Delete, Protect, bureaucrat, administrator
4,588
edits
Changes
no edit summary
=Spontaneous fermentation=
The spontaneous fermentation of lambic is a complex process involving a succession of bacteria and yeasts that progresses along with the chemical changes that occur during fermentation.<ref name=GeuzeKriek>Jef Van den Steen, [[Books#Geuze & Kriek: The Secret of Lambic Beer|Geuze & Kriek: The Secret of Lambic Beer]], 2012</ref> The spontaneous fermentation process has shown considerable variability even among different barrels of beer from the same brewery,<ref name=Spitaels> F. Spitaels, A. D. Wieme, M. Janssens, M. Aerts, H.-M. Daniel, A. Van Landschoot, L. De Vuyst, P. Vandamme [http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0095384 | The Microbial Diversity of Traditional Spontaneously Fermented Lambic Beer], 2000</ref> though all spontaneously fermented beers appear to follow a general sequence of microbes, which can be broken into four distinct stages:<ref name=AWAs>Nicholas A. Bokulich, Charles W. Bamforth, David A. Mills. [http://www.plosone.org/article/info%3Adoi%2F10.1371%2Fjournal.pone.0035507|Brewhouse-Resident Microbiota Are Responsible for Multi-Stage Fermentation of American Coolship Ale], PLoS One, 7(4), 2012</ref><ref name=Oevelen77 >D. Van Oevelen, M. Spaepen, P. Timmermans and H. Verachtert, [http://onlinelibrary.wiley.com/doi/10.1002/j.2050-0416.1977.tb03825.x/abstract|MICROBIOLOGICAL ASPECTS OF SPONTANEOUS WORT FERMENTATION IN THE PRODUCTION OF LAMBIC AND GUEUZE], 1977</ref><ref name=Spitaels />
# An enteric stage, starting around three days after the boil and ending between 30 and 90 days, in which enteric bacteria dominate.
[[file:Salmonella_micrograph.jpg|thumb|left|Electron micrograph of the enteric bacteria Salmonella (pink rods).]]
Lambic wort arrives in the [[koelschip]] at approximately 5% sugar per weight of water, along with an assortment of proteins and fatty acids and other compounds.<ref name=Erbe />T. Erbe and H. Brückner, [http://www.sciencedirect.com/science/article/pii/S0021967300002557| Chromatographic determination of amino acid enantiomers in beers and raw materials used for their manufacture], 2000</ref> Negligible ethanol is present prior to fermentation, nor is there much of the organic acids that will give lambic its characteristic tartness; the wort has a pH around 5, which is similar to the wort of other beers.<ref name=Oevelen77 >D. Van Oevelen, M. Spaepen, P. Timmermans and H. Verachtert, [http://onlinelibrary.wiley.com/doi/10.1002/j.2050-0416.1977.tb03825.x/abstract| MICROBIOLOGICAL ASPECTS OF SPONTANEOUS WORT FERMENTATION IN THE PRODUCTION OF LAMBIC AND GUEUZE], 1977</ref> Enteric bacteria, including Enterobacter hormaechei, E. kobei, Klebsiella pneumoniae, and Escherichia coli, are the first to gain a foothold in this environment, with significant numbers found after three to four days. The enteric bacteria primarily consume glucose, which reduces the gravity of the wort from ~1.050 to ~1.040 after the first three weeks.
