Method for brewing beer

Abstract

The present application generally relates to the use of a non-conventional sourdough yeast strain as the dominant yeast strain in combination with lactic acid bacteria in a brewing process. In particular, the present application relates to a method of brewing beer wherein a wort is fermented with a Kazachstania yeast and a lactic acid bacteria; and / or a sourdough, wherein the sourdough comprises a non-Saccharomyces yeast species, particularly a Kazachstania yeast, as the dominant yeast species, and at least one lactic acid bacteria species.

Description

[0001] METHOD FOR BREWING BEER

[0002] Field of the invention

[0003] The present invention relates to the field of beer brewing. Specifically, the invention relates to a method of brewing beer comprising the fermentation of a wort with a combination of a non- Saccharomyces yeast, particularly a Kazachstania yeast, and lactic acid bacteria, particularly wherein said combination is in the form of a sourdough product.

[0004] Background of the invention

[0005] Traditionally, brewers have distinguished essentially two types of brewer’s yeasts, i.e. ale and lager yeasts, which are used to produce ale beers and lager beers, respectively. Both ale and lager yeasts belong to the genus Saccharomyces. Ale beer yeast or S. cerevisiae is a top fermenting yeast: during fermentation, the yeast grows on and / or migrates to the surface of the wort and can easily be collected from the surface for the next production. S. uvarum, S. eubayanus or S. pastorianus are lager beer yeasts. They have the main advantage to grow at much lower temperatures, ideal to produce lager beers and they sink to the bottom of the fermentation tank where they can be collected at the end of the process. Lager yeast is used to produce pilsner beer, which comprises the large majority of the beers produced in the world. Sour beers, such as Belgian sour beers like gueuze and lambic beers, are another type of beer, typically obtained by spontaneous fermentation. The most common yeast appearing in this type of beer belongs to the Brettanomyces genus (B. bruxellensis). These yeasts are well known for their resistance to acidity coming from a spontaneously developing population of acidifying bacteria in the beer.

[0006] Despite the growing interest in r\or\-Saccharomyces yeasts in food and beverage production, they are still largely overlooked in the beer brewing field. In addition, except for the above- mentioned sour beers, acidifying bacteria, such as lactic acid bacteria, are mostly considered as unwanted in beer brewing.

[0007] There is thus a need in the art for new methods for brewing a high quality, flavour-rich beer, based on a wort fermentation by a microflora that is not dominated by the traditional brewer’s yeasts.

[0008] Summary of the invention

[0009] The inventors have surprisingly found that it was possible to produce a high-quality beer with improved taste and aroma by performing the wort fermentation step in a beer brewing process with a combination of a Kazachstania yeast and lactic acid bacteria and / or with a sourdough comprising a r\or\-Saccharomyces yeast, particularly a Kazachstania yeast, as the dominant yeast and lactic acid bacteria. It was further found that the acidity levels of the obtained beer could be regulated during the fermentation process, resulting in a well-balanced, low alcoholic beer. Advantageously, the obtained beer is a full-bodied beer, despite the lower alcohol content. Due to the lower alcohol content, its calory content will be lower as well, compared to Saccharomyces based beers.

[0010] A first aspect of the present invention provides a method for brewing beer comprising the steps of a) preferably, providing or preparing a wort; and b) fermenting the wort with (i) a Kazachstania yeast and at least one lactic acid bacteria; and / or (ii) a sourdough, wherein the sourdough comprises a non- Saccharomyces yeast as the dominant yeast species, and at least one lactic acid bacteria, wherein the dominant yeast species is present in the sourdough in a concentration that is at least one log CFU / ml or CFU / g higher than any other yeast species, which is optionally present in the sourdough. In particular embodiments, the sourdough comprises a yeast belonging to the genus Kazachstania, Pichia, Torulaspora, Candida, or Lachancea as the dominant yeast species. In more particular embodiments, in step b), the wort is fermented with a sourdough, wherein the sourdough comprises a Kazachstania yeast as the dominant yeast species, and at least one lactic acid bacteria, wherein the dominant yeast species is present in the sourdough in a concentration that is at least one log CFU / ml or CFU / g higher than any other yeast species, which is optionally present in the sourdough. In certain embodiments, in case the sourdough comprises a Saccharomyces yeast, it is present in a concentration that is at least one log CFU / ml or CFU / g, particularly at least two, three or four log CFU / ml or CFU / g, lower than the dominant r\or\-Saccharomyces yeast species. In certain embodiments, the wort is fermented in the absence of a Saccharomyces and / or Brettanomyces yeast. In particular embodiments, the sourdough is an active liquid, paste or stiff, or dried sourdough.

