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Quality Assurance

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Any microorganism that is introduced into a beer unintentionally and can survive in the beer is considered a "beer spoiler". One survey of 38 craft beers in the Spanish market found that 68% of them had some presence of unwanted microbes, with beers under 5% ABV being more susceptible than higher ABV beers, indicating that craft breweries in particular may have a high degree of contamination issues <ref>[https://www.jmbfs.org/issue/june-july-2020-vol-9-no-6/jmbfs_2132_garcia-lopez/?issue_id=7366&article_id=25 CONTAMINANT MICROBIOTA IN CRAFT BEERS. Marta García López, Elena Rocheb, Encarnación Rodríguez. 2020.]</ref>. While most microorganisms cannot survive in beer due to the hops, low pH, alcohol content, relatively high carbon dioxide, and shortage of nutrients, certain species are considered to be beer spoilage organisms due to their ability to adapt to brewing conditions (namely hops, ethanol, and low pH) and sometimes form biofilms that help them resist cleaning. Some are able to survive in beer and make a potential impact on the beer's flavor by producing acidity, phenols, turbidity/ropiness via exopolysaccharides (EPS), and/or super-attenuation (which can cause gushing or in extreme cases exploding bottles/cans) with just a few surviving cells. These effects can sometimes manifest days or even weeks after packaging, and longer storage or non-refrigerated storage can increase the potential for beer spoilers to negatively impact the beer. Bacteria species that have adapted to the brewing environment tend to be hop tolerant, but strains of the same species found outside of breweries are not tolerant of brewing conditions. It is thought that these species evolved to carry the genes to adapt to brewing conditions during the 5th to 9th centuries when hops were first being used in brewing, and that this evolution gave them a specialized adaption to the brewing environment where few competitors can survive <ref name="Suzuki_2012">[https://onlinelibrary.wiley.com/doi/abs/10.1002/j.2050-0416.2011.tb00454.x 125th Anniversary Review: Microbiological Instability of Beer Caused by Spoilage Bacteria. Ken Suzuki. 2012. DOI: https://doi.org/10.1002/j.2050-0416.2011.tb00454.x]</ref>. Hop tolerant lactic acid bacteria have been found on the surfaces of many places in the brewing environment, including the fermentation area, bottling area, and cold storage. Hop tolerant lactic acid bacteria have been isolated from the air in at least one brewery in the fermentation and bottling areas <ref>[https://www.tandfonline.com/doi/abs/10.1094/ASBCJ-2017-4294-01?src=recsys Distribution of Lactobacillus and Pediococcus in a Brewery Environment. Jorge Hugo Garcia-Garcia, Luis J. Galán-Wong, Benito Pereyra-Alférez, Luis C. Damas-Buenrostro, Esmeralda Pérez, and Juan Carlos Cabada. 2017. DOI: https://doi.org/10.1094/ASBCJ-2017-4294-01.]</ref>.
Species of yeast and bacteria that are considered beer spoilers include [[Brettanomyces|''Brettanomyces'']] species, numerous [[Lactobacillus|''Lactobacillus'']] species, ''Pediococcus damnosus'', ''Pectinatus'' species (anaerobe responsible for 20-30% of bacterial contaminations that produces acetic acid, [https://en.wikipedia.org/wiki/Propionic_acid propionic acid], acetoin, and 'rotten egg' like odors in contaminated beer), ''Megasphaera cerevisiae'' (7% of bacterial contaminations; inhibited below pH 4.1 and 2.8% ABV but can produce considerable amounts of [[Butyric Acid|butyric acid]] along with smaller amounts of acetic acid, caproic acid, [[Isovaleric Acid|isovaleric acid]], acetoin, and hydrogen sulphide), ''Selenomonas lactifex'', ''Zymophilus'' spp., [[Saccharomyces#Diastatic_strains_of_Saccharomyces_cerevisiae|diastatic strains of ''Saccharomyces cerevisiae'' var. ''diastaticus'']], and some species from the ''Candida'' and ''Pichia'' genera. Most wild yeasts that can grow in beer in lab conditions are not considered largely impactful because of their limitatons to growing in the presence of ethanol or lack of oxygen, but they can become impactful on barrel aged beers where oxygen is present (''Candida'' species, ''Pichia'' species, ''Torulaspora delbrueckii'', ''Issatchenkia orientalis'', ''Kluyveromyces marxianus'', ''Debaryomyces hansenii'', ''Zygosaccharomyces bailii'', ''Zygosaccharomyces bisporus'', ''Schizosaccharomyces pombe'', and ''Kloeckera apiculata'') <ref name="Bokulich_2013">[http://mmbr.asm.org/content/77/2/157.full The Microbiology of Malting and Brewing. Nicholas A. Bokulich and Charles W. Bamforth. 