Enteric bacteria are responsible for the production of [[acetic acid]], and the pH of the wort falls from around 5 to 4.5 in the first week of fermentation. The 40 to 120 mg/L acetic acid found in the wort after the first week is very close to the amount found in the final product.<ref name=Oevelen77 >D. Van Oevelen, M. Spaepen, P. Timmermans and H. Verachtert, [http://onlinelibrary.wiley.com/doi/10.1002/j.2050-0416.1977.tb03825.x/abstract| MICROBIOLOGICAL ASPECTS OF SPONTANEOUS WORT FERMENTATION IN THE PRODUCTION OF LAMBIC AND GUEUZE], 1977</ref>
<ref name=sour> J. Edwards and A. DiCaprio. [http://www.process-nmr.com/pdfs/Edwards%20-%20SMASH%202014%20-%20MNova%20Users%20Meeting%20-%209-7-14.pdf| When Beer Goes Sour: An NMR Investigation], Mestrelab
MNova Users Meeting, SMASH – Atlanta, GA, September 7, 2014</ref> Significant changes to the concentration of acetic acid should not occur until the ethanol has a chance to oxidize in aging in the bottle over many years or even decades.<ref name=Vanderhaegen1> B. Vanderhaegen, H. Neven, H. Verachtert, G. Derdelinckx [http://www.google.com/url?sa=t&rct=j&q=&esrc=s&source=web&cd=1&ved=0CCAQFjAA&url=http%3A%2F%2Fwww.researchgate.net%2Fprofile%2FGuy_Derdelinckx%2Fpublication%2F222839054_The_chemistry_of_beer_aging__a_critical_review%2Flinks%2F0c960523339c4b25a6000000.pdf&ei=Tq3IVKmfFcGyogSs_YLQCA&usg=AFQjCNFaBrvqDGjqEV2I9uQ73dYh_ParXg&sig2=Z8dY4iDHozbT1eb9JeAdrw&bvm=bv.84607526,d.cGU| The chemistry of beer aging – a critical review], 2006</ref><ref name = Werner> Werner Van Obberghen, '''2. Het algemene productieproces van bier'''</ref> The pellicle that forms on the top of the wort may be the product of acetobacteria during the enteric phase,[9] though most other sources inidcate that the pellicle is the result of Brettanomyces (with Pichia and Candida).<ref name="Guinard">Jean-Xavier Guinard, [[Books#Classic Beer Styles: Lambic|Classic Beer Styles: Lambic]], 1990</ref>
[[file:Saccharomyces_micrograph.jpg|thumb|right|Saccharomyces cerevisiae]]
After the pH falls below ~4.5 and the alcohol content rises over ~2%, [[Saccharomyces| Saccharomyces species]] take over as the dominant organisms in the wort, though Saccharomyces is present in large numbers well prior to the disappearance of the enterobacteria. Saccharomyces will remain dominant until at least 6 to 8 months into fermentation, and will maintain a presence, though no longer active, throughout fermentation. Despite Saccharomyces' importance to the fermentation of lambic, its concentrations remain below 10<sup>7</sup> cells/mL of wort, which is considerably lower than the 10<sup>8</sup> cells/mL found in commercial beers.<ref name=Oevelen77 >D. Van Oevelen, M. Spaepen, P. Timmermans and H. Verachtert, [http://onlinelibrary.wiley.com/doi/10.1002/j.2050-0416.1977.tb03825.x/abstract|MICROBIOLOGICAL ASPECTS OF SPONTANEOUS WORT FERMENTATION IN THE PRODUCTION OF LAMBIC AND GUEUZE], 1977</ref><ref name="Guinard">Jean-Xavier Guinard, [[Books#Classic Beer Styles: Lambic|Classic Beer Styles: Lambic]], 1990</ref>
As in controlled fermentation, Saccharomyces is responsible for most ethanol production and attenuation in lambic. The yeasts consume all the major sugars found in lambic wort (glucose, maltose, and some maltotriose). By the end of the Saccharomyces phase around 8 months, the ethanol content of the beer stabilizes at 5 to 7% by volume and will remain around that value until the end of fermentation.<ref name="Guinard">Jean-Xavier Guinard, [[Books#Classic Beer Styles: Lambic|Classic Beer Styles: Lambic]], 1990</ref><ref name=Oevelen77 >D. Van Oevelen, M. Spaepen, P. Timmermans and H. Verachtert, [http://onlinelibrary.wiley.com/doi/10.1002/j.2050-0416.1977.tb03825.x/abstract|MICROBIOLOGICAL ASPECTS OF SPONTANEOUS WORT FERMENTATION IN THE PRODUCTION OF LAMBIC AND GUEUZE], 1977</ref> Attenuation after Saccharomyces fermentation reaches 60 to 65%, which is known as the "attenuation limit" for conventional beers. Despite being responsible for most of the ethanol in lambic, yeasts of the Saccharomyces genus are not responsible for most of the aroma and flavor compounds that give lambic its distinct sensory characteristics.<ref name = Witrick1> K. A. T. Witrick [https://vtechworks.lib.vt.edu/handle/10919/19203| Characterization of aroma and flavor compounds present in lambic (gueuze) beer], 2012 </ref>
Other bacteria and yeasts also thrive during the Saccharomyces phase. [[Kloeckera]] and [[Debaryomyces]] both survive in considerable numbers alongside Saccharomyces.<ref name=Oevelen77 >D. Van Oevelen, M. Spaepen, P. Timmermans and H. Verachtert, [http://onlinelibrary.wiley.com/doi/10.1002/j.2050-0416.1977.tb03825.x/abstract|MICROBIOLOGICAL ASPECTS OF SPONTANEOUS WORT FERMENTATION IN THE PRODUCTION OF LAMBIC AND GUEUZE], 1977</ref>
<ref name=Spitaels /> In the case of Kloeckera, the growth begins prior to Saccharomyces in the first days of fermentation, but are overgrown by Saccharomyces in the following weeks.