[0011] In some embodiments, the Kazachstania yeast is Kazachstania bulderi, Kazachstania exigua, Kazachstania barnettii or Kazachstania unispora. More in particular, the Kazachstania yeast is K. bulderi or K. exigua. Even more in particular, the Kazachstania yeast is K. bulderi.

[0012] In particular embodiments, the at least one lactic acid bacteria is a sourdough lactic acid bacteria, such as belonging to the genus Lactobacillus, Fructolactobacillus, Lactiplantibacillus, Levilactobacillus, Umosilactobacillus, Pediococcus, Leuconostoc or Weiseiia.

[0013] In particular embodiments, the dominant non-Saccharomyces yeast, particularly the dominant Kazachstania yeast, is present in the sourdough in a concentration between 106and 1O10CFU / ml or CFU / g, and the at least one lactic acid bacteria is present in the sourdough in a concentration between 106and 1O10CFU / ml or CFU / g. In particular embodiments, the sourdough is added to the wort in an amount between 1 and 100 g / l or ml / l wort. In particular embodiments, the yeast and lactic acid bacteria are each added to the wort in an amount between 103and 109CFU / ml wort.

[0014] In particular embodiments, the wort is prepared from the malt of a cereal or pseudocereal, and optionally an unmalted cereal or pseudocereal, such as from malted or unmalted barley, wheat, corn, spelt, rye, rice, oats, sorghum, millet, buckwheat, quinoa, amaranth, or teff.

[0015] In particular embodiments, the method further comprises adding a hop or a hop derivative to the wort.

[0016] A second aspect of the present invention relates to a kit comprising:

[0017] (i) a Kazachstania yeast and at least one lactic acid bacteria; and / or a sourdough comprising a non- Saccharomyces yeast species, particularly a Kazachstania yeast, as the dominant yeast species and at least one lactic acid bacteria; and

[0018] (ii) one or more items or ingredients suitable to produce a beer, wherein the one or more items or ingredients comprise at least one item selected from the group consisting of: prepared wort solution; dry malt extract; one or more cereal or pseudocereal grains, such as barley, wheat, corn, spelt, rye, rice, oats, sorghum, millet, buckwheat, quinoa, amaranth or teff; one or more hop varieties or hop derivatives; and one or more sugar adjuncts.

[0019] In some embodiments, the sourdough in the kit comprises a yeast belonging to the genus Kazachstania, Pichia, Torulaspora, Candida, or Lachancea as the dominant yeast species. In particularly embodiments, the Kazachstania yeast is Kazachstania bulderi, Kazachstania exigua, Kazachstania barnettii or Kazachstania unispora, particularly the Kazachstania yeast is K. bulderi or K. exigua, even more particularly the Kazachstania yeast is K. bulderi’, and / or the at least one lactic acid bacteria is a sourdough lactic acid bacteria, such as belonging to the genus Lactobacillus, Fructolactobacillus, Lactiplantibacillus, Levilactobacillus, Umosilactobacillus, Pediococcus, Leuconostoc or Weiseiia.

[0020] The kit may further comprise instructions for producing a beer.

[0021] A further related aspect of the present invention relates to the use of the combination of a Kazachstania yeast and at least one lactic acid bacteria for the production of beer. The present invention also relates to the use of a sourdough comprising a r\or\-Saccharomyces yeast species, particularly a Kazachstania yeast species, as the dominant yeast species and at least one lactic acid bacteria for the production of beer. In some embodiments, the sourdough comprises a yeast belonging to the genus Kazachstania, Pichia, Torulaspora, Candida, or Lachancea as the dominant yeast species. In particular embodiments, the Kazachstania yeast is Kazachstania bulderi, Kazachstania exigua, Kazachstania barnettii or Kazachstania unispora, particularly the Kazachstania yeast is K. bulderi orK. exigua, even more particularly the Kazachstania yeast is K. bulderr, and / or the at least one lactic acid bacteria is a sourdough lactic acid bacteria, such as belonging to the genus Lactobacillus, Fructolactobacillus, Lactiplantibacillus, Levilactobacillus, Umosilactobacillus, Pediococcus, Leuconostoc or Wei sella.

[0022] Detailed description of invention

[0023] Before the present method and devices used in the invention are described, it is to be understood that this invention is not limited to particular methods, components, or devices described as such methods, components, and devices may, of course, vary. It is also to be understood that the terminology used herein is not intended to be limiting, since the scope of the present invention will be limited only by the appended claims.

[0024] Although any method and material similar or equivalent to those described herein may be used in practice or testing of the present invention, the preferred methods and materials are now described.

[0025] In this specification and the appended claims, the singular forms “a”, “an”, “the” include both the singular and the plural, unless the context clearly indicates otherwise.

