Changes

[[File:Jester King aged hops.jpg|thumbnail|300|[https://web.archive.org/web/20180204131402/http://jesterkingbrewery.com/home-for-our-aged-hops Jester King Brewery in Austin, Texas aging their own hops in an old barn. Image provided by Jeffrey Stuffings.]]]

The primary '''alpha Alpha acids''' (also called "humulones" and abbreviated as "α-acids") in hops are mostly consist of humulone, cohumulone, and adhumulone. Trace amounts of other forms of humulones are also present but are difficult to quantify and currently have limited research: posthumulone, perhumulone, adprehumulone, and acetohumulone <ref name="Hao_2020">[https://www.tandfonline.com/doi/full/10.1080/03610470.2020.1712641 Junguang Hao, R.A. Speers, Heliang Fan, Yang Deng & Ziru Dai (2020) A Review of Cyclic and Oxidative Bitter Derivatives of Alpha, Iso-Alpha and Beta-Hop Acids, Journal of the American Society of Brewing Chemists, 78:2, 89-102, DOI: 10.1080/03610470.2020.1712641.]</ref><ref name="Leker_2022">[https://www.tandfonline.com/doi/abs/10.1080/03610470.2022.2079944 Jeremy Leker & John Paul Maye (2022) Discovery of Acetohumulone and Acetolupulone a New Hop Alpha Acid and Beta Acid, Journal of the American Society of Brewing Chemists, DOI: 10.1080/03610470.2022.2079944 ]</ref>. The ratio of these individual acids to each other can vary based on hop variety much like total iso-α-acid percent, though generally the primary acids are humulone and cohumulone. Cohumulone has been identified by some researchers as a source of a more harsh bitterness, although similar research contradicts this statement <ref>[http://www.scielo.br/scielo.php?pid=S0100-40422000000100019&script=sci_arttext&tlng=es Fundamentals of beer and hop chemistry. Denis De Keukeleire. 1999.]</ref>. While Being hydrophobic, alpha acids are mostly insoluble in wortat typical brewing pH (alpha acids become much more soluble as the pH rises towards 5.9 to 7, which is not typical for wort production <ref name="Bastgen_2019">[https://www.tandfonline.com/doi/full/10.1080/03610470.2019.1587734 Influencing Factors on Hop Isomerization Beyond the Conventional Range. Nele Bastgen, Tobias Becher & Jean Titze. 2019. DOI: https://doi.org/10.1080/03610470.2019.1587734.]</ref>). During boiling, alpha acids are isomerized into iso-alpha acids (also called isohumulones) which are formed during boiling that are soluble. Isomerization leads to roughly a 70%/30% split between diastereomeric isomers called ''cis'' and ''trans'' iso-α-acids respectively, with ''cis'' iso-α-acids being more stable over time and more bitter<ref name="Schönberger and Kostelecky, 2012"> [http://onlinelibrary.wiley.com/doi/10.1002/j.2050-0416.2011.tb00471.x/abstract Schönberger and Kostelecky, 2012]</ref>. Alpha acids themselves do not taste bitter, but isomerized alpha acids (iso-α-acids/isohumulones) contribute to the bitterness of beer and have antimicrobial properties. Isocohumulone is often cited as being more harshly bitter than the other iso-α-acids, but studies of taste perception of individual iso-α-acids have not agreed with this. However isocohumolone , iso-cohumolone is slightly more soluble than the other acids and therefore a hop with a higher cohumulone composition may result in a beer with higher iso-α-acid for hops of equal iso-α-acid percent and use in brewing but different iso-α-acid breakdown<ref name="Schönberger and Kostelecky, 2012"/>. Alpha acids are susceptible to oxidation and the alpha acid content of a hop will decrease with storage.