The lactic acid bacteria increase in number until around month 7, achieving concentrations of 10<sup>4</sup> cells/mL wort, which is thought to coincide with the onset of summer and warmer temperatures.<ref name=Spitaels /> With this in mind, it is thought that the increase in lactic acid bacteria be delayed or hastened by decreasing or increasing the storage temperatures, respectively.<ref name=Spitaels /><ref name=AWAs />
Interestingly, lactic acid bacteria have been implicated in racemizing amino acids in beer, causing lambic (and other beers which make use of lactic acid bacteria such as Berliner Wiesse) to have a high percentage of right-handed amino acid stereoisomers relative to both their starting materials and other beers.<ref name=Erbe >T. Erbe and H. Brückner, [http://www.sciencedirect.com/science/article/pii/S0021967300002557|Chromatographic determination of amino acid enantiomers in beers and raw materials used for their manufacture], 2000</ref>
==Brettanomyces dominance==
Tetrahydropyridines (THPs) produced by Brettanomyces (as well as some Lactobacilli) have a wide variety of odors and give lambic much of its "mousey" aroma, as well as cider- and horse-like aromas, though the concentrations and thus smells of THPs are variable.<ref name=Heresztyn1> T. Heresztyn [http://ajevonline.org/content/37/2/127.short| Formation of Substituted Tetrahydropyridines by Species of Brettanomyces and Lactobacillus Isolated from Mousy Wines], 1986</ref> Other important odor and flavor compounds produced by Brettanomyces include 4-ethylphenol, 4-ethylguaiacol, and isovaleric acid. 4-ethylphenol produces barnyard and horsey flavors which can taste like Band-aids in higher concentrations. 4-ethylguaiacol lends spicier flavors of bacon and cloves and can be smoky, while isovaleric acid gives lambic its sweaty and cheesy flavors and odors.
Around 16 months after the start of fermentation, during this stage, the pH of the beer reaches a minimum of about 3.0, which then rises slightly in the following months to ~3.2 to 3.4,<ref name = EtF> [http://embracethefunk.com/ph-readings-of-commercial-beers/| Embrace the Funk's list of beer pH]</ref><ref name=Oevelen77 >D. Van Oevelen, M. Spaepen, P. Timmermans and H. Verachtert, [http://onlinelibrary.wiley.com/doi/10.1002/j.2050-0416.1977.tb03825.x/abstract|MICROBIOLOGICAL ASPECTS OF SPONTANEOUS WORT FERMENTATION IN THE PRODUCTION OF LAMBIC AND GUEUZE], 1977</ref><ref name="Guinard">Jean-Xavier Guinard, [[Books#Classic Beer Styles: Lambic|Classic Beer Styles: Lambic]], 1990</ref> perhaps due to the enzymatic esterification of organic acids by Brettanomyces.
=Refermentation in the bottle=
[[File:BrasserieCantillon-18.jpg|thumb|left|Lambic fermenting in the bottle at Cantillon]]
Gueuze and other lambic bottled with either some residual sugar left unfermented at the time of bottling or added priming sugars will undergo significant fermentation in the bottle, though all unpasteurized lambic will continue to ferment to some degree there. For lambic bottled after about 8 months without additional sugar, the fermentation in the bottle progresses much as an extension of the Brettanomyces stage of fermentation and negligible carbon dioxide and ethanol production occurs, leaving most of this lambic still. For lambic bottled younger and those with additional fermentable sugars added at bottling, considerable fermentation by Saccharomyces occurs in the bottle, causing marked increases in ethanol and the production of carbon dioxide, leading to a carbonated product.<ref name=Oevelen77 >D. Van Oevelen, M. Spaepen, P. Timmermans and H. Verachtert, [http://onlinelibrary.wiley.com/doi/10.1002/j.2050-0416.1977.tb03825.x/abstract|MICROBIOLOGICAL ASPECTS OF SPONTANEOUS WORT FERMENTATION IN THE PRODUCTION OF LAMBIC AND GUEUZE], 1977</ref>
The reactivation of Saccharomyces fermentation at the addition of additional sugar indicates that the dominance of Brettanomyces is brought about by Brettanomyces' ability to ferment sugars that Saccharomyces cannot. Once Saccharomyces has consumed most of the available glucose and other simple sugars in the wort, it goes dormant, though does not die completely, and Brettanomyces is free to assume the role of primary yeast in the wort. Upon the re-introduction of the simple sugars, the faster-growing Saccharomyces once again flourishes, until the fermentable sugar is again consumed and Brettanomyces and its other associated yeasts can once again resume the slow process of the final fermentation.