[0026] The terms “comprising”, “comprises” and “comprised of’ as used herein are synonymous with “including”, “includes” or “containing”, “contains”, and are inclusive or open-ended and do not exclude additional, non-recited members, elements or method steps. Where this description refers to a product or process which “comprises” specific features, parts or steps, this refers to the possibility that other features, parts or steps may also be present, but may also refer to embodiments which only contain the listed features, parts or steps. The terms “comprising”, “comprises” and “comprised of” also include the term “consisting of”.

[0027] The enumeration of numeric values by means of ranges of figures comprises all values and fractions in these ranges, as well as the cited end points.

[0028] The terms “about” and “approximately” as used when referring to a measurable value, such as a parameter, an amount, a time period, and the like, is intended to include variations of + / - 10% or less, preferably + / -5% or less, more preferably + / -1 % or less, and still more preferably + / -0.1 % or less, of and from the specified value, in so far as the variations apply to the invention disclosed herein. It should be understood that the value to which the term “about” or “approximately” refers per se has also been disclosed.

[0029] Reference throughout this specification to “one embodiment” or “an embodiment” means that a particular feature, structure or characteristic described in connection with the embodiment is included in at least one embodiment of the present invention. Thus, appearances of the phrases “in one embodiment” or “in an embodiment” in various places throughout this specification are not necessarily all referring to the same embodiment, but may. Furthermore, the particular features, structures or characteristics may be combined in any suitable manner, as would be apparent to a person skilled in the art from this disclosure, in one or more embodiments. Furthermore, while some embodiments described herein include some but not other features included in other embodiments, combinations of features of different embodiments are meant to be within the scope of the invention, and form different embodiments, as would be understood by those in the art. For example, in the following claims, any of the claimed embodiments can be used in any combination.

[0030] Unless defined otherwise, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this invention belongs.

[0031] In the context of the present invention, it is understood that the different steps in a traditional process of brewing beer are well known to the skilled person. Briefly, malt is prepared from dried, germinated cereal grains, primarily barley or wheat, and grounded into a grist, which may also contain unmalted adjuncts. The grist is mashed, i.e. mixed with water, steeped and heated, to allow enzymes in the malt to convert the starch into sugars. The remaining grain particles and undissolved adjuncts are separated from the liquid wort in a process called lautering. The malt making and mashing steps can be skipped by adding water to malt extract. After addition of hops and / or other ingredients such as herbs and sugars, the wort is boiled, cooled and aerated. Hops may also be added during or after boiling. The wort is then moved to a fermentation tank and fermented by the addition of a brewer’s yeast. The primary fermentation, lasting typically 5 to 10 days, may be followed by a secondary fermentation step with brewer's yeast. After fermentation the fresh beer or "green" beer, is conditioned, optionally filtrated, carbonated and bottled.

[0032] As used herein, the expression “<genus name> yeast” is in the meaning of and used interchangeably with the expression “a yeast belonging to the genus <genus name>”. For instance, the expression “Kazachstania yeast” refers to and is used interchangeably with “a yeast belonging to the genus Kazachstania". The same holds for the lactic acid bacteria mentioned herein.

[0033] The term “CFU” means “colony forming unit”. This is a unit which estimates the number of microbial cells, such as bacteria or yeasts (fungi) in a sample that are viable and able to multiply under controlled conditions.

[0034] The inventors have prepared a high-quality beer by performing the wort fermentation step in a beer brewing process with a combination of a r\or\-Saccharomyces yeast, particularly a Kazachstania yeast, and a lactic acid bacteria, in particular by performing the wort fermentation step with a sourdough comprising the r\or\-Saccharomyces yeast, particularly the Kazachstania yeast, as the dominant yeast and lactic acid bacteria. The inventors found that during the fermentation an attenuation of more than 70% was obtained, assuring a good stability of the beer over time. Furthermore, the flavour of the obtained beer was balanced and intense, with fruity flavour notes. The flavour profile was also characterized by a full body. Advantageously, the brewing process according to the present invention resulted in a full body tasting beer, with the minimum and purest of brewing ingredients only.

[0035] In a first aspect, the present invention provides a method of brewing beer comprising the steps of a) preferably, providing a wort; and b) fermenting the wort with

[0036] - a Kazachstania yeast and a lactic acid bacteria; and / or

[0037] - a sourdough, wherein the sourdough comprises a r\or\-Saccharomyces yeast as the dominant yeast species, and at least one lactic acid bacteria, wherein the dominant yeast species is present in the sourdough in a concentration that is at least one log CFU / ml or CFU / g higher than any other yeast strain, which is optionally present in the sourdough.

[0038] More in particular, the present invention provides a method of brewing beer comprising the steps of a) preferably, providing a wort; and b) fermenting the wort with a Kazachstania yeast and a lactic acid bacteria; wherein, in step b), the wort is fermented with a sourdough, wherein the sourdough comprises the Kazachstania yeast as the dominant yeast species, and at least one lactic acid bacteria, wherein the dominant yeast species is present in the sourdough in a concentration that is at least one log CFU / ml or CFU / g higher than any other yeast strain, which is optionally present in the sourdough.

