WO2002014461A2

WO2002014461A2 - The brewing of an alcoholic beverage

Info

Publication number: WO2002014461A2
Application number: PCT/IB2001/001417
Authority: WO
Inventors: Barry Clifford Axcell; Brett Ziehl; Sandra Nadine Egbe Van Nierop
Original assignee: The South African Breweries Limited
Priority date: 2000-08-11
Filing date: 2001-08-08
Publication date: 2002-02-21
Also published as: AU2001276602A1; WO2002014461A3

Abstract

A process for brewing or producing a beer includes obtaining a wort from a grain mash comprising grain which includes peroxidase, the wort thus including peroxidase extracted from the grain in the mash. The wort is maintained at a temperature of no greater than 65° C, and then fermented, at a temperature no greater than 25° C, to obtain a beer which includes peroxidase.

Description

THE BREWING OF AN ALCOHOLIC BEVERAGE

THIS INVENTION relates to the brewing of an alcoholic beverage. In particular, it relates to a process for brewing a beer, to a process for brewing an alcoholic beverage, and to an alcoholic beverage.

According to a first aspect of the invention, there is provided a process for brewing a beer, the process including obtaining a wort from a grain mash comprising grain which includes peroxidase, the wort including peroxidase extracted from the grain in the mash and the wort being maintained at a temperature of no greater than 65°C; and fermenting the wort, at a temperature no greater than 25 °C, to obtain a beer which includes peroxidase.

The grain may be barley, or other grain which includes peroxidase. The beer may be a lager beer.

Preferably, the grain mash from which the wort is obtained, is subjected to a heat treatment step. The heat treatment step may include maintaining the grain mash at a temperature of between about 63°C and about 65 °C, e.g. 64°C, for a period of at least 1 1 0 minutes, typically between about 1 10 minutes and about 1 50 minutes, e.g . 1 20 minutes. The grain mash may be agitated during this time to improve mixing and heat transfer. During this time, starch extracted from the grain is gradually hydrolysed into fermentable sugars and smaller dextrins, and the peroxidase is extracted from the grain. Preferably, the grain mash is starch negative to iodine testing, before further processing. The process typically includes mixing crushed grain, e.g. malted barley, and brewing water together to prepare the grain mash from which the wort is obtained . The brewing water and crushed grain or grist may be mixed in a ratio of about 3,2 hectoliter to about 4,2 hectoliter of brewing water per 1 00 kg of grist. The brewing water may have a pH of between about 5 and about 6 and a temperature of between about 50°C and about 60°C, thus typically resulting in the grain mash having a temperature of between 50°C and 60°C, e.g. about 55 °C. Typically, for the heat treatment step, the temperature of the grain mash is then raised, e.g. by using steam, to reach the preferred temperature of between 63°C and 65 °C. The temperature of the mash may be raised at a rate of between 0.8°C/minute and 1 °C/minute, e.g.

1 °C/minute.

Obtaining the wort from the grain mash typically includes straining the wort from insoluble grain and husk material forming part of the grain mash. Brewing water, preferably at about the same temperature as the grain mash, may be used to sparge the grain mash, and forms part of the wort. The brewing water used to sparge the grain mash may have a pH of between about 5 and about 9 and a temperature of between about 65°C and about 67°C, e.g. about 65 °C.

Preferably, the wort so obtained has a density of at least 1 1 ,8°P, e.g . between about 1 1 ,8°P and about 1 2,5°P, ensuring that the wort is not over-diluted, i.e. that the wort has a peroxidase concentration as high as is practically possible and economically feasible.

The process may include adding hops to the wort to obtain hopped wort, without calcium sulphate being added to the hopped wort and without boiling the hopped wort, thus maintaining the temperature of the hopped wort at no greater than 65°C. Preferably, the hops is pre-isomerised. Contrary to conventional beer brewing processes, there is thus no addition of calcium sulphate to the hopped wort. However, other easily dissolving brewing salts, e.g. calcium chloride, may be added. If necessary, to await for capacity in downstream vessels, the hopped wort is kept at a temperature of about 60°C to about 65 °C. Thus, the hopped wort is unboiled. The unboiled hopped wort may have a density of between about 1 0°P and about 1 8°P, preferably between 1 1 ,8°P and 1 3°P, most preferably between 1 1 ,8°P and 1 2,2°P.

