RATE-LIMITING ENZYMES IN THE LIBERATION OF AMINO ACIDS IN MASHING

When 29 malts representing a wide variation in free amino nitrogen of wort were mashed for 3 h at 45°C, the increases in TCA-soluble amino nitrogen varied from 47 to 116 μmoles per gramme of malt. The carboxypeptidase activities of the malts showed only small positive correlations with the increase of amino nitrogen during mashing. Inactivation of the different malt carboxypeptidases by 70 to 90% with diisopropylfluorophosphate decreased the liberation of amino nitrogen by only 5 to 25%. Moreover, addition of purified malt carboxypeptidase I to the mashing suspensions had no effect on the liberation of amino nitrogen. It is therefore evident that the carboxypeptidases do not have a major rate-limiting role in the liberation of amino acids in mashing, although it has earlier been shown that they are essential enzymes in this process. The proteinase activities of the malts, determined using two methods, correlated strongly with the liberation of amino nitrogen in mashing. Addition of crude malt proteinase to the mash increased, and addition of purified barley proteinase inhibitor decreased the measured proteinase activity and the liberation of amino acids roughly to the same extent. These results indicate that the most important rate-limiting enzyme activity in the liberation of amino acids in mashing is the proteinase activity of the malt.     

THE FATE OF AMINO ACIDS DURING MASHING AND HOP BOILING

In an infusion mash system the concentration of all α amino acids in sweet wort reaches a maximum value when approximately one-third of the total volume extracted has been run from the mashing vessel. The relative concentrations of each α amino acid in wort vary throughout the mash tun extraction owing to differences in rate of extraction of the amino acids and to the quantities produced by peptidase activity in the mash tun. The relative proportions of proline, alanine, glutamine and asparagine in the total α amino nitrogen content of wort decrease appreciably as the mashing process proceeds. Copper boiling results in the partial destruction of methionine, glutamic acid, lysine, arginine and valine present in sweet wort together with an increase in the concentration of ammonium ion in the wort.     

糖化过程中二甲基硫的形成及控制

研究了糖化过程中二甲基硫（DMS）及前体的变化，糖化工艺参数对定型麦汁中DMS及前体的影响。结果表明；在浸出法糖化过程中，二甲基亚砜（DMSO）在洗糟之前其含量基本不变，硫甲基蛋氨酸（SMM）含量逐渐上升，DMS的含量逐渐下降，随着辅料比的增大，定型麦汁中SMM和DMSO的含量逐渐下降；煮沸温度越高，时间越长，强度越大，麦汁SMM分解得越彻底，采用抽真空喷淋工艺能消除大部分回旋沉淀过程中产生的游离DMS。     

The Evolution of Dextrins During the Mashing and Fermentation of All‐malt Whisky Production

The production of malt whisky involves the mashing of barley malt, followed by the fermentation of the resulting wort without further treatment. While this process has many parallels to the production of an all‐malt beer, one of the main differentiating steps during substrate preparation is the inclusion of a boiling step for the wort in the production of beer. Other than the destructive action of the boiling process on microorganisms, the boiling also destroys all malt enzyme activity. Since a typical whisky wash is not boiled it carries through a certain proportion of microbial activity associated with the malt, but more importantly it retains some enzyme activity that has been activated during the malting and mashing processes. The changes in sugars and dextrins during both mashing and fermentation of the resulting wash were investigated. Evidence of the continuous amylolytic activity during an unboiled, all‐malt wash fermentation is shown; while no ongoing amylolytic activity could be deduced during the fermentation of a boiled all‐malt wort. Furthermore, the data suggests that the amylolytic activity during mashing and fermentation are different with regards to α‐amylase action linked to its multiple‐attack action pattern as a function of substrate conformation, temperature, and effectiveness of potential hydrolytic events.     

Antimicrobial activity of Enterococcus faecium L50, a strain producing enterocins L50 (L50A and L50B), P and Q, against beer-spoilage lactic acid bacteria in broth, wort (hopped and unhopped), and alcoholic and non-alcoholic lager beers

