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==Microbes and Flavor Compounds==
===Introduction===
While most beer styles are fermented using one culture of ''Saccharomyces cerevisiae'' or ''S. pastorianus'', Flanders Red Ales are fermented with a [[Mixed Fermentation|mixed culture fermentation]]. At one brewery (presumed to be Rodenbach) studied by Martens et al., two beers were produced using mixed fermentation and blended together. The first "light beer" was 11°P and was less acidic, while the second "heavy beer" was 13°P and served as an Old Ale if unblended. Both beers were inoculated with an acid washed yeast slurry that was harvested from a previous fermentation of the "light beer", and contained about 5% lactic acid bacteria. The fermentation of these beers followed three stages: 1. a seven day ethanol fermentation that is dominated by ''Saccharomyces'', 2. a four to five week lactic acid fermentation that was dominated by ''Lactobacilli'', and 3. a twenty to twenty-four month fermentation that was dominated by ''Brettanomyces'', ''Lactobacilli'', ''Pediococcus parvulus'', and acetic acid bacteria. The development of the ''Brettanomyces'' and ''Pediococcus'' stage was similar to the development of these microbes in [[Lambic]] fermentation. The "light beer" was never allowed to go through the third phase of fermentation, and was instead chilled to 0°C and then used to blend with previous batches of the "heavy beer" <ref name="Martens">[http://onlinelibrary.wiley.com/doi/10.1002/j.2050-0416.1997.tb00939.x/abstract MICROBIOLOGICAL ASPECTS OF A MIXED YEAST—BACTERIAL FERMENTATION IN THE PRODUCTION OF A SPECIAL BELGIAN ACIDIC ALE. H. Martens, D. Iserentant andH. Verachtert. 1997.]</ref>. Interestingly, and perhaps frustratingly, Flanders red and brown ales have been the subject of published studies far less than Belgian lambic beers, so the following information is based off of the Martens et al. study (see reference).
====Primary Fermentation====
====Comparison to Lambic====
Lambic is a similar beer produced in Belgium, but is fermented using [[Spontaneous Fermentation|spontaneous fermentation]]. Enterbacteria were not found in the brewery that Martens et al. studied since spontaneous fermentation was not used. However, after the enterobacteria and primary ''Saccharomyces'' fermentation phases of lambic brewing are complete, the microbial populations of lambic and Flanders red/brown beers are similar during their aging processes. Both beers display a dominance by ''Brettanomyces'' and ''Pediococcus'' during the aging phases. Flanders Red Ales differ by having a large portion of the acid production performed by ''Lactobacilli'', where as in lambic the acid production is performed by ''Pediodoccus damnosus''. Flanders Red Ales are also characterized by having ''P. parvulus'' instead of ''P. damnosus''(although this may have been misidentified; see Modern Analysis below), however Martens et al. noted that the two species have no clear difference as far as their effects on fermentation go. Martens also noted that ''Brettanomyces'' began to disappear from old English Porter when the beer moved from wooden casks to metallic ones. It is thought then that the wooden casks are vital to ''Brettanomyces'' in Flanders Red Ale brewing, perhaps due to the presence of ''Pediococci'', with which ''Brettanomyces'' "cooperates" to ferment dextrins in the beer during the aging phase <ref name="Martens"></ref>.
Belgian brewers have even married the Flanders Red Ale and Lambic by blending the two beers together. The Flanders "acid beer" is fermented with cherries, and later on Lambic is added and the blend is allowed to referment in the bottle, creating something truly special <ref name="Verachtert"></ref>.
====Modern Analysis====
A more recent study by Snaewaert et al. (see reference <ref name="snauwaert"></ref>) looked at the microbial and metabolic composition in the finished beer of the same brewery as Martens et al., as well as two other Flanders Red Ale breweries using "state of the art" DNA sequencing methods. Three samples were analyzed (A, B, and C) from each brewery (1, 2, and 3). As expected, there were both similarities and differences between the three breweries tested, as well as some differences between the individual beers from each brewery. The bacterial populations for Breweries 1 and 3 were similar across all samples, whereas Brewery 2's samples had a larger difference in the three samples of its beers. The yeast populations were similar between all samples for Brewery 1, whereas Breweries 2 and 3 had a larger difference in yeast populations between their three samples of beer <ref name="snauwaert"></ref>. In summary, Brewery 1 (presumed to be Rodenbach) had similar populations of bacteria and yeast across all samples of their beer, whereas Brewery 3 only had similar populations of bacteria and not yeast across all samples of their beer, and Brewery 2 had the least amount of similarity in yeast and bacteria populations between their beer samples.
Overall, the samples were mostly dominated by ''Pediococcus damnosus'' and ''Brettanomyces bruxellensis'', except for two samples of Brewery 2 that were dominated by ''Acetobacter'' (smaller amounts of ''Acetobacter'' in Brewery 1 were identified as ''Acetobacter pasteurianus'', and might be the same species in Brewery 2), and one sample from Brewery 2 and two samples from Brewery 3 that were dominated by an unidentified yeast. Note that these findings conflicted with the findings of Martens et al. who identified ''P. parvulus'' as the dominating ''Pediococcus'' species. These two species are closely related, and an explanation of this discrepancy between the two findings was never found. ''Acetobacter'' numbers were higher in Brewery 1 and 2, and lower in Brewery 3, and were associated with the high acetic acid amounts found in the beers. Small numbers of ''Lactobacillus'' were found in all samples. ''Weissella'' and ''Leuconostoc'' were found in some samples from Brewery 2, whereas only trace amounts of ''Wessella'' were found in one samples from Brewery 3. A significant amount of ''Pichia'' yeast were found in two Brewery 2 samples, and trace amounts in other samples, as well as trace amounts of ''Candida'' and ''Kregervanrija'' yeasts <ref name="snauwaert"></ref>.
====Metabolic Composition====
Snaewaert et al. also looked at the metabolic composition of finished beers in the three breweries. Glucose was completely gone from the samples, but substantial concentrations of maltose, matotriose, tetraose, pentaose, and hexaose were still present. This contrasts with the relatively high glucose and fructose found in finished bottled versions of these beers, which indicates that the beers are back-sweetened with young beer or with residual sugar or even possibly just some form of sugar at bottling time <ref name="snauwaert"></ref>.
Across the samples there was a presence of isoamyl alcohol (31-150mg/L) and isoamyl acetate (1.99-6 mg/L), and an absence of 2-phenyl ethanol and 2-phenylethyl acetate in both the matured beers and the bottles versions of those beers.
==See Also==