The isomerization of alpha acids into There is evidence to show that during wort boiling iso-alpha acids is mostly dependent on time humulone and temperatureperhaps also iso-cohumulone bind with the head forming proteins, Lipid Transfer Protein (LTP) and not other variables such as sugar concentrationProtein Z, pH, and calcium concentration <ref>[https://onlinelibrary.wiley.com/doi/pdf/10.1002/j.2050to help form foam-0416.1964.tb06356.x CHANGES IN HOP ACIDS CONCENTRATIONS ON HEATING IN AQUEOUS SOLUTIONS AND UNHOPPED WORTSpositive structures in beer. H. O. Askew. 1964.]</ref><ref>[http://pubs.acs.org/doi/abs/10.1021/jf0481296 Isomerization and Degradation Kinetics of Hop (Humulus lupulus) Acids in a Model WortThese iso-Boiling Systemalpha acids bind less so with LTP than they do with Protein Z. Mark G. Malowicki and Thomas H. Shellhammer. 2005.]</ref>. Significant isomerization of alpha acids can occur in water without sugar at all (temperatures around boiling are still required)The resulting bound structures have been described as "vesicles", which is relevant in the production of [http://www.garshol.priv.no/blog/331.html are protein "hop teabubbles" (but with no gas in traditional farmhouse brewing] where hops are steeped in hot water for some time, and this is said to extract bitterness from the hops them) with thick surface layers <ref>[httphttps://www.ijbbbsciencedirect.org/papers/161-E005.pdf Kinetic Modeling of Hop Acids during Wort Boiling. Yarong Huang, Johannes Tippmann, and Thomas Becker. 2013.]</ref><ref>[http:com/science/www.garshol.priv.noarticle/blogpii/331.html Lars Marius Garshol. "Raw ale". Larsblog. 05/06/2015. Retrieved 12/17/2018.]</ref>. Malowicki S0268005X19325391 Vesicular structures formed from barley wort proteins and Shellhammer determined a calculation that predicts the isomerization rates of alpha acids into iso-alpha acids at different temperatureshumulone. Beginning at the boiling temperature of 100°C/212°FYi Lu, which could be considered a rate of 100%Peter Osmark, at 96°C/205°F the rate is 72%Björn Bergenståhl, and at 90°C/194°F the rate is 43%Lars Nilsson. This rate continues to drop significantly as the temperature of the wort decreases2020. At 82°C/180°F isomerization occurs at a rate of 17%. At a temperature of 50°C/122°F, the isomerization rate is at 1%, and finally 0% at 45°C/113°F. This fact has several impacts on brewing processes. For example, when brewing at higher altitudes where the boiling point of wort is less than 100°C]</212°F, the isomerization of alpha acids into iso-alpha acids will be reduced to whatever the rate is at that lower temperature. "Hop stands" or "whirlpool additions" where hops are left in contact with hot wort that is less than boiling temperature will continue to isomerize alpha acids <ref>. See also [https://www.mbaayoutube.com/publications/tq/tqPastIssues/2017/Pages/TQ-54-3-0806-01.aspx A Look at Isomerization Reduction Due to Altitude. John Palmer. MBAA TQ 2017 http://dx.doi.org/10.1094/TQ-54-3-0806-01.watch?v=5F8vmuTV5Mg Escarpment Labs presentation on the science of beer foam]</ref>.

'''Beta Acids''' (lupulones) are similar in structure to alpha acids and have the analogous individual beta acids (lupulone, colupulone, adlupulone, prelupulone, postlupulone, adprelupukone, and postlupulone acetolupulone <ref name="Dušek_2014">[http://pubs.acs.org/doi/abs/10.1021/jf501852r Qualitative Determination of β‑Acids and Their Transformation Products in Beer and Hop Using HR/AM-LC-MS/MS. Martin Dušek, Jana Olšovská, Karel Krofta, Marie Jurková, and Alexandr Mikyška. 2014.]</ref><ref name="Hao_2020" /><ref name="Leker_2022" />) to individual alpha acids. In their original form, beta acids do not contribute to the flavor of beer because they are not soluble in beerunless the pH of the boiling wort is significantly raised to around 7 pH (which is not typical in brewing conditions) and the original gravity is relatively low (2-8°P) <ref name="Bastgen_2019" />. They are also not able to isomerize during wort boiling. Beta acids do not become soluble in wort or beer unless they are chemically modified by a process such as oxidation <ref name="Algazzali_2014" />, nor are they soluble in beer when dry hopping <ref name="Maye_EBC2017">John Paul Maye. EBC 2017 Presentation. 2017.]</ref>. Oxidized beta acids are soluble and can contribute to bitterness in beer. Oxidized beta acids are discussed more under [[Hops#Acids_2|aged hops]]. ====Isomerization of Alpha Acids==== The isomerization of alpha acids into iso-alpha acids is mostly dependent on alpha acid content of the hops, time (to a certain extent), temperature, original gravity, hop rate (hop weight), and IBU saturation. Other variables also affect isomerization to a lesser extent such as pH and calcium concentration <ref>[https://onlinelibrary.wiley.com/doi/pdf/10.1002/j.2050-0416.1964.tb06356.x CHANGES IN HOP ACIDS CONCENTRATIONS ON HEATING IN AQUEOUS SOLUTIONS AND UNHOPPED WORTS. H. O. Askew. 