Enterococcus faecium L50 produces enterocin L50 (L50A and L50B) (EntL50, EntL50A and EntL50B), enterocin P (EntP) and enterocin Q (EntQ) and displays a broad antimicrobial spectrum against the most relevant beer-spoilage lactic acid bacteria (LAB) (i.e., Lactobacillus brevis and Pediococcus damnosus), which is mainly due to the production of EntL50 (EntL50A and EntL50B). Bacteriocin assays using in vitro-synthesized EntL50 (EntL50A and EntL50B) showed that both individual peptides possess antimicrobial activity on their own, EntL50A being the most active, but when the two peptides were combined a synergistic effect was observed. The only virulence genes detected in E. faecium L50 were efaAfm (cell wall adhesin) and ccf (sex pheromone), and this strain was susceptible to most clinically relevant antibiotics. E. faecium L50 survived but did not grow nor showed antimicrobial activity in hopped and unhopped wort, and alcoholic (1 and 5% ethanol, v/v) and non-alcoholic (0% ethanol, v/v) commercial lager beers. However, when unhopped wort was supplemented with 50% (v/v) MRS broth, E. faecium L50 grew and exerted antimicrobial activity similarly as in MRS broth. The enterocins produced by this strain were bactericidal (5 log decrease) against P. damnosus and Lb. brevis in a dose- and substrate-dependent manner when challenged in MRS broth, wort (hopped and unhopped), and alcoholic (1 and 5% ethanol, v/v) and non-alcoholic (0% ethanol, v/v) lager beers at 32 degrees C, and no bacterial resistances were detected even after incubation for 6-15 days. The enterocins in wort and lager beer (5% ethanol, v/v) withstood the heat treatments commonly employed in the brewing industry during mashing, wort boiling, fermentation, and pasteurization, and retained most of their antimicrobial activity in lager beer (5% ethanol, v/v) after long-term storage at 8 and 25 degrees C.     

Characterisation and Assessment of the Role of Barley Malt Endoproteases During Malting and Mashing1

Barley malt endoproteases (EC.3.4.21) develop as multiple isoforms mainly during grain germination and pass through kilning almost intact. Thermostability, under simulated mashing conditions, varied from low to high depending on the substrate used in the assay. This suggests that individual enzymes respond differently to heat exposure and to protein substrates. The optimal pH with haemoglobin was pH 3.5, with hordein pH 4 and with glutelin pH 5. The optimal temperature with hordein was 40°C, with glutelin 50°C and with haemoglobin 60°C. These differences suggest that it is not possible to comprehensively characterise all malt endoproteases under one set of assay conditions. In brewing, most of the barley protein degradation (> 70 %) occurs during malting. But some proteinases remain active during mashing and contribute to wort soluble proteins and free amino nitrogen. Their contribution to all malt EBC mash total free amino nitrogen was 25 % in Schooner (Australian) and 30 % in Morex (USA). The importance of proteolytic activity during mashing and the possibility that the levels may not be adequate, at high solid adjunct ratios, are acknowledged.     

THE RELATIONSHIP OF DIMETHYL SULPHIDE LEVELS IN MALT,WORT AND BEER

The half-life for the conversion of malt dimethyl sulphide (DMS) precursor to free DMS has been determined at various temperatures and pH values. At pH 5·2 the half-life of the precursor in wort (S.G. 1·060) at its boiling point is 38 min, and is doubled for each 6°C fall in temperature. At pH 5·5 the half-life at the boiling point is 32·5 min. Knowing the stability of the precursor at the various temperatures in the brewing process, the extent of conversion to free DMS in wort at pitching can be predicted for malt of a given precursor content and for a given set of process conditions. The results of DMS analyses of samples taken during brewery trials are in reasonable agreement with the predicted values. This work involved infusion mashing only, but the same principles apply to decoction mashing. The fate of precursor and free DMS during fermentation and conditioning has been followed on a production scale. With some brews, where high levels of free DMS were present at pitching, much free DMS was lost during fermentation. Also, precursor DMS reappeared in the beer after a few days and there was some increase in the level of free DMS. The DMS precursor in green malt has been isolated by ion-exchange and gel-filtration chromatography. A preparation has been obtained which has 0·6 mol potential DMS per mol amino nitrogen. Thin layer chromatography showed that the preparation and its hydrolysis product had the same R_f values as S-methylmethionine and homoserine respectively.     

pH IN MALTING AND BREWING—A REVIEW

Variations in pH during malting, kilning, mashing, lautering, wort boiling and fermentation are reviewed. The fragmentary information available suggests that the pH of cold water extracts of barley falls during steeping, rises during germination and falls again during kilning. Brewing liquor, mashing method, addition of acid, and the nature of the malt influence the pH of unhopped wort, which falls by about 0·3 of a unit during boiling. Fermentation reduces the pH by a further unit yielding ales with pH values of about 3·8-4·2, or lagers of pH 4·2–4·8. Literature evidence does not suggest any obvious reason why ales and lagers should differ in pH.     

