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1.
Variation in the diastatic power of Australian barley, and the relationships between diastatic power and the starch-degrading enzymes contributing to diastatic power, were investigated in 11 cultivars of barley grown at six diverse locations in Australia. Diastatic power varied with genotype and location, with the levels ranging from 3·1 to 16·5 U/kg. For alpha-amylase activity, levels across cultivar and location ranged from 52 to 214 U/g, for beta-amylase activity they ranged from 201 to 1550 U/g; and, for limit dextrinase activity, they ranged from 56 to 636 U/kg. Alpha-amylase (r = 0·64) and beta-amylase (r=0·77) activities were correlated more strongly with diastatic power than was limit dextrinase (r=0·37). Grain nitrogen content was correlated positively with diastatic power (r=0·71), largely because of the relationship between nitrogen content and beta-amylase activity (r=0·82). High grain nitrogen contents were also associated with small grain sizes (r=−0·76) and low hot-water extracts (r=−0·75). The levels of alpha-amylase activity were correlated more closely with limit dextrinase activity (r=0·65) than with beta-amylase activity (r=0·28). The results indicate the need to select barley cultivars separately for alpha-amylase and beta-amylase activities to achieve high levels of diastatic power.  相似文献   

2.
The activities of endogenous (R-type) and exogenous acting (D-type) protein inhibitors ofalpha-amylase and the activities ofalpha- and total amylase were determined in milling fractions of rye. High D-type amylase inhibitor activities were detected in the embryo (255 IU/g) and in the endosperm fraction (64·9 IU/g), low inhibitor activities were found in the aleurone layer fraction (25·9 IU/g). The highest R-typealpha-amylase inhibitor activity was found in the aleurone layer fraction (32·6 IU/g), and the lowest value in the epidermis containing fraction (5·0 IU/g). The D- and R-typealpha-amylase inhibitor activities varied with growing conditions. D-type amylase inhibitor activities were found to be high in those samples which grew under drought conditions and low in samples cultivated under wet and cool weather. Higher R-typealpha-amylase inhibitor activities were found in rye genotypes cultivated under wet conditions and lower values under dry weather. There were small variations inalpha-amylase inhibitor activities between sprout-stable and sprout-sensitive rye genotypes. The D- and R-typealpha-amylase inhibitor activities of all varieties were stable during 72 h of germination. Similar soil conditions will therefore lead to differentialalpha-amylase inhibitor activities depending on weather conditions during growth.  相似文献   

3.
Proteolytic degradation of barley proteins is examined in green (unkilned) malt and germinating seeds from Hordeum vulgare L. cv. Harrington. Zymographic analysis of the Harrington green malt extracts using commercial preparations of barley beta-amylase incorporated as a proteolytic substrate in 2-D SDS gels shows multiple proteolytic activities. A developmental study shows that the several green malt beta-amylase-degrading activities appear at around day 2 of germination. The several activities appear to increase and decrease through 7 days of germination in a coordinated fashion. Gels treated with class-specific proteinase inhibitors show that serine-class proteinase activities are responsible for barley beta-amylase degradation seen on the zymograms. Western blot analysis also shows that proteolytic enzymes recovered from 1-D electrophoretic gels degrade barley beta-amylase, and that the degradation is inhibited by PMSF. This is the first demonstration that malt proteinases are capable of degrading important metabolic enzymes in germinating barley, and the first postulated physiological role for the serine class proteinases in barley malt.  相似文献   

4.
Diastatic power (DP), a measure of joint alpha- and beta-amylase activities, is the most important quality criterion of sorghum malt. There is a need for a rapid method to estimate sorghum malt DP. Such methods have been developed using both the Falling Number (FN) and Rapid Visco Analyser (RVA) instruments, which measure alpha-amylase activity. Maize starch is used as substrate at a ratio of malt to maize starch of 1:29. Good estimates of DP can be obtained with malts prepared from grain of a single cultivar (FN, r = −0·872; RVA, r = −0·993). The estimate is less good with malts prepared from different cultivars (FN, r = −0·759; RVA, r = −0·759), probably a result of the different cultivars having varying proportions of alpha-amylase relative to DP. The methods are well suited, therefore, to quality control in maltings and breweries, but less suitable for evaluating the malting quality of different cultivars.  相似文献   