2013. DOI: 10.1128/MMBR.00060-12]</ref>. Hop tolerant lactic acid bacteria make up the majority of contamination issues in breweries, with ''L. brevis'' making up more than half of the reported contaminations, and all lactic acid bacteria making up 60-90% of reported contaminations. A new species of ''Lactobacillus'' was recently identified called ''L. acetotolerans'' and was [https://www.facebook.com/groups/MilkTheFunk/permalink/1363048380390039/ responsible for contaminating Goose Island's Bourbon County Stout], which is 60 IBU and 11% ABV. In sour beers with a pH below 4.3, only some lactic acid bacteria, ''Brettanomyces'', and some wild ''Saccharomyces cerevisiae'' strains (which sometimes produce phenols, haze, over-attenuation, and/or over-carbonation) have the potential for unwanted growth, while beers with low alcohol, a small amount of hops, lower CO<sup>2</sup> volumes (cask ales and beers dispensed with nitrogen, for example), and higher pH (4.4-4.6) are the most susceptible to contamination <ref name="Vaughan_2005">[https://onlinelibrary.wiley.com/doi/full/10.1002/j.2050-0416.2005.tb00221.x Enhancing the Microbiological Stability of Malt and Beer — A Review. Anne Vaughan, Tadhg O'Sullivan, Douwe Van Sinderen. 2005. DOI: https://doi.org/10.1002/j.2050-0416.2005.tb00221.x.]</ref>. ''Pectinatus'' and ''Megasphaera'' are Gram-negative anaerobic species that produce a number of off-flavors in ales but not lagers (probably due to their preference for warmer temperatures). They are somewhat tolerant of hops (they can grow in beers with IBU's as high as 33-38 with one strain isolated from pickles reported to grow in beer up to 5% ABV and 80 IBU <ref>[https://www.sciencedirect.com/science/article/pii/S0740002020300514 Comparative genetic and physiological characterisation of Pectinatus species reveals shared tolerance to beer-associated stressors but halotolerance specific to pickle-associated strains. Timo Kramer, Philip Kelleher, Julia van der Meer, Tadhg O’Sullivan, Jan-Maarten A.Geertman, Sylvia H. Duncan, Harry J. Flint, Petra Louis. 2020. DOI: https://doi.org/10.1016/j.fm.2020.103462.]</ref>) and often survive within the biofilms of other species in the brewing environment where the biofilm creates an anaerobic environment for them. They are sometimes found contaminating low ABV beers (under 5.2%) during packaging. They are not tolerant of pH below 4 and are killed at relatively low temperatures (58–60°C for one min) <ref name="Suzuki_2012" />. ''Zymomonas mobilis'' is a microaerophilic Gram-negative acetic acid bacteria that can withstand hops and can grow in bottled beer or casks where priming sugar is added and small amounts of air is present and produces high levels of acetaldehyde and hydrogen sulphide <ref name="Vaughan_2005" />. While the Gram-positive ''Staphylococcus xylosus'' bacteria, which grows on the skin of humans and animals, is not normally considered a beer contaminant, one strain was isolated from craft beer and was identified as the cause of increased turbidity, lactic acid, and succinic acid. It could grow at a pH between 3-7 (although only produced turbidity at a pH of 4-7), a temperature between 4–37°C, and as much as 8% ABV. This demonstrates that it is possible for species to adapt to living in beer other than the more typical beer spoilers <ref>[https://onlinelibrary.wiley.com/doi/pdf/10.1002/fsn3.1256 Beer‐spoilage characteristics of Staphylococcus xylosus newly isolated from craft beer and its potential to influence beer quality. Zhimin Yu, Qiuying Luo, Li Xiao, Yumei Sun, Rong Li, Zhen Sun, Xianzhen Li. 2019. DOI: 10.1002/fsn3.1256.]</ref>.