The unboiled hopped wort may have a pH of between about 5, 1 and about 5,5, e.g. between 5,2 and 5,4. If it is desired to reduce the effect of amylases, the pH may be even lowered to between about 4 and 4,4. The pH may be lowered by adding a suitable acid, e.g. lactic acid (80%) or phosphoric acid, to the hopped wort.

If hops is added to the wort, the process typically includes separating trub from the unboiled hopped wort, whilst preferably still maintaining the unboiled hopped wort at a temperature of 60°C to 65 °C.

Prior to fermenting the wort, the wort may be cooled and, if desired, oxygenated. Typically, the wort is cooled to a temperature of about 8°C to about

1 4°C, preferably about 9°C to about 1 3°C, e.g. 1 0°C. The oxygenated cold wort may have a dissolved oxygen concentration of about 1 0 ppm to about 20 ppm, typically about 1 2 ppm to about 1 8 ppm, e.g . about 1 6 ppm to about 1 8 ppm.

Typically, fermenting the wort includes pitching the wort with a yeast slurry comprising fresh yeast, i.e. not yeast that has been previously used for the process of the invention, although this possibility is not excluded. Preferably, the yeast is healthy and is removed promptly after fermentation. The wort may be pitched at a ratio of between about 0,8 kg yeast per hectoliter of wort and about 1 ,2 kg yeast per hectoliter of wort, e.g . 1 kg yeast per hectoliter of wort.

During fermentation, the temperature of the wort may be allowed to rise to a temperature no greater than 25 °C. The temperature of the wort may thus rise to about 1 2°C to about 1 8°C, preferably about 1 3°C to about 1 7°C, e.g.1 5°C, as a result of the heat released by the fermentation process. Typically, the wort is rapidly fermented for a period of about 5 days to about 7 days, in order to obtain maximum peroxidase activity in the fermenting wort. After fermentation of the wort for the desired period, the resulting beer, which is a "green" beer and which includes peroxidase, is typically chilled back to a temperature of about 3 °C to about 5 °C, e.g. about 4°C. The green beer may be chilled back at a rate of about 0, 1 5 °C/h to about 0,35 °C/h, typically about 0,2°C/h to about 0,3°C/h. Preferably, the green beer is chilled back when it reaches a diacetyl concentration of less than 60 ppb, more preferably less than 50 ppb. As mentioned hereinbefore, this is typically after about 5 days to 7 days of fermentation. Carbon dioxide may be collected for purification.

After fermentation, the green beer may be centrifuged to remove the yeast and traces of trub, before the green beer is matured. Preferably, the yeast count should be reduced to 0,5 X 1 0⁶ cells/ml. The green beer may be matured for a period of about 1 day to about 7 days, at a temperature of between about -2°C and about

0°C, e.g. -1 ,5 °C.

The process may include adding silica hydrogel to the centrifuged green beer to increase colloidal stability of the beer.

The process may also include adding Kieselguhr to the green beer, and filtering the green beer through a diatomaceous earth filter, rendering the beer bright.

The process may further include carbonating the bright beer, e.g . by adding purified CO₂ gas, typically at a temperature of between about -1 °C and about 0°C.

It is to be appreciated that the activity of the peroxidase is negatively affected by excessive heat and the use of stabilisers to remove proteinaceous material from the beer. Excessive pasteurisation also causes a reduction in peroxidase activity. Furthermore, certain types of silica hydrogel, such as PCSX, Lucilite and PC5, also adsorb peroxidase and remove it from the beer during filtration. The addition of proteolytic enzymes to the beer may also reduce the activity of the peroxidase. The process may include blending the beer which includes peroxidase with another beer to produce a beer having a peroxidase concentration of at least 0,04 U/ml.

According to a second aspect of the invention, in a process for brewing an alcoholic beverage from raw materials and additives and which process comprises obtaining a wort from a grain mash, subjecting the wort to a heat treatment step, and converting the heat treated wort into the alcoholic beverage, there is provided the improvement whereby, at any stage in the process after the heat treatment of the wort, an enzyme is added as an additive in order to inhibit the presence of hydroxyl radicals in the alcoholic beverage obtained.

The alcoholic beverage may be a beer, such as a lager beer, or an ale. Thus, typically, the grain of the mash is barley, and the wort is typically obtained from a malted barley mash.

The enzyme may be peroxidase.

The peroxidase may be obtained from a source selected from the group consisting of grain, fungi and bacteria. Instead, the peroxidase may be synthesized .