1964.]</ref><ref name="Malowicki_2005">[http://pubs.acs.org/doi/abs/10.1021/jf0481296 Isomerization and Degradation Kinetics of Hop (Humulus lupulus) Acids in a Model Wort-Boiling System. Mark G. Malowicki and Thomas H. Shellhammer. 2005.]</ref><ref name="justice_2018">[https://www.mbaa.com/publications/tq/tqPastIssues/2018/Pages/TQ-55-3-1205-01.aspx Tracking IBU Through the Brewing Process: The Quest for Consistency. Aaron Justus. Director of R&D and Specialty Brewing, Ballast Point Brewing. MBAA TQ 2018; vol. 55, no.3. https://doi.org/10.1094/TQ-55-3-1205-01.]</ref>. The higher the gravity of wort above 1.050 SG, the more proteins coagulate and drop iso-alpha acids out of solution (lower gravity worts are not affected by this). During fermentation, yeast cells can absorb iso-alpha acids, which results in further loss of iso-alpha acids in the finished beer <ref name="Bastgen_2019" />. Lower flocculating yeast strains tend to reduce the IBU in finished beer more than high flocculating yeast <ref name="justice_2018" />. Significant isomerization of alpha acids can occur in water without sugar at all (temperatures around boiling are still required), which is relevant in the production of [http://www.garshol.priv.no/blog/331.html "hop tea" in traditional farmhouse brewing] where hops are steeped in hot water for some time, and this is said to extract bitterness from the hops <ref name="Malowicki_2005" /><ref>[http://www.ijbbb.org/papers/161-E005.pdf Kinetic Modeling of Hop Acids during Wort Boiling. Yarong Huang, Johannes Tippmann, and Thomas Becker. 2013.]</ref><ref>[http://www.garshol.priv.no/blog/331.html Lars Marius Garshol. "Raw ale". Larsblog. 05/06/2015. Retrieved 12/17/2018.]</ref>. Aside from boiling hops in wort, [https://www.frontiersin.org/articles/10.3389/fnut.2022.843808/full Hydrodynamic Cavitation] at a 90°C temperature is another method that results in the isomerization of alpha acids into iso-alpha acids; although an additional 10 minutes of boiling at 100°C was needed to remove DMS and achieve hot break. This method reportedly had a 33% savings in energy costs in [https://www.sciencedirect.com/science/article/abs/pii/S0960308524000166 one study].  Malowicki and Shellhammer determined a calculation that predicts the isomerization rates of alpha acids into iso-alpha acids at different temperatures. Beginning at the boiling temperature of 100°C/212°F, which could be considered a rate of 100%, at 96°C/205°F the rate is 72%, and at 90°C/194°F the rate is 43%. This rate continues to drop significantly as the temperature of the wort decreases. At 82°C/180°F isomerization occurs at a rate of 17%. At a temperature of 50°C/122°F, the isomerization rate is at 1%, and finally 0% at 45°C/113°F (note that [https://www.tandfonline.com/doi/full/10.1080/03610470.2021.1878684 Kishimoto et al. (2021)] reported that isomerization begins somewhere between 60-70°C <ref name="Kishimoto_2021">[https://www.tandfonline.com/doi/full/10.1080/03610470.2021.1878684 Toru Kishimoto, Satoko Teramoto, Akiko Fujita & Osamu Yamada (2021) Evaluation of Components Contributing to the International Bitterness Unit of Wort and Beer, Journal of the American Society of Brewing Chemists, DOI: 10.1080/03610470.2021.1878684.]</ref>). This fact has several impacts on brewing processes. For example, when brewing at higher altitudes where the boiling point of wort is less than 100°C/212°F, the isomerization of alpha acids into iso-alpha acids will be reduced to whatever the rate is at that lower temperature. "Hop stands" or "whirlpool additions" where hops are left in contact with hot wort that is less than boiling temperature will continue to isomerize alpha acids <ref name="Malowicki_2005" /><ref>[https://www.mbaa.com/publications/tq/tqPastIssues/2017/Pages/TQ-54-3-0806-01.aspx A Look at Isomerization Reduction Due to Altitude. John Palmer. MBAA TQ 2017 http://dx.doi.org/10.1094/TQ-54-3-0806-01.]</ref>.  