The Beer Brewer

The diagram below shows a typical evolution of lambic fermentation over time.

Key to the diagram: 1 : Ethanol, 2: Lactic acid, 3: Ethyl Acetate, 4: pH, 5: Extract content, 6: Acetic acid

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The following table shows the characteristics of three differnent styles of Lambic Beer:

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The soft and hard classifications are due primarily to the acid contents of the beer, whereas the rope characteristic describes the oily consistency of the beer due to high lactic acid levels.

The finished product is characterised by high levels of lactic acid, acetic acid, ethyl acetate and ethyl acetate. Final gravities are in the region of 1.008 (2.2 oP) 1.012, 3 oP, DMS levels are reduced to approx 100pb and diacetyl concentrations between 45-80ppb. Final 2,3 butanediol levels are not available.

Beers of the lambic style are described as having vinous aromas and tastes accompanies by ‘horse blanket’ or ‘goaty’ characterisitics, which appears to be attributable to the contained acids and products from Brettanomyces activity. Thin mouth feel is also noted, due to the low levels of residual dextrins.

The spontaneous nature of lambic fermentation and the varied spectra of bacteria and yeasts encountered in differing breweries that are responsible for such fermentation, may produce beers with differing characterisitics.

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Brettanomyces species, particularly B. limbicus and B. bruxellensis also become active during this period, from approximately 8 months onward (figure to follow). These yeasts are capable of fermenting dextrins (found in relatively high levels in lambic wort), maltotriose and maltotetraose, producing small amounts of acetic acid and ethanol. However, it is the production of unique flavour and aroma compounds that characterises the activity of Brettanomyces. Brettanomyces exhibits a high esterase activity, primarily resulting in the synthesis of large amounts of ethyl acetate and ethyl lactate. This esterase  also cleaves iso- amyl acetate esters, thus accounting for its low concentration in lambic beer.

Tetrahydropyridines are also produced from ethanol and the amino acid lysine, which impart a ‘mousy’ or ‘horsy’ aroma to the beer. Volatile phenolics, with medicinal, ‘barnyard’ or ‘animal’ type odours are also from from 4-ethyl phenol and 4-ethyl guaiacol compounds.

Brettanomyces activity also leads to a significant increase in caprylic (C8) and caprix (C10) acids (small amounts are formed during earlier Saccharomyces acitivity) along with their associated esters, ethyl caprylate and ethyl caprate. These compounds produce a characteristic ‘goaty’ aroma/flavour to the final product. Capric and caprylic acids are short chain fatty acids and are thought to be by-products of yeast metabolism, produced during lipid synthesis by a yeast cell and released into the medium via leakage through membranes damaged by ethanol, or as a consequence of an autolytic mechanism. Higher temperatures, aeration and agitation during fermentation reduce the amounts of fatty acids in beer. Reduced aeration/agitation produced higher concentrations. The combined threshold for C6-C12 acids is 10ppm, beyond which ‘goaty’ aromas arise. C8 acid levels in gueuze as 12.4 - 21.85 ppm and C10 acids as 2.3 - 3.9p ppm.

Brettanomyces is active until the end of the fermnetaion period. Acetic acid bacteria of the genus Acetomonas and a number of oxidative yeasts (Pichia, ) are also found during lambic fermentations, however their influence is often minimal.

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As the Saccaromyces species decline once the primary fermentation is completed (2 months), the secondary or lambification (acidifcation) phase, initially involving the hop tolerant Pediococcus damnosus, takes place. P. damnosus is the primary producer of lactic acid in lambic beer, as it is a homofermentative organism producing only lactic acid from the metabolism of glucose, via the EMP pathway. As lactic acid is a key component in lambic flavour, its production is considered here: Glucose undergoes enzymatic cleavage and substrate level phosphorlyation, producing glcyeraldehyde 3 - phosphate, a proportion of which is converted to pyruvate. Lactic acid is produced from pyruvate by the the enzyme lactate dehydrogenase. No gas is evolved. The taste threshold for lactic acid is 400ppm. Final concentrations are indicated as between 492 - 3677ppm.

Fermentation at temperatures above 20oC are required for P. damnosus growth, therefore increasing temperatures beyond this level will stimulate lactic acid production as cell numbers increase. P. damnosus is a also a facultative anaerobe, thus the exclusion of air during P. damnosus activity will presumably facilitate the further production of lactic acid.

P. damnosus is also a producer of diacetyl, prodcuing up to 200ppb during this period. Acetoin is also produced as a consequence . The production of large amounts of lactic acid result in a drop in beer pH from 4 to 3, with a gradual lowering of residual extract.