In one embodiment of the invention, the peroxidase is added in the form of an intermediate peroxidase-containing product of the process in accordance with the first aspect of the invention.

In another embodiment of the invention, the peroxidase is obtained from at least a portion of the grain mash from which the wort is obtained. The process may thus include separating the grain mash into a primary mash portion and a secondary mash portion with the primary mash portion providing the wort which is subjected to the heat treatment step in order to obtain the alcoholic beverage, and the secondary mash portion being treated to provide a peroxidase-containing additive. The volumetric ratio of the secondary mash portion to the primary mash portion may be between about 0, 1 : 1 and about 0,3: 1 , e.g. 0,2: 1 .

The grain mash may be prepared as hereinbefore described, and the heat treatment step to which the primary mash portion is subjected may be as hereinbefore described .

A liquid phase, hereinafter referred to as a green wort, may be separated from an insoluble phase of the secondary mash portion. As will be appreciated, the green wort includes some of the peroxidase extracted from the grain in the grain mash.

The peroxidase may thus be added in the brewing process in the form of the green wort, which thus provides a peroxidase- containing additive.

The insoluble portion may be re-suspended in hot water (65°C to 72°C), and returned or recycled to the primary mash portion, in order to maximise the amount of extract recovered from the grain.

The primary grain mash portion may be further treated in conventional manner, apart from the addition of the enzyme at some stage in the process, in order to produce an alcoholic beverage. Typically, for a lager beer, the treatment of the primary mash portion includes completing conversion of starch extracted from the grain into fermentable sugars and smaller dextrins, separating wort and spent grain, sparging the spent grain with brewing water to extract as much sugar as possible and adding the brewing water used as sparging water to the wort, boiling the wort with hops, separating trub from the boiled hopped wort, cooling the hopped wort and oxygenating the hopped wort to provide an oxygenated cold wort, pitching the oxygenated cold wort with a yeast slurry and fermenting the pitched wort to provide a green beer, cooling the green beer, centrifuging the green beer to remove traces of trub and yeast, maturing the green beer, adding Kieselguhr to the green beer and filtering the green beer with a diatomaceous earth filter, and adding C0₂ to the green beer to provide a bright beer. As will be appreciated, the peroxidase, contained in the green wort, or in another form, can be added as an additive at a number of points in or between stages of the further treatment of the primary grain mash. Examples of addition points where the peroxidase can be added are at the cooling of the hopped wort, during fermentation of the pitched wort, immediately prior to centrifuging the green beer, immediately after maturing the green beer, and into the bright beer.

The process may include treating the green wort in similar, though not identical fashion, to the treatment of the wort obtained from the primary mash portion. The green wort may be treated in a manner described hereinbefore for the treatment of wort according to the first aspect of the invention.

The invention extends to an alcoholic beverage which includes peroxidase.

The beverage may have a peroxidase-concentration of at least 0,04 U/ml, preferably between about 0,04 U/ml and about 0,06 U/ml e.g. 0,05 U/ml.

The alcoholic beverage may be a beer, e.g. a lager beer, or an ale.

The invention will now be described, by way of example, with reference to the accompanying diagrammatic drawings, in which

Figure 1 shows a process in accordance with the invention for brewing a peroxidase-containing beer; and Figure 2 shows another process in accordance with the invention for brewing a beer.

Referring to Figure 1 of the drawings, reference numeral 1 0 generally indicates a process in accordance with the invention for brewing a peroxidase- containing beer. The process 1 0 comprises, connected in series by a flow line, a mashing tun or vessel 1 2, a lauter tun or mash filter 1 6, an underback 1 8, a wort kettle 20, a whirlpool 24, a wort cooler 28, an oxygenator 32, a yeast pitching stage 36, a fermenter 40, a beer cooler 46, a beer centrifuge 48, a storage and maturation vessel 52, a buffer tank 56, a diatomaceous earth filter 60, and a bright beer tank 64.

The process 1 0 further includes a mash feed line 1 4 leading into the mashing vessel 1 2, a brewing water feed line 1 7 leading into the lauter tun 1 6, a hops feed line 22 leading into the wort kettle 20, a trub removal line 26 leading from the whirlpool 24, a cooling water line 30 passing through the wort cooler 28, an oxygen feed line 34 leading into the oxygenator 32, a yeast slurry feed line 38 leading into the yeast pitching stage 36, a CO₂ and volatiles removal line 42, and a solids removal line 44, leading from the fermenter 40, a solids removal line 50 leading from the beer centrifuge 48, a silica hydrogel feed line 54, a Kieselguhr feed line 58, and a carbonated, deaerated water feed line 62.