[0039] As used herein, the term "wort" has the conventional meaning in the field of brewing beer and refers to the sugary liquid extracted from the mashing process in a beer brewing process. The term "beer" as used herein refers at least to beers prepared from mashes prepared from malted and / or unmalted cereals, and optionally from mashes prepared with adjuncts. The term “beer” is not limited to a particular alcohol content. Advantageously, the beer obtained by a brewing method according to the present invention has a reduced alcohol content, such as ranging from 0.5 vol% to 5.0 vol%, more particularly ranging from 1.2 vol% to 4.5 vol% or ranging from 2.5 vol% to 4.0 vol%.

[0040] In particular embodiments, the wort is fermented with a Kazachstania yeast and at least one lactic acid bacteria. More in particular, the Kazachstania yeast and the lactic acid bacteria comprise species that occur in and are isolated from a sourdough. Particular examples of Kazachstania species and lactic acid bacteria are described below. The wort may be inoculated with a co-culture of the Kazachstania yeast and the lactic acid bacteria, or the Kazachstania yeast and lactic acid bacteria may be added as separate individual purified strains, either at the same time or at different moments in the fermentation.

[0041] In particular embodiments, the wort is fermented with a sourdough comprising a non- Saccharomyces yeast species as the dominant yeast species, and at least one lactic acid bacteria. As used herein, the dominant yeast species or strains in a sourdough refers to a yeast species or strain that is present in a concentration that is at least one log CFU / ml or CFU / g higher than any other yeast species or strain that may be present in the sourdough. Surprisingly, despite the complex microflora of a sourdough, comprising a variety of yeasts and bacteria, and the presence of lactic acid bacteria in the sourdough, a well-balanced beer with good beer quality properties and without off-flavours was obtained when fermenting a wort with a sourdough as defined elsewhere herein.

[0042] The term “sourdough” as used herein is defined as a mixture comprising flour and water, such as a dough or batter, which is fermented by lactic acid bacteria, particularly in combination with one or more yeasts, and which is optionally dried. The flour used for sourdough production and propagation includes but is not limited to flour from wheat, durum wheat or rye. In the context of the present invention, the sourdough is particularly fermented by lactic acid bacteria in combination with a r\or\-Saccharomyces yeast as the dominant yeast species. It is understood that a sourdough typically contains a complex microflora containing different lactic acid bacteria and yeast species.

[0043] In general, there is a large variety in which yeast is the dominant yeast species in a sourdough. In many sourdoughs, traditionally used in the production of baked products, such as bread Saccharomyces cerevisiae is the dominant yeast. However, in other sourdoughs, non- Saccharomyces yeasts dominate the yeast microflora. Examples of such yeasts include but are not limited to yeasts belonging to the genus Kazachstania, Pichia, Torulaspora, Lachancea, Candida, Meyerozyma, or Metschnikowia. Accordingly, in the context of the present invention, in particular embodiments, the wort is inoculated with a sourdough comprising, as the dominant yeast species, a Kazachstania yeast, a Pichia yeast, a Torulaspora yeast, a Lachancea yeast, a Candida yeast, a Meyerozyma yeast, a Metschnikowia yeast or another r\or\-Saccharomyces yeast. Particularly preferred are sourdoughs comprising, as the dominant yeast species, a Kazachstania yeast, a Pichia yeast, a Torulaspora yeast, a Lachancea yeast, or a Candida yeast, or stated differently, a yeast belonging to the genus Kazachstania, Pichia, Torulospora, Lachancea, or Candida. Even more particularly preferred are sourdoughs, comprising, as the dominant yeast species, a Kazachstania yeast or a Lachancea yeast, such as Lachancea thermotolerans. In even more particular embodiments, the sourdough comprises a Kazachstania yeast as the dominant yeast species. Accordingly, in particular embodiments, the wort may be inoculated with a sourdough comprising a Kazachstania yeast as the dominant yeast species and lactic acid bacteria.

[0044] In particular embodiments, the Kazachstania yeast, when used in an isolated form, e.g. as a purified strain or occurring as the dominant species in a sourdough, may be any Kazachstania yeast that occurs naturally in sourdoughs. Examples of Kazachstania yeast species include but are not limited to K. bulderi, K. exigua, K. barnettii, or K. unispora. Particularly preferred are K. bulderi or K. exigua, even more particularly preferred is K. bulderi.