5.
The cultivar and environmental variation of beta -amylase activity was studied using two barley cultivars with contrasting growth properties. There was a significant difference in beta -amylase activity between the two cultivars used, 92-11 being significantly higher than Xiumai 3. A significant variation in beta -amylase activity was detected between grains at different positions within a spike. The two cultivars showed the same pattern, with top grains showing the highest and bottom ones the lowest activities. The relative difference within a spike varied between the cultivars, with 92-11 being larger than Xiumai 3. Both seeding rate and timing of N application dramatically affected the beta -amylase activity. With N application at the booting stage, beta -amylase activity increased, mainly due to the significantly increased beta-amylase activity in the topmost grains. The bottom grains showed a lower response to timing of N application. The variation in protein content and grain weight between cultivars and among the various treatments was also examined. The possible influence of these factors on beta -amylase activity are discussed.  相似文献   

6.
The effect of hydrogen peroxide (HP) and ozone (O3) treatment during barley steeping on the quality of malt produced from two barley varieties (GrangeR and AC Metcalfe) by micro-malting was investigated. The two steeping oxidation treatments that was observed to promote barley acrospire growth. Ozone treatment improved the malt enzyme activity of endo-protease, α-amylase, free beta-amylase and total limit dextrinase to differing extents, with GrangeR improving to a greater degree. HP treatment contributed to the increase of α-amylase, β-glucanase and endo-protease. Surprisingly, HP or ozone oxidation during malting resulted in different and novel outcomes for total beta-amylase in GrangeR and AC Metcalfe. In GrangeR, total beta-amylase activity reduced with respect to the control in both treatments. In comparison with AC Metcalfe there was a substantial increase of 78% with HP and 90% O3 in total beta-amylase activity. Malt quality including wort free amino nitrogen, β-glucan, turbidity and diastatic power was differentially increased by the oxidation induction treatment during steeping in malting. Gene expression analysis indicated that the effects of the steep oxidation treatments on enzyme and malt quality were putatively linked with the up-regulation of certain genes involved in GA synthesis (GA20ox1) and ABA catabolism (ABA8′OH). Barley grain germination assay results also showed that moderate HP induction could improve barley germination tolerance to the ABA effect. Malting including steep oxidation induction was shown to be beneficial to malt quality by improving the resultant wort quality and the efficiency of the beer brewing process. These observations point the way towards improving malt quality and the efficiency of the malting process.  相似文献   

7.
Thermostability assays in conjunction with IEF and molecular mapping were used to identify three beta-amylase alleles (Bmyl-Sd1, -Sd2L, -Sd2H) in cultivated barley and an additional allele (Bmy1-Sd3) in an accession of wild barley Hordeum vulgare ssp. spontaneum. The four forms of beta-amylase exhibit different rates of thermal inactivation in barley extracts. This variation was shown to persist after the proteolytic processing of the enzyme that occurs during germination. Three forms of beta-amylase representing the range of thermostabilities were purified and shown to have T50 temperatures of 56·8°C for the Sd2L enzyme, 58·5°C for the Sd1 enzyme, and 60·8°C for the Sd3 beta-amylase from wild barley. Analysis of the relationship between beta-amylase thermostability and fermentability, i.e. the yield of fermentable sugars obtained from starch hydrolysis during brewing in 42 commercial malt samples suggests that increased thermostability results in more efficient starch degradation. Screening for specific beta-amylase alleles is proposed as a method for increasing fermentability in malting barley.  相似文献   

8.
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9.
The frequency and mechanisms of four modes of alpha -amylase enzyme accumulation in U.K. wheat, retained pericarp alpha -amylase activity (RPAA), pre-maturity alpha -amylase activity (PMAA), pre-maturity sprouting (PrMS) and post-maturity sprouting (PoMS), were investigated in field and laboratory experiments. Of 56 cultivar site year combinations (four model cultivars grown at up to four sites from 1994–1997), enzyme activity was detected in 32 cases, in 23 cases sufficient to reduce Hagberg falling number (the usual industry measure of alpha -amylase) below the commercial criterion (250 s). The frequency of occurrence of different modes of enzyme accumulation was in the order PoMS>PMAA>PrMS>RPAA. Both PMAA and PrMS were more common than expected and the most usual pattern was for alpha -amylase to accumulate by several modes. Although green grains are rejected as impurities, study of grain colour in relation to pericarp alpha -amylase activity showed that the enzyme could persist in non-green grains in levels sufficient to affect the Hagberg value. Two factors thought to promote PMAA, grain drying rate and transient changes in temperature in early development, were studied in the field and controlled environment cabinets. No significant difference was found in grain drying rate between samples where PMAA was or was not identified. However, out of 19 transfers from a cool (16/10 °C) to a warm (26/20 °C) temperature regime, six led to significant increases in PMAA. No transfers after 45% grain moisture increased PMAA. PrMS occurred as early as 67% grain moisture and susceptibility usually increased with stage of development, being greatest in the grain dough stage. PrMS susceptibility varied with cultivar (in the same order as PoMS sensitivity) and was affected by environmental factors.  相似文献   