The process 1 0 is used to produce or brew a lager beer which includes peroxidase. Crushed barley malt is mixed with brewing water in a malt mill or premasher (not shown) in conventional manner, before being fed to the mashing vessel 1 2 along the mash feed line 1 4. Thus, typically, the barley malt is milled for about 30 minutes, using a roller mill with roller gaps of between about 0.3 mm and 1 .2 mm. If desired, the malt may be a mixture of black malt and pale malt, e.g . a mixture containing 0.4 kg of black malt per 1 00 kg of pale malt. If available, the malt is preferably Pilsen malt, since Pilsen malt has been found to give high peroxidase concentrations in the beer. Malting conditions that preserve enzymes, such as long gentle kilning at lower temperatures, are preferred for the barley malt. The brewing water and crushed barley malt are mixed in a ratio of 4 hectoliters of brewing water per 1 00 kilograms of grit to provide a mash. The mash has a target calcium content of 80 mg per liter. The brewing water has a pH of about 5,5 and a temperature of about 60°C, providing the mash at a temperature of about 55 °C.

In the mashing vessel 1 2, it is intended that as much peroxidase as possible is extracted from the mash. The mash is heated in a controlled way with steam to a temperature of 64°C, and maintained at this temperature for about 1 20 minutes. It is imperative that the temperature of the mash does not exceed 65 °C in the mashing vessel 1 2. It has been found that peroxidase is stable at 65°C for up to 5 hours, but that its activity declines rapidly above 65°C. The mash temperature is raised at a rate of about 1 °C per minute from 55 °C to 64°C. The mash should have a pH of about 5,5, and lactic or phosphoric acid and/or a salt such as calcium chloride or calcium sulphate is used to correct the pH of the mash. Preferably, the mash is starch negative when tested with fresh, cold iodine.

No attempt is made during mashing to control fermentability, since wort which will be obtained from the mash in the mashing vessel 1 2 will contain sufficient

^-amylase to allow fermentation to go to completion. In the mashing vessel 1 2, starch extracted from the barley in the mash is gradually hydrolyzed into fermentable sugars and smaller dextrins, and peroxidase is also extracted from the barley.

From the mashing vessel 1 2, the mash is transferred at a transfer velocity of about two meters per second to the lauter tun 1 6 where wort is obtained from the mash by straining the wort from the insoluble barley and husk material forming part of the mash. The lauter tun 1 6 is loaded with the mash at a loading of about 240 kg of mash per m² of the lauter tun 1 6. Brewing water is used as sparging water in the lauter tun 1 6 and is fed into the lauter tun 1 6 along the brewing water feed line 1 7. The brewing water fed to the lauter tun 1 6 should have an alkalinity, expressed as milligrams CaCO₃ per liter, of 25, a pH of about 5,5 and a temperature of about 65°C, but no greater than 65°C. The wort obtained from the lauter tun 1 6 should be as bright as possible. The amount of sparging water used is restricted so as not to over- dilute the wort, i.e. to provide a wort with a peroxidase concentration as high as possible. It is important not to run the wort off too fast, particularly in the final stages of lautering, to allow the peroxidase to diffuse into the sparging water.

The wort and sparging water from the lauter tun 1 6 are collected in the underback 1 8, together forming a green wort. The gravity of the wort collected in the underback 1 8 should be kept above 1 1 ,8°P. From the underback 1 8 the green wort is transferred to the wort kettle 20. In the wort kettle 20, hops is added to the green wort. The temperature of the green wort should be maintained above 60°C . Preferably, this is not achieved by heating, but by minimizing heat losses from the wort kettle 20. No calcium sulphate is added to the green wort in the wort kettle 20, since there is no boiling of the green wort in the wort kettle 20 which would allow the calcium sulphate to adequately dissolve and mix into the green wort. Pre-isomerized hops is used for its bacteriostatic effect and to add bitterness to the beer. The hopped wort in the wort kettle 20 should have a pH of about 5,3 and a gravity of about 1 2°P as well as a colour and bitterness in accordance with specifications, before the hopped wort is transferred to the whirlpool 24.