[0045] Similar as for the yeasts, there is a large variety of lactic acid bacteria naturally occurring in sourdoughs. Lactic acid bacteria fermentation in the sourdough generates lactic acid and / or acetic acid that give the acidity to the sourdough. In the context of the present invention, the lactic acid bacteria is not particularly limiting. Examples of lactic acid bacteria, either used in isolated form, e.g. as a purified strain or as present in the sourdough, include but are not limited to sourdough lactic acid bacteria belonging to the genus Lactobacillus, Fructolactobacillus, Lactiplantibacillus, Levilactobacillus, Umosilactobacillus, Pediococcus, Leuconostoc or Weisella. In this context, it is noted that the genus Lactobacillus has recently been the subject of a taxonomic reorganisation, resulting in the classification of the old Lactobacillus species in 23 novel genera (Zheng et al., 2020, DOI 10.1099 / ijsem.0.004107). In the context of the present invention, the lactic acid bacteria may also be a Lactobacillus bacteria, according to the older taxonomic classification. In the methods as envisaged herein, the lactic acid bacteria includes but is not limited to Fructolactobacillus sanfranciscensis, Lactiplantibacillus plantarum, Levilactobacillus brevis, Umosilactobacillus fermentum, Umosilactobacillus pontis, Lentilactobacillus kefiri, Lac bacillus delbrueckii, Leuconostoc mesenteroides or Weisella confusa. In certain embodiments, the lactic acid bacteria may be a probiotic bacteria, which may provide further health benefits.

[0046] In particular embodiments, the wort is fermented with a Kazachstania yeast, such as K. bulderi or K. exigua, in combination with Fructolactobacillus sanfranciscensis and / or Lactiplantibacillus plantarum, following inoculation of the wort either with the microorganisms in isolated or purified form, or either with a sourdough comprising the Kazachstania yeast as the dominant yeast species and F. sanfranciscensis and / or Lactiplantibacillus plantarum as the lactic acid bacteria. Advantageously, in sourdoughs, when a yeast species belonging to the genus Kazachstania is dominant, it often appears synergistically with F. sanfranciscensis and / or Lactiplantibacillus plantarum.

[0047] Advantageously, the combination of lactic acid bacteria and a sourdough yeast as described herein, particularly a Kazachstania yeast as described herein, contributes positively to the flavour profile of the beer, such as the beer having a full body flavour profile, despite the reduced alcohol content, and an intense and balanced flavour with fruity flavour notes. Without wishing to be bound by theory, the sourdough yeasts as described herein, particularly a Kazachstania yeast as described herein, produce higher amounts of secondary alcohols and ester molecules from organic acid and alcohol. The lactic acid bacteria also add extra flavour during the combined fermentation.

[0048] In particular embodiments, the dominant non- Saccharomyces yeast species, as described elsewhere herein, particularly the dominant Kazachstania yeast species, is present in the sourdough in a concentration between 106and 1O10CFU / ml or CFU / g, and / or the at least one lactic acid bacteria is present in the sourdough in a concentration between 106and 1O10CFU / ml or CFU / g. It is further understood that a Saccharomyces yeast may be present in the sourdough, particularly in view of the sourdough typically being the result of a spontaneous fermentation of a dough or batter. In this case, i.e. when the sourdough contains a Saccharomyces yeast, it is present in a concentration that is at least one log CFU / ml or CFU / g, particularly at least two, three or four log CFU / ml or CFU / g, lower than the dominant non- Saccharomyces yeast species.

[0049] The wort fermentation is initiated by adding (a) an effective amount of the combination of a Kazachstania yeast, as described elsewhere herein, and a lactic acid bacteria, as described elsewhere herein, or (b) an effective amount of the sourdough, as described elsewhere herein, comprising a r\or\-Saccharomyces yeast, particularly a Kazachstania yeast, as the dominant yeast species and a lactic acid bacteria. Typically, the sourdough is added to the wort in an amount between 1 and 100 g / l or ml / l wort and / or wherein the yeast and lactic acid bacteria species are each added to the wort in an amount between 103and 109CFU / ml wort. It is understood that adding a lower concentration of the inoculum will increase the fermentation time, while adding a higher concentration will lower the fermentation time. In certain embodiments, the wort fermentation is performed for at least 5 days, such as between 5 and 14 days or between 5 and 10 days, but may be carried out for several weeks, such as up to 21 or 28 days or even for longer periods.

[0050] In certain embodiments, the fermentation is performed at a temperature between 12°C and 30°C, such as between 18°C and 25°C.

[0051] In certain embodiments, particularly in case when the wort is fermented with purified strains of the r\or\-Saccharomyces yeast, particularly the Kazachstania yeast, and lactic acid bacteria, the wort is fermented in the absence of a Saccharomyces and / or Brettanomyces yeast. In particular embodiments, the sourdough is an active sourdough. By this is meant that the sourdough product contains active and / or viable microorganisms, i.e. active and / or viable yeasts, particularly a non- Saccharomyces yeast, more particularly a Kazachstania yeast, and lactic acid bacteria.