10.
The barley (Hordeum vulgare L.) varieties, Franklin and Schooner, contain two different allelic forms of beta -amylase (EC 3.2.1.2) encoded on chromosome 4H by the Bmy 1-Sd1 and Bmy 1-Sd2L alleles, respectively. The corresponding enzymes, referred to as Sd1 and Sd2L, were purified from both mature barley grain and germinated barley (green malt), and their physical and kinetic properties studied. Approximately 4 kDa were cleaved from both Sd1 and Sd2Lbeta -amylases after germination. The Kmvalue for green malt beta -amylase was less than that of mature grain beta -amylase for both varieties when potato starch was used as a substrate, although Vmaxwas similar. This indicated that proteolysis after germination increased the affinity of beta -amylase for potato starch. No significant kinetic differences were observed between beta -amylase from mature grain and green malt of the two barley varieties when amylose (degree of polymerisation 100 and 18) and maltopentaose were used as substrates. Kinetic differences were also observed between the two allelic forms of beta -amylase. Sd1 beta -amylase from green malt exhibited a lower Kmvalue for potato starch than Sd2L beta -amylase, demonstrating that at non-saturating starch concentrations Sd1 beta -amylase is better able to hydrolyse starch than Sd2L beta -amylase. As the degree of polymerisation of the substrates decreased from approximately 740 (potato starch) to 5 (maltopentaose), the Kmvalues for beta -amylase increased, whereas Vmaxvalues decreased. Maltose, the hydrolytic product of beta -amylase, was found to be a weak competitive inhibitor of both Sd1 and Sd2L green malt beta -amylases with respect to potato starch and amylose. Taken together the kinetic observations for bet a-amylase suggest that the allelic differences and C-terminal proteolysis might be exploited to improve the efficiency of starch hydrolysis during the mashing stage of the brewing process.  相似文献   

11.
Two barley cultivars differing in grain size and protein content were used to investigate the effects of nitrogen nutrition, cultivar and their interaction on grain protein content, hordein content and beta-amylase activity and the relationship between hordein content and beta-amylase activity during in vitro spike culture. The content of protein and hordein fraction, and beta-amylase activity in barley grains increased as the nitrogen level in culture solution increased. Grain protein content was significantly affected by nitrogen treatment and cultivar, and there was no significant interaction between nitrogen treatment and cultivar. Hordein content and beta-amylase activity were significantly affected by nitrogen treatment and cultivar as well as their interaction. Beta-amylase activity was positively correlated with grain protein and hordein contents, and the ratio of hordein B:C was negatively correlated with total protein content and beta-amylase activity.  相似文献   

12.
A double antibody, sandwich enzyme-linked immunosorbent assay (ELISA) was developed using polyclonal antibodies specific tobeta-amylase to estimate the amount of ‘free’ (soluble in aqueous saline solution) or ‘combined’ (extracted with saline solution including reducing agent)beta-amylase protein in barley grain and malt. This ELISA was used to quantify the amount ofbeta-amylase in barley grain and malt from four varieties grown at nine sites in South Australia in 1993. The antibody used to develop the ELISA reacted differently withbeta-amylase depending on whether the source was barley grain or malt, and on thebeta-amylase band pattern in isoelectric focussing (IEF) of the barley variety. On the basis of their IEF band patterns barley varieties were divided into two types, designatedBmy1-Sd1 andBmy1-Sd2. Malting resulted in proteolytic cleavage of thebeta-amylase peptide with a reduction in the apparent molecular weight of up toMr4000 and the appearance of new maltbeta-amylase IEF bands that were more basic. The new maltbeta-amylase IEF band patterns still allowed the identification of theBmy1-Sd1 andBmy1-Sd2 IEF types despite the change in molecular weight and pI. The data obtained using thebeta-amylase ELISA were highly correlated withbeta-amylase activity for both the free and combined fractions when the IEF band pattern and its source, barley grain or malt, were taken into account.  相似文献   