酒精发酵中应用酸性蛋白酶的研究

对酸性蛋白酶在酒精发酵中的应用进行了研究。添加酸性蛋白酶有利于原料中蛋白质的水解，增加醪液中酵母可吸收性氮，促进酵母菌生长繁殖，提高酒精发酵速率。而且，a-氨基氮的增加，减轻了酵母菌细胞氨基酸合成代谢的负荷，使底物（葡萄糖）更多地转向发酵生成乙醇，提高原料出酒率。接种酵母后，在醪液中添加0．01％的酸性蛋白酶，发酵时间由原来的52h缩短到32h，发酵成熟醪酒精含量为10．2％（V）。着延长发酵时间至44h，则发酵成熟醪中酒精含量由原来的9．8％（V）增加到10．7％（V），并对添加酸性蛋白酶和（NH4）2SO4的发酵结果进行了比较，认为酸性蛋白酶是实现高效率酒精发酵的优选促进剂。     

Importance of Endosperm Modification for Malt Wort Fermentability1

Malt wort fermentability is dependent on an adequate supply of the essential nutrients required by yeast. The barley and malt factors affecting this supply of nutrients are not well understood. This study used two doubled haploid populations (Arapiles × Franklin, TR251 × HB345), the latter with a hulless barley parent, to investigate effects of barley and malt quality on fermentability. Populations were grown and malted at different locations resulting in a broad range in malt quality with significant differences in extract, modification and enzyme levels. Fermentability, as indicated by apparent attenuation limit and ethanol levels, also showed significant differences among samples. Modification was the most important factor for good fermentability. There appeared to be several different mechanisms by which modification affected fermentability. High viscosities, slow starch release during mashing, increased glucose supply from better β‐glucan breakdown and increased free amino nitrogen levels all affected fermentability. Effects of starch‐degrading enzymes on fermentability became more significant in better modified malts with α‐amylase showing stronger effects than diastatic power. The poorer fermentability of hulless barley malt was predominately due to low levels of α‐amylase, although, free amino nitrogen also appeared to be an important factor.     

探讨革新的糖化生产技术与设备

<正> 当今先进的技术已将啤酒酿造工艺带进了另一个新国度。今天,高质量的麦芽糖化工艺和低麦汁浊度已成为啤酒质量和味道稳定性的重要指标。 以融汇新技术知识为基础,针对于设备的功能性和有效利用性,ZIEMANN公司不断在改进其糖化设备     

Studies on the mashing conditions of teff (Eragrostis tef) malt as a raw material for lactic acid‐fermented gluten‐free beverage

Formation of extracts and fermentable sugars during mashing can be limited by incomplete starch gelatinisation. The aim of this research was to develop mashing programme for 100% teff malt as a potential raw material for gluten‐free lactic acid‐fermented beverage. Isothermal mashing at temperatures ranging between 60 and 84 °C was conducted, and the highest extract (85%) was observed for the wort samples produced at temperatures higher than 76 °C. Sixty‐minute rest at 71 °C resulted in higher fermentable sugars than other tested conversion rest temperatures. Inclusion of lower mashing‐in temperature in the mashing programme also substantially improved the concentrations of free amino nitrogen (128 mg L^?1) and fermentable sugar (58 g L^?1) in the final wort. Therefore, 30‐min rest at 40 °C followed by 60‐min rest at 71 °C and 10‐min rest at 78 °C was found to be a suitable mashing programme for teff malt.     

INVESTIGATION OF TRANSAMINASE ACTIVITY DURING MASHING

Malt peptidase activity has been determined by Jones & Pierce as the difference in quantities of free amino acids produced under standard conditions between an enzymic mash and one in which the enzymic activity has been destroyed with boiling absolute alcohol. The measurement is a reasonable assessment of individual mash tun peptidase activity provided no interconversion of amino acids takes place during the mashing. Any such interconversion would involve the participation of transmination reactions in the malt. As a means of detecting transaminase activity, malt was mashed at 50° C. in the presence of ¹⁵N-l -glutamic acid and ¹⁵N-l -alanine, and the extracted amino acids were analysed for ¹⁵N by mass spectrometry. Little transaminase activity was detected.     