[0052] In the context of the present invention, the sourdough as envisaged herein may be provided in many forms, particularly defined by its dry matter content. In certain embodiments, the sourdough as envisaged herein is a liquid sourdough, particularly an active liquid sourdough. Liquid sourdoughs are a commonly used type of sourdough with guaranteed homogeneity in quality and processing. A liquid sourdough product is characterized by having a dry matter between 5% (w / w) and 50% (w / w), preferably between 10% and 40% (w / w), more preferably between 10% and 35% (w / w). Other types of sourdoughs may be used as well, such as paste or stiff, or dry sourdoughs. In certain alternative embodiments, the sourdough as envisaged herein is a dry sourdough, particularly an active dry sourdough. Dry sourdoughs are typically obtained by drying a liquid sourdough. A dry sourdough product is characterized by a dry matter content of at least 85% (w / w), preferably at least 90%, or at least 92% or more. In certain embodiments, the sourdough as envisaged herein, the sourdough is a paste or stiff sourdough, typically having a dry matter content of higher than 50% (w / w), but lower than 85% (w / w).

[0053] It is further understood that the methods according to the present invention may include one or more of the steps of a traditional beer brewing process, such as (malt) grinding, mashing, lautering, wort boiling, hop addition, wort cooling, fermentation, maturation, filtering and bottling.

[0054] In certain embodiments, the method further comprises adding a hop or hop derivative to the wort.

[0055] In general, two kinds of hops are used in beer brewing: bitter hops and aroma hops. Bitter hops are typically used for lager beers to give extra bitterness to the beer. Aroma hops are used for specialty beers to enhance the flavor. The hop or hop derivative may be added prior to or at the start of wort boiling, particularly a bitter hop for increasing the bitterness, or during or at the end of wort boiling, particularly an aroma hop for aromatization. In addition, for extra hop flavour, hop may be added during the maturation phase, also known as dry-hopping. Many hop varieties are known to the skilled person. It is understood that the present method is not limited to one or more particular hop varieties.

[0056] In certain embodiments, the method further comprises preparing a wort from a cereal malt, and optionally an unmalted cereal or pseudocereal. The term "cereal" generally refers to plants of the botanical family of the Poaceae, particularly to the edible components (seeds) thereof, including but not limited to species such as wheat, barley, oat, spelt, rye, sorghum, corn, triticale, millet, teff and rice. The term “pseudocereal” refers to plants not belonging to the Poaceae family, but being able to be processed and used as a cereal. Examples of pseudocereals include buckwheat, amaranth and quinoa.

[0057] In a further related aspect, the present invention provides a kit comprising the yeast strain, particularly the Kazachstania yeast, the lactic acid bacteria and / or the sourdough as described elsewhere herein.

[0058] In particular embodiments, the kit comprises:

[0059] (i) a Kazachstania yeast and at least one lactic acid bacteria; and / or a sourdough comprising a r\or\-Saccharomyces yeast species, particularly a Kazachstania yeast species, as the dominant yeast species and at least one lactic acid bacteria species; and

[0060] (ii) one or more items or ingredients suitable to produce a beer.

[0061] In particular embodiments, the kit comprises a Kazachstania yeast, such as K. bulderi or K. exigua, particularly K. bulderi, and a lactic acid bacteria, as described elsewhere herein, as isolated strains, or as a co-culture.

[0062] In certain embodiments, the sourdough comprises a Kazachstania yeast, a Pichia yeast, a Torulaspora yeast, a Candida yeast or a Lachancea yeast as the dominant yeast species. In certain embodiments, the Kazachstania yeast is Kazachstania bulderi or Kazachstania exigua, more in particular is K. bulderi. In certain embodiments, the at least one lactic acid bacteria is a sourdough lactic acid bacteria, such as belonging to the genus Lactobacillus, Fructolactobacillus, Lactiplantibacillus, Levilactobacillus, Umosilactobacillus, Pediococcus, Leuconostoc or Weiseiia.

[0063] Said one or more items or ingredients suitable to produce a beer include, but are not limited to a prepared wort solution; a dry malt extract; one or more cereal or pseudocereal grains, such as barley, wheat, corn, spelt, rye, rice, oats, sorghum, millet, buckwheat, quinoa, amaranth, or teff; one or more hop varieties or hop derivatives; and one or more sugar adjuncts.

[0064] In certain embodiments, the kit may further comprise instructions or instructional materials for producing a beer using the components of the kit. The instructions or instructional material includes a publication, a recording or any other medium of expression that can be used to communicate the use of the kit or a component present therein. Delivery of the instructions can be a physical delivery of the publication or medium of expression, but may also be achieved by electronic transmission, e.g. a download from a website or by electronic mail. In a further related aspect, the present invention relates to the use of the combination of a Kazachstania yeast, as described elsewhere herein, and at least one lactic acid bacteria, as described elsewhere herein, for the production of beer, in particular for the fermentation of a wort.