13.
Ileal endogenous N losses (ENL) were measured, using the15N isotope dilution technique, in piglets (17 kg) fed different barley genotypes (naked, spring, winter with low/high beta -glucan content) or diets containing 330, 530, 730 or 930 g of a blend of barleys/kg diet. The apparent protein and amino acid digestibilities of the naked variety and the winter variety with a high beta -glucan content were, on average, significantly higher than those for the other two varieties. The ENL were inversely correlated (p<0·01) with the apparent digestibilities but the difference between each of them was not significant (p>0·05). The ENL increased linearly with the inclusion level of barley in a N-free basal diet (2 mg endogenous N/g barley). Isolated hulls added to a N-free diet at the rate of 100 or 200 g/kg diet exerted no significant effect on the ENL (1·80 g endogenous N/kg diet in both cases vs. 1·76 g for the basal level). On the contrary, the effect of isolated bran, measured under similar conditions, was significantly higher and dependent on fibre intake (2·59 and 3·31 g N/kg diet, respectively). It is concluded that the ENL are affected by the insoluble bran fibre but not by the hulls, nor by the level of beta -glucan.  相似文献   

14.
The acid extract viscosities and β-glucan contents of ten two- and six-rowed barley cultivars grown at seven locations in three consecutive years in Spain were studied in the present work. The viscosities varied from 2·4 to 24·8 centistokes (cSt) and the mean value was 6·4 cSt. The average β-glucan content of barleys determined by HPLC was 3·5% with a range of 1·9–5·5%. Significant differences were found in both β-glucan content and acid extract viscosity between different cultivars, locations and years. The β-glucan contents and viscosities of winter cultivars were higher than those of spring. Cvs. Barbarrosa and Hatif de Grignon were the genotypes with the highest values for both parameters, while cv. Beka had the lowest viscosity and β-glucan content. Environmental factors influenced both parameters. The acid extract viscosities of barleys were correlated negatively with the amount of precipitation (r=−0·754;P<0·05). Barleys grown in wet and rainy areas (Girona and La Coruña) had lower viscosity values.  相似文献   

15.
Response surface methodology was used to determine the levels ofalpha-amylase,beta-amylase and limit dextrinase enzymes required for efficient conversion of starch to fermentable sugars during mashing. Micro-scale mashes with purified barley starch and malt enzymes were performed in a Brewing Research Foundation mash bath, and mash liquors were analysed for solubilised starch, reducing sugars (neocuproine assay) and fementable sugars (anion exchange HPLC). Fermentable sugars in the mash liquor were positively correlated with reducing sugars (R2=0·94) and the percentage of starch solubilised during mashing (R2=0·68). A multiple regression equation relating the levels of the three starch degrading enzymes to the percentage of starch hydrolysed to fermentable sugars gave a good fit to the second order response surface (R2=1·00, RMSE=1·37%). Addition of limit dextrinase to the mashes resulted in a substantial increase in levels of fermentable sugars, and limit dextrinase showed a synergistic effect in increasing levels of maltose in the mash liquor when combined with high levels ofbeta-amylase. The efficiency of any one starch degrading enzyme in a mash is influenced by the presence of other starch degrading enzymes. Commercial malts contain excess levels ofbeta-amylase and below optimal levels of limit dextrinase. Malt extract may not be a good indicator of the level of fermentable carbohydrates produced during mashing.  相似文献   

16.
The influence of milled grain particle size on the kinetics of enzymatic starch digestion was examined. Two types of cereals (barley and sorghum) were ground, and the resulting grounds separated by size using sieving, with sizes ranging from 0.1 to 3 mm. In vitro enzymatic digestion was performed, using pancreatic alpha-amylase, amyloglucosidase and protease, to determine fractional-digestion rates over 24 h. The resulting glucose production rate data were well fitted by simple first-order kinetics. For each sieve screen size, the digestion rate of barley was always higher than that of sorghum. The rate coefficients for digestion showed a decrease with increasing size, and could be well fitted by an inverse square relationship. This is consistent with the supposition that starch digestion in these systems is controlled by diffusion of enzyme through the grain fragment. Apparent diffusion coefficients of alpha-amylase obtained by fitting the size dependence were 0.76 (sorghum) and 1.7 (barley) × 10−7 cm2 s−1, 9 (sorghum) and 4 (barley) times slower than predicted for a molecule of the size of alpha-amylase in water.  相似文献   