Other species of microbes do not grow in beer but can become contaminants earlier on in the brewing process prior to primary fermentation (for example during kettle souring). These species include enterobacteria such as species of ''Clostridium'', ''Obesumbacterium'', ''Citrobacter'', and ''Rahnella aquatilis'', and wild ''Saccharomyces'' species that might not be able to grow in finished beer. These microbes produce [[Dimethyl Sulfide|dimethyl sulfide]], organic acids, and 2,3-butanediol in high amounts which gives the beer an unpleasant rotting fruity or vegetal aroma <ref name="Bokulich_2013" />. Other species are considered "indicator" species because they do not directly cause spoilage of beer, but indicate that there is a hygiene problem. These include ''Acetobacter'', ''Gluconobacter'', ''Escherichia '', and ''Klebsiella'', as well as aerobic yeasts, all of which usually don't have an impact when present unless oxygen is also present. They can also produce slime that protects other microorganisms that can have a greater impact on the beer's stability <ref name="Wirtanen_2001">[https://www.researchgate.net/publication/273439407_Disinfectant_testing_against_brewery-related_biofilms. Disinfectant testing against brewery-related biofilms. Erna Storgårds, Gun Wirtanen. 2001.]</ref><ref name="Bokulich_2018">[https://www.tandfonline.com/doi/abs/10.1094/ASBCJ-2012-0709-01 A Review of Molecular Methods for Microbial Community Profiling of Beer and Wine. Nicholas A. Bokulich, Charles W. Bamforth & David A. Mills. 2018.]</ref>. Some species can contaminate yeast pitches. ''Pediococcus damnosus'' is frequently the cause of such contaminations and can cause diacetyl problems, as well as ''Pediococcus inopinatus'', ''Pediococcus claussenii'', ''L. casei'', ''Selenomonas lacticifex'', and ''Zymophilus raffinosivorans'' (although these are rarer to find in finished beer). ''Obesumbacterium proteus'' (which gives a parsnip-like smell and flavor) and ''Rahnella aquatilis'' can contaminate yeast pitches, and can inhibit fermentation and result in the beer finishing at a higher pH <ref name="Vaughan_2005" />.
* Clean first using an effective cleaner, and then apply a disinfectant/sanitizer as a second step.
* Use 180°F (82°C) hot water for 60 minutes to disinfect stainless steel and other heat tolerant materials (check with your manufacturer to make sure that the vessel is rated to withstand fast hot/cold cycles; vacuum or pressure relief valves should be used in order to avoid imploding due to fast temperature shifts for some equipment).
* For any plastics that cannot be treated with heat, especially tubing, keep separate plastics for use with potential contaminants such as ''[[Lactobacillus]]'', ''[[Pediococcus]]'', ''[[Brettanomyces]]'', and ''[[Saccharomyces#Diastatic_strains_of_Saccharomyces_cerevisiae|diastatic strains of ''Saccharomyces cerevisiae'' var. ''diastaticus]]''.
* Replace rubber and plastic parts such as gaskets as often as recommended by the manufacturer or when wear is apparent.
* When operating a commercial brewery, invest in a quality control lab and procedures to identify inefficient hygiene practices.
Several generalized procedures are used for limiting the number of unwanted microorganisms. These include acid washing yeast that is re-pitched (kills bacteria but not wild yeast that is tolerant of low pH), keeping beer cool (slows the growth of microbes in general), filtration (removes yeast), pasteurization (kills vegetative cells in the finished beer, but not spores - most beer spoilers are killed at 15 [http://wiki.zero-emissions.at/index.php?title=Pasteurization_in_beer_production pasteurization units (PU)] and all are killed at 30 PU using a recommended pasteurization temperature of 66°C ), and aseptic or hygienic packaging. Packaging systems should be frequently flooded with hot water between 80-95°C or saturated steam (every 2 hours in the summer and every 4 hours in the winter). UV light or disinfecting chemicals are also used. The filler and crowner should be disinfected frequently as well. Packaging in an aseptic room with HEPA filtration and higher air pressure within the room compared to outside, along with special clothing, is another method that larger breweries use to remain aseptic <ref name="storgards_2000" />.