The whirlpool 24 is used to separate any solid material, i.e. trub from the hopped wort. The trub is removed by means of the trub removal line 26. The hopped wort is allowed to stand in the whirlpool 24 for a period of about 20 minutes before it is transferred to the wort cooler 28, where the hopped wort is cooled to a temperature of about 1 6^CC by means of cooling water flowing through the wort cooler 28 along the cooling water line 30. The cooling water entering the wort cooler 28 has a temperature of about 3°C to 5 °C, and exits at a temperature of about 55°C to 60°C.

Cooling of the hopped wort is effected in less than 75 minutes.

The cooled wort is passed through the oxygenator 32, where oxygen is added along the oxygen feed line 34 to oxygenate the wort to a dissolved oxygen concentration of about 1 6 ppm to 1 8 ppm. The oxygenated wort is transferred from the oxygenator 32 to the yeast pitching stage 36 where it is pitched with fresh yeast slurry fed along the yeast slurry feed line 38. The yeast should be healthy and is pitched at a rate of 1 kilogram per hectoliter of oxygenated wort, at 60% consistency by mass.

The pitched wort is fermented in the fermenter 40 to produce green beer. Fermentation occurs rapidly at a temperature of about 1 3°C to about 1 7°C over a period of about 5 to 7 days. The fermenter 40 is operated at a back-pressure of less than about 5 kPa at a fermentation rate of 3,5 °P per day. The alcohol concentration of the beer produced in the fermenter 40 is about 6% by volume. CO₂ and other volatiles are removed from the fermenter 40 by means of the CO₂ and volatiles removal line 42, whereas yeast and trub are removed from the fermenter 40 by means of the solids removal line 44. The yeast is removed promptly at the end of fermentation.

After fermentation, the green beer is cooled in the fermenter 40 at a rate of about 0.25 °C per hour to a temperature of about 4°C. The green beer is cooled when it reaches a diacetyl concentration of less than 50 ppb. Thus, the process 1 0 strives to have a fermented green beer with a diacetyl concentration of less than 50 ppb, by the time the peroxidase activity peaks, which is usually around 5 days.

The green beer is transferred from the fermenter 40 to the beer cooler 46 to further reduce the beer temperature to about -1 ,5°C. From the beer cooler 46, the green beer is transferred to the beer centrifuge 48 for removing any remaining yeast via the solids removal line 50, and then to the storage and maturation vessel 52. The green beer is typically kept in the storage and maturation vessel 52 for a period of between 1 and 7 days, at a temperature of about -1 ,5 °C. Silica hydrogel is then added to the green beer by means of the silica hydrogel feed line 54, in order to increase colloidal stability of the beer whereafter the green beer is fed into the buffer tank 56. It passes through the buffer tank 56 with a residence time of about 1 5 minutes.

From the buffer tank 56, the green beer is passed through the diatomaceous earth filter 60 to render the bear bright. Before entering the diatomaceous earth filter 60, Kieselguhr is added to the green beer by means of the Kieselguhr feed line 58.

Carbonated, deaerated water is fed into the bright beer exiting the diatomaceous earth filter, before the bright beer is fed into the bright beer tank 64. The beer is carbonated to a CO₂-level of about 2,4 volumes CO₂/volume of beer and should not be kept in the bright beer tank 64 for more than about four days. In total, the process for producing the beer is completed in less than 1 8 days. The peroxidase-containing beer produced in accordance with the process 1 0 is blended into a conventional beer at a concentration of about 5 % to 1 0% by volume. Blending may be achieved in a storage vessel, or in-line during racking of the conventional beer. The objective of blending is to blend as little as possible of the peroxidase-containing beer into the conventional beer while still achieving a peroxidase concentration of about 0,05 U/ml. The blended beer may then be matured and processed in conventional fashion.

It should be appreciated that the process 1 0, as described above, may be interrupted at one of many points or stages during the process, with the peroxidase- containing material present at that stage in the process being used as an additive in a process for brewing beer, with the purpose of inhibiting the presence of hydroxyl radicals in the beer to which it is added. Examples of such removal points or interruption points are the hopped wort obtained from the whirlpool 24 after the cooler 28, the green beer produced by the fermenter 40, the green beer from the storage and maturation vessel 32, and the bright beer from the diatomaceous earth filter 60.