17.
The three beta -amylase genes (Bmy1, 2 and 3) in cultivated barley were mapped to chromosomes 4HL, 2HL And 4HL respectively using RFLP analysis. No recombinants between Bmy1 andBmy3 were detected among 264 DH lines. Polymorphism of the Sd1 and Sd2 isoenzymes of beta -amylase co-segregated with the Bmy loci on chromosome 4HL in a doubled-haploid population of the cross Chebec (Sd2)×Harrington (Sd1). This locus also explained 90·5% of the variation in the level of free enzyme between the two parents. Two cDNAs ofbeta -amylase were isolated by RT-PCR from the developing grains of Harrington (Sd1) and Galleon (Sd2). Alignment of the deduced amino acid sequences identified three amino-acid substitutions between the Sd2 and Sd1 forms of beta -amylase (Arg115 – Cys, Asp165 – Glu, and Val430 – Ala). Three allele-specific PCR primer pairs based on the three amino acid substitutions were used to amplify the beta -amylase genes in genomic DNA of sixteen barley cultivars/lines. Only the Arg115(Sd2)/Cys(Sd1) substitution was consistent with the isoenzyme form. This amino acid replacement reduced the pI of the Sd1 beta -amylase consistent with the fact that the Sd2 form is more basic than the Sd1 form when separated by IEF. The mutation from Arg115 to Cys in the Sd1 form also provides one more -SH group to form S-S-bridges. As bound beta -amylase is linked to the insoluble proteins of the endosperm and its inhibitor via disulphide bridges this could explain the higher level of binding exhibited by Sd1 vs Sd2. Thus a single amino acid substitution determines both the isoenzyme type and beta -amylase binding.  相似文献   

18.
A method using methanolic sulphuric acid as transmethylating reagent was developed for determining the fatty acid composition of lipids of oats. The method was optimised for reaction conditions and applied to the determination of the fatty acid composition of lipids of a number of varieties of Australian oats grown in several locations. Thirteen fatty acids were detected with oleic, linoleic and palmitic acids comprising more than 95% of the total fatty acids. Total lipid content of the oats was positively related to the proportion of stearic (r=0·32) and oleic (r=0·81) acids and negatively correlated with the proportion of palmitic (r=−0·64), linoleic (r=−0·39) and linolenic (r=−0·65) acids. Significant positive correlations were found between total lipid content and absolute content of the major fatty acids (r=0·670·98), except for linolenic acid (r=0·12). Environment had significant effects on fatty acid composition, but variety was the controlling factor. The broad sense heritability estimated from individual plot ranged from 69 to 73% and that from the average of three replications and eight locations ranged from 94 to 98% for the major fatty acids. It is possible to improve fatty acid composition of oats by breeding procedures.  相似文献   

19.
Endoproteases play an important role in barley germination by controlling the hydrolysis of the grain's storage proteins into peptides and amino acids that are needed by the young plant. During malting, the commercial version of this process, many high Mr barley biopolymers are converted into malt nutrients that can be utilized by yeasts during brewing. However, barley and malt both contain endogenous proteins that inhibit the enzymatic activities of these proteases. High levels of these inhibitors can cause brewing problems by preventing the proteases from producing optimal levels of soluble proteins and amino acids. Both high and low Mr inhibitors of cysteine proteases occur in barley and malt. Two of the high Mr inhibitors, lipid transfer protein 1 (LTP1) and LTP2, have been purified and studied. Recently, members of the trypsin/alpha-amylase inhibitor protein family (CM proteins) have been shown to inhibit the activity of SEP-1, a purified serine class barley protease. No inhibitors of aspartic proteases or metalloproteases have yet been purified, but it has been reported that endogenous metalloprotease inhibitors do exist. The inhibitors of the cysteine proteases and metalloproteases are probably the ones most important for brewing, because members of these two protease classes apparently catalyse most of the protein hydrolysis that occurs during malt mashing and, presumably, also during malting. More biochemical studies are needed to clarify how these proteins interact with the proteases to control protein hydrolysis during germination.  相似文献   

20.
A group of low Mr wheat proteins with characteristic extractability behavior was isolated using two different isolation procedures. The proteins were extractable with water, salt solution and 70% (v/v) ethanol. After water extraction of flour and separation of gluten, a substantial proportion of these proteins was still extractable from gluten using 70% (v/v) ethanol. Based in their amino acid compositions, Mrs and IEF patterns, the isolated proteins resemble closely most of the alpha -amylase/protease inhibitors described in the literature. This was confirmed by enzyme inhibition studies in which it was shown that they inhibited mammalian, but not wheat, bacterial and fungal alpha-amylases. All proteases tested were inhibited by the low Mr proteins. Their Mrs and their high cysteine contents (6·5-8·1 mol%) indicated that the proteins contain four to five disulphide bonds. Free thiol groups were not detected in the proteins. Upon reduction, the Mr increased from 7-8000 to 14-19000. Furthermore, the disulphide bonds were highly reactive as determined by their reaction with the thiol-specific label monobromobimane. This suggests that the low Mr wheat proteins may play a role in thiol group/disulphide bond exchange in wheat proteins.  相似文献   

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