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The beer at this point contains 1000ppm lactic acid, 1000ppm 2,3-butanediol, 250ppm succinic acid, 200ppm acetic acid and 100ppm formic acid. Flacours identified as sweet, honey, fruity, vegetbale or faecal are associated with compounds produced by the Enterobacteriaceae. The slow emegence of the the predominant Saccharomyces species, S. cerevisiae and possibly S. globus is a result of the depletion of wort amino acids by Enterobacteriaceae activity. This signifies the primary period of alcohol production, during which 80% of the wort sugars (glucose, fructose, sucrose, maltose, and maltotriose) are metabolised via the EMP pathway. Ethanol and CO2 are the primary end products. Pre-formed volatile compounds (esters and DMS) may be driven out of the beer by CO2 evolution during this phase.

Higher, or Fusel alcholol synthesis is a feature of Saccharomyces fermentation, occurring during ethanol production. The fusel alchohols impart a plastic or solvent like flavour to the beer. Elevated levels of propanol, amyl alchohol, isobutanol, isoamyl alcohol and phenylethyl alchohol are not noted in lambic beer and are ths not distinctive flavour/aroma components of the style.

A significant proportion of the lambic esters, particularly ethyl acetate and ethyl lactate are produced during this period. Ester impart a full-bodied, floral/fruit flavour/aroma to beers. Excessive levels (noted in lambic beers) may lead to off or overly fruity flavours. Ethyl acetate is formed by a cellular esterification pathway within the yeast cell by a reaction between acetyl-CoA compounds and alcohol, catalysed by the acyl-alcohol transferase enzyme. The resulting ester produces a fruity/solventy taste in the beer and has a threshold detection level of 33pm. The increased ester formation is promoted under the following conditions: Wort density about 13o P, increasing attenuation levels, increasing fermentation temperatures, increasing agitation during fermentation and reducing wort aeration levels. The inverse of these conditions reduce ester formation. A profile of beer components at this stage indicates wort gravity decreased to approx 1.012, alchohol content is 4.5 - 5% v/v, butanediol content is unchanged whilst DMS levels are reduced.

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Continuing our article on Lambic Beers …

The primary phase of lambic fermentation occurs after the “infected” wort is transferred into the fermentation casks. Typical initial temperatures are between 18-20oC, with initial specific gravities ranging from 1.048 - 1.072, depending upon lambic style produced. Initial fermentation during this period is due to the activity of bacteria from the Enterbobacteriaceae family and Kloeckera yeast strains. The cooling wort is readily colonised by the Enterbobacteriaceae, with strains of Enterobacter cloacae and E. aerogene being indicated as the predominant species. Strains of Klebsiella aerogene, Escherichia coli, Citrobacter freundii and Hafnia alevi are also active at this point. All species are capable of fermenting glucose and/or lactose via the Embden-Meyerhof-Parnas (EMP) pathway, but cannot utilise maltose or maltotriose.

The Enterobacter, Keibsiella and Hafnia species utilise a butanediol fermentation, producing 2,3-butanediol, acetoin and ethanol from the metabolism of glucose. E. coli and C. freundii employ a mixed acid fermentation to primarily produce lactic acid and lesser amounts of acetic, succininc and formic acids, ethanol and CO2. The activity of the latter species results in the formation of the majority of acetic acid found in lambic beer. This bacterial activity also results in the production of large amounts (500ppb) of DMS.

The activity of Kloeckera apiculata, a yeast capable of mixed acid fermentation from the metabolism of glucose, during this phase has little impact upon the lambic flavour, as any volatile esters it may produce are lost due to subsequent CO2 evolution. The combination of bacteria and yeast growth within this period has minimal impact upon the worts fermentable sugars, a maximum attenuation of 15% may be realised, however a significant drop in wort pH is apparent, from 5.1 to 4.6 as a resul primarily of acid formation. K. apiculata is gradually supplanted after two weeks by the apperance of the Saccharomyces species, whilst the Enterobacteriaceae disappear after 30-40 days, due to their sensitivy to ethanol concentration above 2% and pH leveles below 5.5 .

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An Introduction to Lambic Beers

The Belgian lambics are unique beers, traditionally produced in the Payottenland region of Belgium, via a process of spontaneous fermentation, as opposed to the controlled inoculation and fermentation procedures used in the production of other beer styles. This type of traditional, spontaneous fermentation is employed in combination with differing raw materials and production processes, to produce beers with unique flavour and aroma profiles, which have not been reproducible via other methods. Spontaneous fermentation is facilitated by overnight exposure of the cooling (un-inoculated) wort to the predominant air borne microflora of the brewery, which rapidly establish themselves within the wort.

There are two major phases of fermentation underaken in lambic beer, both involving the activities of bacteria and yeast; a primary phase, lasting for 3-4 months, characterized by the production of high amounts of ethanol; and a second, slower period of “lambification” or acidification, lasting for 12-24 months. A subsequent third (bottle) fermentation phase may be used to produce a gueuze style lambic.

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