Referring to Figure 2 of the drawings, reference numeral 1 00 generally indicates another process in accordance with the invention for brewing a beer. The process 1 00 includes, in a first leg, many of the features of the process 1 0, and the same reference numerals are thus used to indicate the same or similar features forming part of the first leg. However, in addition, the process 1 00 includes, also connected in series by a flow line and forming a second leg, a lauter tun 1 1 6, and underback 1 1 8, a wort kettle 1 20, a whirlpool 1 24, a wort cooler 1 28, an oxygenator 1 32, a yeast pitching stage 1 36, a fermenter 1 40, a beer cooler 1 46, a beer centrifuge 1 48, a storage and maturation vessel 1 52, a buffer tank 1 56, a diatomaceous earth filter 1 60 and a bright beer tank 1 64.

The process 1 00 further includes a hops feed line 1 22 leading into the wort kettle 1 20, a cooling water line 1 30 passing through the wort cooler 1 28, an oxygen feed line 1 34 leading into the oxygenator 1 32, a yeast slurry line 1 38 leading into the yeast pitching stage 1 36, a CO₂ and volatiles removal line 142, and a solids removal line 1 44, leading from the fermenter 1 40, a solids removal line 1 50 leading from the beer centrifuge 1 48, a silica hydrogel feed line 1 54, a Kieselguhr feed line 1 58, and a carbonated, deaerated water feed line 1 62.

The process 1 00 is also used to produce or brew a lager beer which includes peroxidase. A mash is prepared in the same fashion as described above for the process 1 0, with mashing taking place in the mashing vessel 1 2 at a temperature of less than 65 °C, e.g . 64°C. However, about 3,6 hectoliters of brewing water is used per 1 00 kg of barley malt. The conditions in the mashing vessel 1 2 of the process 1 00 are the same as the conditions in the mashing vessel 1 2 of the process 1 0.

From the mashing vessel 1 2, after about 45 minutes, a 1 6% by volume secondary portion of the mash is transferred by means of a flow line 1 02 to the lauter tun 1 1 6, where wort is obtained from the secondary mash portion by straining the wort from the insoluble barley and husk material forming part of the secondary mash portion. The wort obtained from the lauter tun 1 1 6 is collected in the under back 1 1 8

^■as a green wort. Grain remaining in the lauter tun 1 1 6 is returned by a flow line 1 04 to the mashing vessel 1 2 after having been resuspended in hot water (65 °C - 72°C) . The mash portion in the mashing vessel 1 2 forms a primary mash portion. After starch conversion in the mashing vessel 1 2, the primary mash portion in the mashing vessel 1 2 is transferred to the lauter tun 1 6, where wort is obtained from the primary mash portion and collected in the underback 1 8 as a green wort.

Between the underback 1 1 8 and the bright beer tank 1 64, the process 1 00, as far as the green wort obtained from the lauter tun 1 1 6 is concerned, is identical to the process 1 0 described above. However, since the secondary mash portion in the lauter tun 1 1 6 is not sparged, the gravity of the wort obtained from the lauter tun 1 1 6 is higher than the gravity of the wort obtained from the lauter tun 1 6 in the process 1 0, i.e. about 1 6°P. It is to be appreciated that the process 100, between the underback 1 18 and the bright beer tank 1 64 of the second leg, can be interrupted at any point or stage of the second leg with the peroxidase-containing material present at that stage in the second leg being used as an additive in the first leg of the process 1 00 between the beer cooler 28 and the bright beer tank 64. Examples of such interruptions in the first leg of the process 1 00, and additions to the second leg of the process 1 00, are indicated by the broken feed lines 1 80, 1 82, 1 84, 1 86 and 1 88. If it is desired to counter the effect of amylases preserved in the green wort of the second leg owing to the fact that the green wort in the second leg is not boiled, the wort in the first leg, after addition of the peroxidase-containing additive from the second leg, may be acidified to a pH of about 4. Acidification is however unnecessary if the peroxidase- containing additive is added to the first leg when the pH and temperature in the first leg is already low, e.g. when using the flow line 1 84.

The treatment of the green wort in the underback 1 8, apart from the possible addition of a peroxidase-containing additive from the second leg of the process 1 00, is conventional. Thus, operating conditions are conventional between the underback 1 8 and the bright beer tank 64. For example, in the wort kettle 20, the hopped wort is boiled, in contrast to the situation in the wort kettle 1 20. However, as soon as the peroxidase-containing additive from the second leg of the process 1 00 has been added to the first leg of the process 1 00, e.g . along one of the flow lines

1 80, 1 82, 1 84, 1 86 or 1 88, care must be taken to ensure that operating temperatures from then onwards along the first leg of the process 1 00 do not exceed 65 °C.