Most brewing equipment should be designed for good hygiene. Pits and crevices should be avoided, and all surfaces should be smooth when possible. All equipment and pipelines should be self-draining. Valves are a typical source of contamination because they are not easily CIP'ed, especially plug valves and ball valves (although butterfly, gate, and globe valves are also difficult to CIP) <ref name="storgards_2000" />. Horizontal surfaces and wet surfaces are more prone to biofilm formation. In one study that compared biofilm formation in bottling lines versus canning lines, it was found that canning lines develop less microbial biofilms and contaminations than bottling lines due to not having rinsing stations, labeling stations, and simpler constructions than the bottling lines that were studied <ref name="Storgårds_2006" />. However, some canning lines cannot use caustic for cleaning, or it is not common practice, but use foaming agents instead which are less effective at removing biofilms (see [[Quality_Assurance#Efficacy_of_Cleaning_Agents|efficacy of cleaning agents below]]). The lack of use of caustic cleaners in canning lines has been identified as a source of contamination issues with [[Saccharomyces#Diastatic_strains_of_Saccharomyces_cerevisiae|diastatic strains of ''Saccharomyces cerevisiae'' var. ''diastaticus'']] in canning lines <ref>[https://www.facebook.com/groups/MilkTheFunk/permalink/1561762887185253/?comment_id=1791471917547681&reply_comment_id=2017381731623364&comment_tracking=%7B%22tn%22%3A%22R9%22%7D Caroline Smith from Lallemand. Milk The Funk Facebook group post on diastaticus contamination. Feb 2018.]</ref>.
Some other methods have been proposed by scientists as being novel ways to reduce unwanted microorganisms. These include exploiting naturally produced toxins. For example, some lactic acid bacteria [[Lactobacillus#Bacteriocins|produce bacteriocins]] which can kill other bacteria. Some strains of wine yeast can [[Saccharomyces#Killer_Wine_Yeast|produce zymocins]] that kill other species of yeast. Such methods are viewed as being fairly extreme. Advances in genetic engineering techniques make these approaches technically possible, however, there currently exists a commercial stigma against genetic modification. Additionally, there are many types of toxins which target only specific species, so anticipating which species should be targeted could be challenging <ref name="Vaughan_2005" />.
A popular alternative to Star San is iodophor, which is an iodine-based sanitizer.
[http://masterbrewerspodcast.com/096-efficacy-of-sanitizers-in-the-brewery Elliot Parcells & Josh Pohlmann from Bells Brewery] tested the efficiency of various sanitizers to kill ''Lactobacillus'', ''Brettanomyces'', and diastatic strains of ''Saccharomyces cerevisiae'' var. ''diastaticus'', and claimed that iodophor was ineffective at concentrations of 25 ppm, which is the maximum concentration of iodophor for it to be considered a no-rinse sanitizer. The methodology of this experiment has received some criticism that might invalidate its results. They diluted yeast/bacteria cultures on a 1:10 ratio into iodophor, thus diluting the iodophor below its recommended dilution and exposing it to concentrations of microbes far beyond what it was designed to kill. Testing the ability to sterilize a bulk solution does not translate to the ability of the sanitizer to sanitize trace surface microbes after proper cleaning <ref>[https://www.facebook.com/groups/MilkTheFunk/permalink/2172245389470330/?comment_id=2172685769426292&comment_tracking=%7B%22tn%22%3A%22R%22%7D Dr. Bryan Heit. Milk The Funk Facebook thread on an MBAA podcast about the efficacy of iodohor as a sanitizer. 07/10/2018.]</ref>.
'''Five Star PBW'''
* [https://www.mbaa.com/publications/tq/tqPastIssues/2020/Pages/TQ-57-1-0222-01.aspx MBAA TQ article, "​Building a Quality Control Lab: An Introduction to Starting and Growing a Quality Program" by Melissa Antone.]
* [https://file.scirp.org/pdf/AiM_2016032215023444.pdf Review of Gram negative bacteria in brewing.]
* [http://suigenerisbrewing.com/index.php/2019/12/04/contamination-detection-3/ Dr. Bryan Heit of Sui Generis blog explains his PCR method for detecting diastaticus diastatic yeast.]
* Books:
** [https://my.mbaa.com/ItemDetail?iProductCode=52206 "Illustrated Guide to Microbes and Sediments in Wine, Beer and Juice" by George G. Edwards.]

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