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http://www.biokemi.org/biozoom/issues/522/articles/2368
Forside
BioZoom
2008
Nummer 2
Enzymes in brewing
Enzymes in brewing
Publiceret April 2008 Even for an old industry like beer brewing new industrial processes benefit from using enzymes developed from microbial sources. In the last years quality issues like flavour control, beer stability and general cost savings in the industry go hand in hand with efficient solutions of environmental problems. Future aspects focus on a wider application of enzymes to brew with high amounts of inexpensive raw materials like barley. Alternative beer processes for production of wort and beer with higher productivity and reduced amounts of waste and by-products are under development.
Introduction
Beer and wine are both alcoholic beverages which have been part of our social life for thousands of years. Both beverages are produced by yeast fermentation of sugars. Wine is based on grapes, and beer is traditionally based on barley. The matured grapes already contain the sugars needed for the fermentation, while barley contain starch that has to be broken down to fermentable sugars before the yeast can make alcohol. Therefore, traditional brewing contains and extra step compared with wine-making, namely malting in which enzymes needed for the degradation of starch into fermentable sugars are produced.
Figure 1: Germinating barley kernel Malt is germinated barley or other cereals like wheat and sorghum: First the grains are "steeped" bringing the water content from about 12% to 45%, then they are allowed to germinate for 4-6 days and finally the germination is stopped by heating (kilning) reaching a final moisture content of about 4%. Some enzymes are already present in the barley, e.g. -amylases, but the majority of enzymes are produced during the germination, e.g. -amylases and proteases, and in the final malt all the enzymes needed for the conversion of "grains" into a fermentable liquid (wort) is present (Figure 1 and 2)
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Figure 2. Enzyme production during malting, ref. Aastrup et al. (2004) In former days, production of malt was an integrated part of every brewery, but to day most malt is produced outside the brewery in large malt factories, and malt has become a purchased raw material, like other raw materials. This means that the breweries to day are more flexible in the use raw materials, and for that matter for the source of enzymes. The malt enzymes do have some limitations. They can only work at certain temperatures, pH values etc., and the activities might be too low to do a proper job in proper time. In contrast, commercial exogenous enzymes can be designed to work at preferred temperatures and pH values, to have more enzymatic power, or to express wanted enzyme activities that are not present in malt. Addition of exogenous enzymes at various steps during the brewing process can therefore make brewing easier, faster and more consistent. It gives the brewmasters extra flexibility in the choice of raw materials due to less dependence on malt enzymes, as well as providing opportunity to create new products, which is not possible to make with malt enzymes alone. Also the possibility to improve beer quality by avoiding off-flavours is possible with commercial enzymes. The increasing concern on resources and CO2- emission has also put the use of commercial enzymes within the brewing industry in focus. By the use of exogenous enzymes more can be extracted from the raw materials, more local raw materials can be used, and more unmalted grains can be used, saving significant amounts of energy and transport.
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Figure 3: The traditional mashing temperature profile is determined by the temperature optima for the various malt enzymes. Larger version If -glucan and protein are properly broken down during malting, single temperature mashing at 65-71C has shown to be sufficient, as in the case of traditional ale brewing. During mashing the starch is degraded to dextrin and fermentable sugars. -amylase liquefy the gelatinized starch by hydrolysis of the -1,4 linkages at random. -amylases are exo-enzymes which attack the liquefied starch chains resulting in successive removal of maltose units from the non-reducing end. After mashing, the mash is sieved in a lauter tun or on a mash filter. The resulting liquid, known as sweet wort, is then transferred to the copper, where it is boiled with hops. The hopped wort is cooled and transferred to the fermentation vessels, where yeast is added. In normal wort 2/3 of the carbohydrates are fermentable sugars. After fermentation, the so-called green beer' is matured before final filtration and bottling. Fig. 4 shows a diagram of the brewing process and where external enzymes are used for process aids.
Figure 4: The processing steps in brewing where exogenous enzymes can be added. Larger version.
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the beer filtration rate will be reduced; and the flavour and stability of the beer will be inferior. Exogenous enzymes are used to supplement the malt's own enzymes in order to prevent these problems. Furthermore, industrial enzymes are used to ensure better adjunct liquefaction, to produce low-carbohydrate beer (light beer'), to shorten the beer maturation time, and to produce beer from cheaper raw materials. The various steps of the brewing operations, where microbial enzymes are occasionally added, are shown in table 1. Enzymes, enzymic action and their functions are summarized.
Enzymes at work
Quality and supply constraints on malt, and doubling of malt prices have given increased interest for enzyme solutions in 2007 and 2008. Many breweries has run programs within the last two years in order to increase efficiency and optimize raw material usage, and many of them have focused on commercial enzymes to shorten the production time, increase capacity, and to allow use of raw material alternative to malt. Three important examples are mentioned: Exchanging part of the malt with barley has been popular because using barley in combination with commercial enzymes gives the same beer quality as with malt. Introducing a higher content of starch hydrolysing enzymes offer the possibilities of producing "light beer" also called "low calorie beer". An enzyme solution for diacetyl control after fermentation improves vessel utilization, save energy and ensures a high beer quality after a reduced maturation time.