新型添加剂Brewbrite在麦汁制造中的应用

对Brewbrite在麦汁中的应用进行了初步研究．通过麦汁外观和其他指标的分析。结果表明,在麦汁煮沸结束前10min添加Brewbrite 30mg／L,可以有效地澄清麦汁,降低麦汁中的可凝固性氮含量。与卡拉胶、单宁、HD-003,DHG,PVPP等添加剂相比,Brewbrite可降低麦汁和啤酒中的总多酚含量、可凝固性氮含量、麦汁色度、麦汁浊度等,减少高分子氮的比例,有助于提高啤酒的非生物稳定性。（孙悟）     

Evaluation of the potentials of millet,sorghum and barley with similar nitrogen contents malted at their optimum germination temperatures for use in brewing

Studies on the malting physiology of barley have led to similar studies on millet and sorghum. This study compares the outcomes of the malting physiology of millet, sorghum and barley. Results show that optimal development of diastatic power, soluble nitrogen, hot water extract and the wide range of amino acids of these three cereals is related to optimal malting conditions and appropriate mashing procedures. Transfer of the nitrogen/extract/soluble nitrogen/diastatic concepts of barley malt do not apply to millet and sorghum. However, all the cereals studied produced the range of amino acids required by yeast for fermentation. Sorghum malt released the highest amounts of group 1 amino acids, usually taken up faster by yeast. It also produced and released the highest amounts of amino acids, classified as group 2, which are assimilated more slowly than group 1 amino acids. It also produced and released more of the amino acids that are slowest to be assimilated during fermentation, as well as very high levels of proline. Optic barley malt produced and released the least amount of proline. The fate of proline during yeast fermentation is not clear, but it is believed that proline is not utilized during fermentation. Copyright © 2013 The Institute of Brewing & Distilling     

WORT-BOILING IN RELATION TO BEER QUALITY

Comparisons of beers from worts mashed with pale ale malts (nitrogen 1·3%) at 150°F. (65·5°C.) and boiled for 1/2, 1 and 2 hr. show that increase in boiling time leads to slightly increased hop-utilization, decreased head-retention, improved non-biological stability and slight differences in flavour. Findings were very similar when the mashing temperature was either 145°F. (63°C.) or 155°F. (68°C.) except that, in the latter case, no difference in flavour was detectable between beers from long and short boiled worts. Stirring can replace vigorous boiling to secure normal hop-utilization and shelf-life but simmered worts give beers of characteristic flavour probably as a result of enhanced contents of hop oil. Use of a lightly kilned malt of high nitrogen content (1·8%) produced a characteristically different flavour from that obtained with pale-ale malt and reduced the utilization of hop substances by provoking a greater loss of this material during fermentation.     

CHANGES IN THE CONTENTS OP BARLEY PROTEOLYTIC INHIBITORS DURING MALTING AND MASHING

Barley contains three types of large-molecular proteolytic inhibitors, viz., inhibitors active against malt endopeptidases, a specific trypsin inhibitor, and a group of inhibitors of microbial proteinases and chymotrypsin. The inhibitors of malt endopeptidases disappear at an early stage of germination, just before the endopeptidase activity begins to increase. The total amount of these inhibitors is so small, however, that the great increase in endopeptidase activity occurring during germination cannot be due to the breakdown of an “equivalent” amount of inhibitors. The contents of the other two types of inhibitor in barley are much higher; they decrease slowly during germination, but high activities are still present in malt. The inhibitors of microbial proteinases are progressively inactivated during mashing. The trypsin inhibitor retains full activity through mashing, but is completely inactivated during boiling and hopping.     

THE INFLUENCE OF MASHING CONDITIONS ON WORT AND BEER QUALITY

From a comparison of the performance of ale and lager malts mashed by constant temperature infusion, by temperature programmed infusion, and by decoction procedures it emerges that malt modification and the pattern of kilning are as important in determining the composition of wort as are the mashing conditions. Worts with appropriate levels of amino and total nitrogen and of fermentable sugars may be obtained by selecting lightly-dried well-modified malts and mashing with a constant temperature infusion rather than by using less modified malt in conjunction with more complex mashing programmes. Ale malts yield worts of lower pH which is reflected in a slight reduction in hop utilization. Head retention is improved by the use of undermodified malts but colloidal stability is improved when well-modified malts are used. In the case of well-modified malts a high final curing temperature is not a prerequisite for achieving good colloidal stability in beer.     

TRACE VOLATILE CONSTITUENTS OF BEER DIMETHYL SULPHIDE FORMATION IN MALT

Dimethyl sulphide formation in malt occurs primarily during the kilning process, and is significantly dependent upon the maximum kilning temperature. During mashing a considerable fraction of the dimethyl sulphide of malt is transferred to the wort. Since this is not always completely removed by the wort boiling process, the remaining dimethyl sulphide, although decreased during fermentation, may be sufficient to impart a noticeable dimethyl sulphide aroma to the finished beer.     

糖化车间节能技术探讨

我们知道,糖化车间的能量消耗约占啤酒厂总能量的50%。如何降低糖化车间的能耗,目前已经成为各啤酒厂的中心议题。本文主要从追加热水、回收乏汽、采用新型煮沸技术、麦汁一段冷却等四个方面对节能技术作了探讨。