If the second leg of the process 1 00 is not interrupted, the bright beer in the bright beer tank 1 64 can be used to blend into the bright beer in the bright beer tank 64, at a concentration of about 5 % by volume, to produce a peroxidase- containing beer.

The following Example illustrates an assay method which can be used to determine peroxidase activity in beverages, such as beer. Example

Peroxidase Assay with Pyroqallol

Materials:

1 ml quartz cuvette(s) (zero on water) Spectrophotometer set at 31 8 nm wavelength

Micropipettes and tips Timer

Solutions:

0.1 M Acetate buffer pH4 Stock A:5.78 ml acetic acid (32%) /L

Stock B: 1 3.6 g CH₃COONa.3H₂O /L

Mix 41 ml A and 9 ml B, make up to 1 00 ml. Check pH.

This is the stock solution and must be stored at 4°C. It has a long shelf life if sterile filtered into a sterile bottle. Check before use that it is clear. For the assay mix 4 parts water and 5 parts stock. This solution has a shelf life of about 1 month (also check that this is still clear before use) .

Pyrogallol (254002 Aldrich)

0.1 g/ml dissolved in 0.01 M HCl. Rotate slowly at room temperature until dissolved. (0.01 M HCl; 86 ml 32% hydrochloric acid in 1 00 ml water) .

Aliquot into Eppendorfs, seal with parafilm and store at 4°C (keep in the dark) . It is important only to use this solution after it has been stored for 24 hours. It has a shelf life of up to 1 month.

H₂O₂ (UniLab Saarchem, 3063800 1 00 vols) 3 g H₂O₂ (30%) in 1 00 ml water.

Aliquot into Eppendorfs and store at 4°C.

Use each aliquot for one set of assays then discard. Peroxidase stock (Boehringer Mannheim, 41 3470)

0.0036 g Peroxidase ( ± 50 mg = 1 0000 U) made up in 1 00 ml water.

Take 5 ml and dilute into 50 ml.

Sample preparation: Fresh samples only, do not freeze, can measure samples stored at 4°C for up to 3 days. Make sure samples have not been exposed to filter aids. Samples can be filtered through 0.45 mm acetate filters but usually the sample is allowed to settle and then taken from the top. When yeast is present it should be removed by filtration or centrifuge (just enough to remove the yeast, 4500-rpm, 5 min)

Method:

Pipette the following directly into the quartz cuvette in the same order:

900 or 820* μl buffer

50 μl pyrogallol solution

20 or 1 00 * μl sample 30 μl H₂0₂ (to start the reaction)

*for samples with low peroxidase activity use larger sample volume i.e. 1 00 μl

1 . Invert cuvette twice to mix, insert into spectrophotometer and start timer while recording absorbance (31 8nm) .

2. Record absorbance at zero time and after 30 seconds. 3. Calculate change in absorbance over 1 minute and compare to a standard to get a value. 4. Use 20 μl sample for green wort and in process samples, use 100 μl sample for inpackage beer. Select volume of sample to ensure that the absorbance is less then 1 .5 during the assay. 5. All assays are done in duplicate, if these do not agree repeat again.

6. Calculation without using a standard:

1 . Average Abs_318nm per 30s X 2 = Abs_318nm./min.

2. Abs_318nm./min) / 5 = Abs_318nm/min./20 μl sample (100 μl assay)

3. Abs_318nm/min./20 μl sample X 1 .371 82 = U/ml peroxidase activity ( 1 .371 82 is the average factor calculated from using standard peroxidase - this varies a little )

When new solutions are made up compare the peroxidase activity of a sample using both old and new solutions to confirm that the assay is giving the same values (old pyrogallol solution may give slightly lower values but this is not more than a decrease of 1 0% over a month or more of storage, the new solution will also read lower after about a week.)