Operation
Enzymes
Enzyme action
Function
Hydrolyse starch
Hydrolyse glucans.
-amylase
Hydrolyse starch.
Malt improvement.
Amyloglucosidase
Mashing Debranching enzyme Hydrolyse -1,6 branch points of starch. Secures maximum fermentability of the wort.
Proteases
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(FAN).
-glucanase
Hydrolyse glucans.
Pentosanase/xylanase
Fungal -amylase
Fermentation
-glucanase
Hydrolyze glucans.
-acetolactatedecarboxylase (ALDC)
* Adjunct is starchy cereals such as maize, rice, wheat, sorghum, barley or pure starch materials added to the mash.
Table 1. Steps of the brewing operations where microbial enzymes are used.
Raw materials:
Malt
475 kg
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TermamylBrewQ is an enzyme preparation containing a thermophilic -amylase. CeremixPlus is an enzyme preparation containing -glucanase, xylanase, -amylase and protease UltrafloMax is an enzyme preparation containing -glucanase and arabinoxylanase.
Figure 5. Mashing diagram for barley brewing (an example). The mashing diagram is shown in figure 5. The maize grits are liquefied separately with help of the -amylase TermamylBrewQ at 96C for 30 minutes, through a short holding time at 70C. It is stabilised by approximately 100 ppm Ca++ at a water-to-adjunct ratio of approximately 4:1. Milled malt and barley are mashed-in at a temperature of 50C. After 30 minutes the adjunct mash from the decoction vessel is added to increase the temperature to 63-66C. After 60 minutes the mash is heated to hold at 76-78C until starch-negative (no blue colour is formed with iodine in potassium iodide). Hereafter the wort separation is made in the lauter tun. Brewing with high amounts (>50%) of barley instead of malt is now possible thanks to the introduction of the new enzyme system UltrafloMax (2).
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Figure 6. Total fermentable sugar production with different dosages of Attenuzyme (kg per ton malt) and extended mashing at 63C Beer types with very high attenuation ("light beer" or "low calorie beer") are most often produced using amyloglucosidase alone. Extended mashing at 63C and high dosages of enzymes is necessary to produce extremely high attenuated beer (see figure 6). Fungal -amylases are used to produce mainly maltose and dextrins whereas amyloglucosidase produces glucose from both linear and branched dextrins.
Diacetyl control
An important question for brewers is "When exactly is a beer mature?", because this determines when they can "rack" the beer to make way for the next batch. The simple answer to the above question is when the diacetyl level drops below a certain limit (about 0.07 ppm). Diacetyl gives beer an off-flavour like buttermilk and one of the main reasons for maturing a beer is to allow the diacetyl to drop to a level where it can't be tasted. Diacetyl is formed by the non-enzymatic oxidative decarboxylation of -acetolactate, which is produced by the yeast during primary fermentation. The yeast removes the diacetyl again during the beer maturation stage by conversion to acetoin, which has a much higher flavour threshold value. In fact, acetoin is almost tasteless compared with diacetyl. By adding the enzyme -acetolactate decarboxylase (ALDC) (e.g. Novozymes' Maturex) at the beginning of the primary fermentation process, it is possible to bypass the diacetyl step (Figure 7) and convert -acetolactate directly into acetoin. Most of the -acetolactate is degraded before it has a chance to oxidise and less diacetyl is therefore formed. This makes it possible to shorten or completely eliminate the maturation period (3) and (4). The brewery enjoys greater fermentation and maturation capacity without investing in new equipment.
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shorter cooking cycles, no risk of residual starch being carried over into the mashing vessel and more flexibility to change adjunct ratios and types.
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Better waste water control: Water and wastewater management constitutes a practical problem for the brewing industry, and exogenous enzymes can play a significant role in waste water treatment. An overview of significant improvement and operation processes and economic reality was described by Fillaudeau et al. (6). ...or something completely different!
References
(1) Aastrup, Sten, Noel Bautista, Elmar Janser and Kurt Drreich. "Choice of enzyme solution should determine choice of raw materials and process". Presentation given at World Brewing Conference, San Diego, USA, 2004 (2) Aastrup, Sten, Claudio Visigalli, Jrg Obrecht, Marcel Mischler, Niels Elvig, Stefan Kreisz, "Rethink beer filtration with new xylanase", Paper submitted for publication in Brauwelt (2008) - special filtration issue. (3) Aunstrup, K. and Olsen, F. Alpha-acetolactate decarboxylase enzyme and preparation thereof. U.S.Patent 4,617,273. (1986) (4) Rostgaard Jensen, B., Svendsen, I., Ottesen, M. Isolation and characterization of -acetolactate decarboxylase useful for accelerated beer maturation. Proceedings of the European Brewery Convention Congress, p. 393-400. (1987). (5) Olsen, H. S. and Bisgaard-Franzen, H. Beer mashing process", WO 2004/050820 A1. (6) Fillaudeau, L, Pascal Blanpain-Avet, Georges Daufin, Water, wastewater and waste management in brewing industries", Journal of Cleaner Production 14 (2006) 463-471.
Forfatter
Steen Aastrup Novozymes Hans Sejr Olsen Novozymes hso@novozymes.com
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