==Geographical variation==
The rapid reproduction of the microorganisms found in lambic also lends to them the ability to evolve on much shorter timescales than those of macroorganisms. Combined with the flora found in lambic (and other spontaneous fermentations) being largely resident inside each brewery,<ref name=AWAs /><ref name=Oevelen77 >D. Van Oevelen, M. Spaepen, P. Timmermans and H. Verachtert, [http://onlinelibrary.wiley.com/doi/10.1002/j.2050-0416.1977.tb03825.x/abstract|MICROBIOLOGICAL ASPECTS OF SPONTANEOUS WORT FERMENTATION IN THE PRODUCTION OF LAMBIC AND GUEUZE], 1977</ref> it is reasonable to assume that even the slight geographic separation found between the facilities within the Pajottenland will lead to markedly different strains of bacteria and yeasts responsible for lambic fermentation after several years of operation. Further, brettanomyces isolated from different sources from the same location have shown significant genetic variability<ref name=Crauwels1> S. Crawels et. al. [http://link.springer.com/article/10.1007%2Fs00253-015-6769-9| Comparative phenomics and targeted use of genomics reveals variation in carbon and nitrogen assimilation among different Brettanomyces bruxellensis strains], 2015</ref> <ref name=Borneman1> A. Borneman et. al. [http://journals.plos.org/plosgenetics/article?id=10.1371/journal.pgen.1004161| Insights into the Dekkera bruxellensis Genomic Landscape: Comparative Genomics Reveals Variations in Ploidy and Nutrient Utilisation Potential amongst Wine Isolates], 2014</ref>, indicating that slightly different handling methodologies and even the surrounding architecture may have an effect on the final product. Thus, it is reasonable to assume that at least part of the distinct flavors found in the lambics of each brewery are due to divergent evolution in their respective microbiomes,<ref name = Witrick1> K. A. T. Witrick [https://vtechworks.lib.vt.edu/handle/10919/19203| Characterization of aroma and flavor compounds present in lambic (gueuze) beer], 2012 </ref> though little literature currently exists in this area. For other breweries, "house strains" developed by propagating favorite yeasts often over a period of years and even decades is commonplace, and often accounts for much of a brewery's specific taste.
Conversely, it has been found that spontaneous fermentations in similar worts in other parts of the world follow similar overall trends.<ref name=AWAs /> Yeasts such as Saccharomyces and Brettanomyces and bacteria such as the enterobacteria and Pediococcus are cosmopolitan throughout the world, and all are similar enough to be classified into the same genus by both phenotype and genotype. However, the large degree of geographic separation coupled with the rapid evolutionary rates of these organisms will still lead to measurable changes in both. This is highlighted in the fact that while many lambic-like beers have been brewed in other parts of the world, and are often very similar to lambic itself, they still possess noticeably different sensory characteristics. This can be likened to terroir in wine, as the local microbiome in one location will produce a similar, but not identical, product to that in another location.
Spontaneously fermented beers from the United States form a group of beers referred to as American Coolship Ales (or ACAs), American Wild Ales (or AWAs), among other names.<ref name=AWAs /> Spontaneously fermented beers from other parts of the world are occasionally referred to as "American Wild Ales" as well, though it is unclear if there is any reason to associate any arbitrary spontaneous or even mixed fermentation beer that doesn't fall into another existing style with the United States. At present, there are few satisfactory naming conventions for these beers.
ACAs vary in their intended similarity to lambic, with some American producers even going so far as to label their beers "lambics", while other ACAs bear little in common with lambic besides spontaneous fermentation. Due to the geographical separation between the United States and Belgium and the large variations in yeast and bacteria genetics between these different populations, even an ACA wort carefully prepared to be very similar to that of lambic will yield notably different results after fermentation, even if the overall experience of the two styles of beer is similar.<ref name=AWAs /> <ref name=sour> J. Edwards and A. DiCaprio. [http://www.process-nmr.com/pdfs/Edwards%20-%20SMASH%202014%20-%20MNova%20Users%20Meeting%20-%209-7-14.pdf| When Beer Goes Sour: An NMR Investigation], Mestrelab MNova Users Meeting, SMASH – Atlanta, GA, September 7, 2014</ref> <ref name=Oevelen77 >D. Van Oevelen, M. Spaepen, P. Timmermans and H. Verachtert, [http://onlinelibrary.wiley.com/doi/10.1002/j.2050-0416.1977.tb03825.x/abstract|MICROBIOLOGICAL ASPECTS OF SPONTANEOUS WORT FERMENTATION IN THE PRODUCTION OF LAMBIC AND GUEUZE], 1977</ref>
==Spontaneous fermentation in other traditional drinks and foods==