It is an advantage of the processes 1 0, 1 00 of the invention, as illustrated, that they use typical conventional brewing plant and equipment while producing a beer which includes peroxidase, leading to an improved flavour stability for the beer

Claims

CLAIMS:

1 . A process for brewing or producing a beer, the process including obtaining a wort from a grain mash comprising grain which includes peroxidase, the wort including peroxidase extracted from the grain in the mash and the wort being maintained at a temperature of no greater than 65 °C; and fermenting the wort, at a temperature no greater than 25 °C, to obtain a beer which includes peroxidase.

2. A process as claimed in claim 1 , in which the grain is barley and the beer is a lager beer.

3. A process as claimed in claim 1 or claim 2, in which the beer which includes peroxidase is blended with another beer to produce a beer having a peroxidase concentration of at least 0,04 U/ml.

4. A process as claimed in any one of the preceding claims, in which the grain mash from which the wort is obtained, is subjected to a heat treatment step.

5. A process as claimed in claim 4, in which the grain mash, during the heat treatment step, is maintained at a temperature of between 63°C and 65 °C for a period of at least 1 1 0 minutes.

6. A process as claimed in any one of the preceding claims, in which the wort has a density of at least 1 1 ,8°P.

7. A process as claimed in any one of the preceding claims, in which hops is added to the wort to obtain hopped wort, without calcium sulphate being added to the hopped wort and without boiling the hopped wort.

8. A process as claimed in claim 7, in which the unboiled hopped wort has a density of between 1 0°P and 1 8°P.

9. A process as claimed in claim 7 or claim 8, in which the unboiled hopped wort has a pH of between 5, 1 and 5.5.

1 0. A process as claimed in claim 7 or claim 8, which includes acidifying the unboiled hopped wort to a pH of between 4 and 4,4.

1 1 . A process as claimed in any one of the preceding claims, in which the wort is oxygenated to have a dissolved oxygen concentration of between 1 0 ppm and 20 ppm.

1 2. A process as claimed in any one of the preceding claims, in which the wort is fermented at a temperature no greater than 22°C, and in which the wort is fermented for a period of 5 to 7 days.

1 3. In a process for brewing an alcoholic beverage from raw materials and additives and which process comprises obtaining a wort from a grain mash, subjecting the wort to a heat treatment step, and converting the heat treated wort into the alcoholic beverage, there is provided the improvement whereby, at any stage in the process after the heat treatment of the wort, an enzyme is added as an additive in order to inhibit the presence of hydroxyl radicals in the alcoholic beverage obtained.

1 4. A process as claimed in claim 1 3, in which the alcoholic beverage is a beer or an ale.

1 5. A process as claimed in claim 1 3 or claim 1 4, in which the enzyme is peroxidase.

1 6. A process as claimed in claim 1 5, in which the peroxidase is obtained from a source selected from the group consisting of grain, fungi and bacteria.

1 7. A process as claimed in claim 1 5, in which the peroxidase is synthesized .

1 8. A process as claimed in claim 1 5, in which the peroxidase is added in the form of an intermediate peroxidase-containing product of the process as claimed in any one of claims 1 to 1 2 inclusive.

1 9. A process as claimed in claim 1 5, in which the peroxidase is obtained from at least a portion of the grain mash from which the wort is obtained.

20. A process as claimed in claim 1 9, which includes separating the grain mash into a primary mash portion and a secondary mash portion with the primary mash portion providing the wort which is subjected to the heat treatment step in order to obtain the alcoholic beverage, and the secondary mash portion being treated to provide a peroxidase-containing additive.

21 . A process as claimed in claim 20, in which the volumetric ratio of the secondary mash portion to the primary mash portion is between 0, 1 : 1 and 0,3: 1 .

22. A process as claimed in claim 20 or claim 21 , in which a liquid phase is separated from an insoluble phase of the secondary mash portion, with the liquid phase forming the peroxidase-containing additive, and the insoluble phase being recycled to the primary mash portion.

23. An alcoholic beverage which includes peroxidase.

24. An alcoholic beverage as claimed in claim 23, which is a beer or an ale.

25. An alcoholic beverage as claimed in claim 23 or claim 24, which has a peroxidase concentration of at least 0,04 U/ml.

26. A process for brewing or producing a beer as claimed in claim 1 , substantially as herein described and illustrated.

27. A process for brewing an alcoholic beverage as claimed in claim 1 3, substantially as herein described and illustrated.

28. An alcoholic beverage as claimed in claim 23, substantially as herein described and illustrated .

29. A new process for brewing or producing an alcoholic beverage or a beer, or a new alcoholic beverage, substantially as herein described.