[0065] The present invention also further relates to the use of a sourdough, as described elsewhere herein, i.e. comprising a non-Saccharomyces yeast species, particularly a Kazachstania yeast species, as the dominant yeast species and at least one lactic acid bacteria species, for the production of beer, in particular for the fermentation of a wort. In particular, the Kazachstania and / or the lactic acid bacteria, particularly in isolated form, and / or the sourdough comprising a r\or\-Saccharomyces yeast, particularly the Kazachstania yeast, and a lactic acid bacteria are used as a starter culture or inoculum for initiating the wort fermentation.

[0066] In certain embodiments, the sourdough comprises a Kazachstania yeast, a Pichia yeast, a Torulaspora yeast, a Candida yeast or a Lachancea yeast as the dominant yeast species.

[0067] In certain embodiments, the Kazachstania yeast is Kazachstania bulderi or Kazachstania exigua, more in particular is K. bulderi. In certain embodiments, the at least one lactic acid bacteria is a sourdough lactic acid bacteria, such as belonging to the genus Lactobacillus, Fructolactobacillus, Lactiplantibacillus, Levilactobacillus, Umosilactobacillus, Pediococcus, Leuconostoc or Weiseiia.

[0068] Examples

[0069] Example 1 - preparation of a sourdough beer

[0070] A / The beer was prepared from the following raw materials:

[0071] • Cereals: Barley malt : 2.25Kg; Spelt flour : 0.25Kg

[0072] • Water: Tap water

[0073] • Hops: Saaz (4,3 % a-alpha): 39.54g, added during cooking; Citra (13% a-alpha) : 5g, added 5min before end of the cooking

[0074] • Inoculum: a liquid durum wheat sourdough containing Kazachstania bulderi as the dominant yeast and Lactilactobacillus plantarum. It was directly added to the wort in an amount providing about 107to 5 x 109CFU of K. bulderi per liter wort and providing about 107to 5 x 109CFU of L. plantarum per liter wort.

[0075] The salt content of the tap water is shown in Table 1. Table 1.

[0076] B / The brewing process comprises the following steps

[0077] The ground cereals (2.5 kg) were mixed with 121 water. The cereal mash was subject to a stepwise heating process, in particular first 60 min at 65 °C, followed by 10 min at 72 °C to reach the sugar level and finally 10 min at 78°C.

[0078] Filtration was performed at a temperature of 80°C and the wort had the desired density of 7.5 °P at the end of filtration.

[0079] Next, the wort was cooked for 60 minutes (after the appearance of the first bubbles). At the start of the cooking, 1.7 g / l of Saaz hop was added to the wort (or 39.45 g for 23I wort), and 5g of Citra hop was added 5 min before the end of cooking. At the end of cooking, the density of the wort was 8.5 °P.

[0080] After cooking, the wort was clarified by centrifugation.

[0081] C / The fermentation was started by addition of 3ml / l of a durum wheat based sourdough containing Kazachstania bulderi as the dominant yeast strain and Lactilactobacillus plantarum. The fermentation was performed at a temperature of 24 °C for 8 days, followed by a maturation step for 2 days at 4 °C.

[0082] D / The characteristics of the final beer were as follows.

[0083] • Alcohol level : 3.5% v / v

[0084] • Bitterness : 31 IBU

[0085] • Color : 6.3 EBC

[0086] • pH : 3.97

[0087] During the fermentation of 8 days at 24°C, the sugars in the wort were metabolized by the sourdough yeasts and bacteria, to reach an attenuation of more than 70%. This assures a good stability of the beer over time / during storage. A high attenuation is a basic requirement for a beer process, but it has never been reported before for a brewing process comprising the co-fermentation by a sourdough yeast and a lactic acid bacteria. The consumption of the sugars ha to be controlled during the brewing process. The beer had a reduced alcohol level. However, its flavour profile was very full bodied, probably due to the combined fermentation with the lactic acid bacteria adding extra flavour through the fermentation. The flavour was also very intense with nice fruity flavour notes. This is presumably due to the use of the special sourdough yeast (Kazachstania), which produces higher amounts of secondary alcohols and ester molecules compared to S. cerevisiae. The flavour was very well balanced and did not show any unacceptable flavours.

[0088] Due to the low alcohol content of the beer, it is also expected to have a low calory content. Normally, low calory drinks are very flat in taste or need to be pimped with a lot of flavours or additives. The present sourdough based brewing process allowed to obtain a beer with a full body taste with the purest ingredients of brewing only. Furthermore, by selecting specific lactic acid bacteria, other health promoting properties could be obtained.

