by the use of data from appropriate stages of selection and the use of appropriate statistical models.

Dissection of a malting quality QTL region on chromosome 1 (7H) of barley

Malting and brewing are major uses of barley (Hordeum vulgare L.) worldwide, utilizing 30–40% of the crop each year. A set of complex traits determines the quality of malted barley and its subsequent use for beer. Molecular genetics technology has increased our understanding of genetic control of the many malting and brewing quality traits, most of which are quantitatively inherited. The objective of this study was to further dissect and evaluate a known major malting quality quantitative trait locus (QTL) region of about 28 cM on chromosome 1 (7H). Molecular marker-assisted backcrossing was used to develop 39 isolines originating from a ‘Steptoe’ / ‘Morex’ cross. Morex, a 6–row malting type, was the donor parent and Steptoe, a 6–row feed type, was the recurrent parent. The isolines and parents were grown in four environments, and the grain was micro-malted and analyzed for malting quality traits. The effect of each Morex chromosome segment in the QTL target region was determined by composite interval mapping (CIM) and confirmed and refined by multiple interval mapping (MIM). One QTL was resolved for malt extract content, and two QTLs each were resolved for α-amylase activity, diastatic power, and malt β-glucan content. One additional putative malt extract QTL was detected at the plus border of the target region by CIM, but not confirmed by MIM. All QTLs were resolved to intervals of 2.0 to 6.4 cM by CIM, and to intervals of 2.0 cM or less by MIM. These results should facilitate marker-assisted selection in breeding improved malting barley cultivars.

Effect of Germination Moisture and Time on Pearl Millet Malt Quality - With Respect to Its Opaque and Lager Beer Brewing Potential

The effect of germination moisture and time on pearl millet malt quality was investigated. Two pearl millet varieties SDMV 89004 and 91018 were germinated at 25°C under three different watering regimes for 5 days. As with sorghum malting, diastatic power, beta-amylase activity, free α-amino nitrogen (FAN), hot water extract and malting loss all increased with level of watering. However, pearl millet malt had a much higher level of beta-amylase and higher FAN than sorghum malt and a similar level of extract. Malting losses were similar or lower than with sorghum. Thus, it appears that pearl millet malt has perhaps even better potential than sorghum malt in lager beer brewing, at least as a barley malt extender, especially in areas where these grains are cultivated and barley cannot be economically cultivated. Also, its increased use in commercial opaque beer brewing, where sorghum malt is currently used, could be beneficial.

Sorghum malting quality as affected by availability of suitable cultivars

Malting quality of grain samples of commercially available malting class sorghum cultivars was determined over five seasons and three localities for each season. The number of cultivars included in the National Sorghum Cultivar Trials has declined in recent years. The aim of the study was to determine the malting status of current South African sorghum cultivars and to determine whether less suitable cultivars have lowered malting standards. All malting parameters were significantly influenced by season. Seasonal effects on diastatic power were greater than locality and cultivar effects. Malting quality during the 2000/01 season was poor. The malting quality of the total group of GM Class cultivars for each season and three cultivars included in trials over all five seasons were found to be similarly influenced by seasonal effects. The lower malting quality of cultivars recorded during seasons such as 2000/01 was not due to cultivars being less suitable, but was caused by seasonal effects.

Malting Behaviour of Barleys Grown in Canada and Spain as Related to Hordein and Enzyme Content

To continue our effort to analyse the genetic (varietal) and environmental (sites and years) effects on malting quality of barley, we have field-tested four barley varieties, two- and six-rowed, European and North American, in Spain and Canada in 1998 and 1999. The Spanish trials were autumn-sown whereas the Canadian ones were spring-sown. Barley grain was analysed for total protein and hordein contents and micromalted. Canadian-grown barleys had significantly lower contents of grain protein and all-three hordein fractions than the Spanish ones. They also had lower malt respiratory losses, wort β-glucan and viscosity but lower fine- and coarse-ground malt extract yield, friability, free amino nitrogen, Kolbach index, α-amylase and diastatic power. In other words, the Canadian-grown barleys, despite showing lower protein and hordein contents, produced malt of inferior quality than their Spanish counterparts, which, overall, produced higher quantities of degrading enzymes (amylolytic, proteolytic and cytolytic) during germination, thus being able to attain higher extract yield levels.

α-グルコシダーゼを利用したビールの高濃度醸造 (第4報)

High gravity brewing has come to be widely used for increased brewery efficiency. However, it is not easy to maintain good yeast performance during fermentation because of the stress on yeastcaused by high sugar and/or ethanol concentrations. Therefore, sugar attenuation tends to be insufficientand the finished beer contains high concentrations of acetic acid and esters.We previously reported on the development of a new fermentation method by which severaldifficulties in high gravity brewing could be overcome. In this method, α-glucosidase added at thebeginning of fermentation enables the conversion of wort sugars by glycosyl-transfer and hydrolyticreactions to proceed in parallel with ethanol formation by yeast. By using α-glucosidase, we succeededin high gravity brewing with a top fermentation brewer's yeast at a constant fermentation temperatureof 15°. However, bottom fermenting brewer's yeast is more widely used for commercial brewing aroundthe world. In this paper we report results of using the method with bottom fermenting brewer's yeast. We confirmed that the method could be successfully applied to high gravity wort fermentation withbottom fermenting yeast at low temperature. When α-glucosidase was added to 20% Plato wort at aconcentration of 800mg/l, the wort was successfully fermented at 10° with a bottom fermenting brewer's yeast, W 34/70. The real degree of fermentation was 77%, which was 16% higher than that ofthe control fermentation, and the concentration of acetic acid was 200mg/l, which was 85mg/l lowerthan that of the control. The addition of α-glucosidase was also effective for increasing the degree offermentation of dextrinous wort. When α-glucosidase was added at the concentration of 400mg/l to 20% Plato wort containing 6.2% dextrin, the content of dextrin decreased to 3.7% by the end of fermentationwith a top fermenting brewer's yeast, NCYC 1245; the real degree of fermentation was 75%, whichwas 19% higher than that of the control. It is suggested that the use of α-glucosidase is applicable forbrewing good beers from wort that have a low proportion of fermentable sugars, meaning that malt withlower diastatic power could be utilized. The use of enzymes in fermentation would open several newpossibilities to add to existing enzyme methods, such as the use of acetolactate decarboxylase to shortenthe conditioning period or glucoamylase for dry beer brewing.