Claims

Claims1 . A method of brewing beer comprising the steps of a) providing or preparing a wort; and b) fermenting the wort with a Kazachstania yeast and at least one lactic acid bacteria species.

2. The method according to claim 1 , wherein the Kazachstania yeast is K. bulderi, K. exigua, K. barnettii orK. unispora, particularly wherein the Kazachstania yeast is K. bulderi or K. exigua, more particularly wherein the Kazachstania yeast is K. bulderi.

3. The method according to claim 1 or 2, wherein the at least one lactic acid bacteria species is a sourdough lactic acid bacteria species, belonging to the genus Lactobacillus, Fructolactobacillus, Lactiplantibacillus, Levilactobacillus, Umosilactobacillus, Pediococcus, Leuconostoc or Weiseiia.

4. The method according to any one of claims 1 to 3, wherein, in step b), the wort is fermented with a sourdough, wherein the sourdough comprises a Kazachstania yeast species as the dominant yeast species, and at least one lactic acid bacteria species, wherein the dominant yeast species is present in the sourdough in a concentration that is at least one log CFU / ml or CFU / g higher than any other yeast strain, which is optionally present in the sourdough.

5. The method according to claim 4, wherein in case the sourdough comprises a Saccharomyces yeast, it is present in a concentration that is at least one log CFU / ml or CFU / g, particularly at least two, three or four log CFU / ml or CFU / g, lower than the dominant Kazachstania yeast species and / or wherein the wort is fermented in the absence of a Saccharomyces and / or Brettanomyces yeast.

6. The method according to claim 4 or 5, wherein the sourdough is an active liquid sourdough, an active stiff sourdough or an active dried sourdough.

7. The method according to any one of claims 4 to 6, wherein the dominant Kazachstania yeast species is present in the sourdough in a concentration between 106and 1O10CFU / ml or CFU / g, and wherein the at least one lactic acid bacteria is present in the sourdough in a concentration between 106and 1O10CFU / ml or CFU / g.

8. The method according to any one of claims 4 to 7, wherein the sourdough is added to the wort in an amount between 1 and 100 g / l or ml / l wort.

9. The method according to any one of claims 1 to 8, wherein the yeast and lactic acid bacteria are each added to the wort in an amount between 103and 109CFU / ml wort.

10. The method according to any one of claims 1 to 9, wherein the method further comprises adding a hop or a hop derivative to the wort, in particular during cooking of the wort and / or maturation of the fermented wort.11 . The method according to any one of claims 1 to 10, wherein the wort is prepared from the malt of a cereal or pseudocereal, and optionally an unmalted cereal or pseudocereal, such as from malted or unmalted barley, wheat, corn, spelt, rye, rice, oats, sorghum, millet, buckwheat, quinoa, amaranth or teff.

12. A kit comprising:(i) a Kazachstania yeast and at least one lactic acid bacteria species; and / or a sourdough comprising a Kazachstania yeast species as the dominant yeast species and at least one lactic acid bacteria species; and(ii) one or more items or ingredients suitable to produce a beer, wherein the one or more items or ingredients comprise at least one item selected from the group consisting of: prepared wort solution; dry malt extract; one or more cereal or pseudocereal grains, such as barley, wheat, corn, spelt, rye, rice, oats, sorghum, millet, buckwheat, quinoa, amaranth or teff; one or more hop varieties or hop derivatives; and one or more sugar adjuncts.

13. The kit according to claim 12, wherein the Kazachstania yeast is K. bulderi, K. exigua, K. barnettii or K. unispora, particularly wherein the Kazachstania yeast is K. bulderi or K. exigua, more particularly wherein the Kazachstania yeast is K. bulderr, and / or wherein the at least one lactic acid bacteria is a sourdough lactic acid bacteria, belonging to the genus Lactobacillus, Fructolactobacillus, Lactiplantibacillus, Levilactobacillus, Limosilactobacillus, Pediococcus, Leuconostoc or Weiseiia.

14. The kit according to claim 12 or 13, further comprising instructions for producing a beer.

15. Use of the combination of a Kazachstania yeast and at least one lactic acid bacteria species for the production of beer.

16. Use of a sourdough comprising a Kazachstania yeast species as the dominant yeast species and at least one lactic acid bacteria species for the production of beer.

17. The use according to claim 15 or 16, wherein the Kazachstania yeast is K. bulderi, K. exigua, K. barnettii orK. unispora, particularly wherein the Kazachstania yeast is K. bulderi or K. exigua, more particularly wherein the Kazachstania yeast is K. bulderi’, and / or wherein the at least one lactic acid bacteria is a sourdough lactic acid bacteria, belonging to the genus Lactobacillus, Fructolactobacillus, Lactiplantibacillus, Levilactobacillus, Limosilactobacillus, Pediococcus, Leuconostoc or Weiseiia.

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