The changes of β-glucan content and β-glucanase activity in barley before and after malting and their relationships to malt qualities

The cultivar and environmental variations of β-glucan content and β-glucanase activity in grains and malt were investigated in 8 barley cultivars grown at 7 locations. On average, for all cultivars and locations, approximately 80% of β-glucan present in grains was degraded after malting, but there was great variation among both cultivars and locations. β-glucanase activity was much lower but detectable in grains, and it dramatically increased after malting. The cultivar and environmental variations of β-glucan content were much higher in malt than in grains, and malt β-glucan content was more dependent on malt β-glucanase activity than the original level of β-glucan in grains. The correlation analysis of β-glucan content, β-glucanase activity and four malt quality parameters (Kolbach index, diastatic power, viscosity and malt extract) showed that malt-glucan content was highly significantly correlated with all quality parameters, while the association between grain β-glucan content and these quality parameters was relatively weak. Malt β-glucan content was negatively and highly significantly correlated with malt β-glucanase activity, and positively and significantly correlated with grain β-glucan content. It was also found that there was a positive and significant correlation between grain and malt β-glucanase activity.

The Impact of the Thermostability of α-Amylase, β-Amylase, and Limit Dextrinase on Potential Wort Fermentability

Diastatic power (DP), a measure of the level of activity of starchhydrolyzing enzymes, is a term that refers to the combined activity levels of β-amylase, α-amylase, limit dextrinase, and α-glucosidase. The result of the action of these enzymes during brewing is to break down starch into component sugars that are readily fermented into alcohol by yeast. Although DP is a reasonable predictor of fermentability, it does not always accurately estimate the level of fermentable sugars generated during mashing or the subsequent fermentability of the resultant wort. Previously, it has been shown that allelic variation in β-amylase thermostability has a significant impact on wort fermentability. The thermostability of the DP enzymes is critical in determining fermentable sugar yield during mashing, where the mash temperature profile is a balance between the temperature required for starch gelatinization to enable efficient hydrolysis and the rate of thermal inactivation of the DP enzymes. This study demonstrates that there is significant variation in the thermostability of α-amylase and limit dextrinase in malts made from a wide range of Australian and international malting barley varieties. This variation in α-amylase and limit dextrinase thermostability combined with that of β-amylase provides an opportunity for barley breeders to produce divergent malting varieties that better fulfill brewers' fermentability requirements. The implications of the varying DP enzyme thermostability on the selection of malt by brewers to optimally suit different brewing styles and regimens are outlined.

Influence of nitrogen and irrigation on malt and wort quality in barley

Nitrogen and irrigation effects on malt and wort quality traits were studied in two and six row barleys. Varietal effects were highly significant for malt yield, hot water extract (HWE), diastatic power (DP), wort viscosity, Kolbach index (KI), wort filtration rate, malt friability and homogeneity. The increase in nitrogen levels increased the DP, HWE, and decreased the wort filtration rate but was within the permissible limits even at 90 kg N/ha. Irrigation significantly increased the DP, malt yield, KI and malt homogeneity. The increase, though not significant was also observed in wort filtration rate and malt friability with more irrigation.

Evaluation of malting barley quality using exploratory data analysis. I. Extraction of information from micro-malting data of spring and winter barley

Abstract This paper presents an exploratory multivariate approach for analysis of malting barley quality data. By using principal component analysis (PCA) and partial least squares regression (PLSR) complex malting quality data are combined into functional factors which are used for malting barley quality characterisation. Fifty barley samples were used in this investigation, representing 15 spring barley and 10 winter barley varieties grown at two locations in Denmark. The samples were micro-malted and mashed and analysed for 13 quality parameters according to official methods of the European Brewery Convention. These data were combined and reduced into a few latent (functional) factors using PCA by which it is demonstrated that the modification of β-glucan plays a major role in both spring and winter barleys. Additionally, the spring barley and winter barley samples display different covariate latent structures, mainly in the nitrogen and diastatic power patterns. It is furthermore shown that graphic display as facilitated by exploratory data analysis, can be utilized in order to evaluate genotype-environmental interactions by considering the position and movements of the individual objects (genotypes in this instance) in the score plots. Thus, in contrast to the classical analysis of variance, the samples can be individually evaluated and the corresponding loadings can be used to validate the genetic and environmental effect of a given sample in a quality perspective. Several of the investigated malting quality parameters are highly intercorrelated. This fact is utilized by applying PLSR to barley and malt data for the prediction of wort quality in order to exclude the mashing step. This approach was successful for the modification-dependent wort parameters, extract, wort β-glucan and viscosity.