The hopping rate (weight) and IBU saturation have a drastic effect on IBU's. Aaron Justice reported a higher utilization when using Polaris hops (17.6% alpha acids) versus Tettnang hops (1.9% alpha acids). Two beers were brewed using enough of each of the hop varieties to target a calculated 40 IBU. However, the beer brewed with Polaris hops had 42.6 IBU and the beer brewed with Tettnang hops had only 28 IBU. It was hypothesized that the higher surface area of more hop matter reduces IBU's by binding to iso-alpha acids and possibly other compounds that register on the standardized IBU test. Justice also reported that more IBU pickup from whirlpooling was possible in beers without hops added in the boil, indicating that IBU saturation can limit IBU's, with a maximum IBU being around 100 <ref name="justice_2018" />. The pH of the wort appears to have no significant effect on the isomerization of alpha acids to iso-alpha acids. Aaron Justice reported a trending slight rise in the conversion of alpha acids to iso-alpha acids when the boil pH was raised from 5.05 (~40% of aa's converted to iso-aa's) to 5.35 (~50% of aa's converted to iso-aa's). However, an Eppendorf BioSpectrometer using ASBC Wort-23 and ASBC Beer-23A was used to measure IBU'S, not isomerized alpha acids specifically (chromatography must be used to measure iso-alpha acids without measuring other bittering compounds) <ref name="justice_2018" />. Bastgen et al. (2019) found that at a boil pH of 5.6, the percentage of iso-alpha acids increased by 32% by extending the boil from 60 minutes to 120 minutes. However, there was no increase at all in iso-alpha acids when the boil pH was 7, but a pH of 7 is not typical in the brewing process <ref name="Bastgen_2019" />. Time is thought to play a large role in isomerization, however, Justice reported that the majority of the IBU from iso-alpha acids in 60 minute additions and in whirlpool additions occurs within the first 10 minutes, with only a 12-30% increase after another 50 minutes of boiling/whirlpooling (higher gravity beers had more isomerization during the final 50 minutes while lower gravity beers had less isomerization during the final 50 minutes of boiling) <ref name="justice_2018" />. Blogger and "garage scientist", John Paul Hosom, who developed the so-called "[https://alchemyoverlord.wordpress.com/2021/11/10/ibus-and-the-smph-model/ SMPH Model for estimating IBU's], hypothesized that non-iso-alpha acid bittering compounds that impact spectrophotometer IBU measurements such as oxidized alpha acids are affected by wort pH, rather than the isomerization rate of alpha acids into iso-alpha acids <ref>[https://alchemyoverlord.wordpress.com/2021/11/10/ibus-and-the-smph-model/ John Paul Hosom. "The Effect of pH on Utilization and IBUs". Alchemyoverlord blog. Retrieved 01/29/2023.]</ref>. See also:* [http://www.thebrewingnetwork.com/brew-strong-tracking-ibu/ Brew Strong podcast; "Tackling the IBU" with Aaron Justice.]

Hops are known to have antimicrobial properties against Gram-positive bacteria. This includes bacteria which that can be present in beer both as spoilage organisms and as intentionally added in sour and mixed fermentation beer such as ''[[Lactobacillus]]'' and ''[[Pediococcus]]''. Certain other Gram-negative bacteria found in beer , such as ''Acetobacteraciae'' are Gram-negative , and are not susceptible to the antimicrobial properties of hops<ref name="Hough_1957">[https://onlinelibrary.wiley.com/doi/pdf/10.1002/j.2050-0416.1957.tb06267.x J. S. Hough, B.Sc, Ph.D., G. A. Howard, M.Sc., Ph.D., and C. A. Slater, Ph.D. 1957.]</ref><ref name="Macrae_1964">[https://onlinelibrary.wiley.com/doi/abs/10.1002/j.2050-0416.1964.tb02001.x SIGNIFICANCE OF THE USE OF HOPS IN REGARD TO THE BIOLOGICAL STABILITY OF BEER: I. REVIEW AND PRELIMINARY STUDIES. R. M. Macrae. 1964.]</ref>. Certain Gram-positive bacteria strains that have adapted to the brewing environment, such as some strains of ''Lactobacillus brevis'' and ''L. paracasei'' <ref>[https://proceedings.science/slacan-2023/papers/prospects-for-the-development-of-a-new-hopped-and-functional-sour-beer-survival?lang=en Marcos Edgar Herkenhoff; Susana Marta Isay Saad. PROSPECTS FOR THE DEVELOPMENT OF A NEW HOPPED AND FUNCTIONAL SOUR BEER: SURVIVAL OF PROBIOTIC STRAINS OF LACTICASEIBACILLUS PARACASEI SUBSP. PARACASEI IN HIGH HOPPED BEERS (HUMULUS LUPULUS L.).. In: CADERNO DE RESUMOS DO 15° SLACAN - SIMPóSIO LATINO AMERICANO DE CIêNCIA DE ALIMENTOS E NUTRIçãO, 2023, Campinas. Anais eletrônicos... Campinas, Galoá, 2023. Disponível em: <https://proceedings.science/slacan-2023/trabalhos/prospects-for-the-development-of-a-new-hopped-and-functional-sour-beer-survival?lang=en> Acesso em: 23 nov. 2023.]</ref>, are known to be more resistant to the antimicrobial effects of hops. Multiple mechanisms The antimicrobial effect is characterized as inhibiting the growth and lactic acid production of lactic acid bacteria, however, this does not always also include cell death as ''Lactobacillus'' that has been inhibited by hops can later be revived <ref name="Macrae_1964" />. The effectiveness of hops to inhibit Gram-positive bacteria is also dependent on pH; at a lower pH, hops have a greater effect on inhibiting bacteria <ref name="Almaguer_2015" />. Hop extracts have also been proposed demonstrated to explain why hops are antimicrobially activebe antimicrobial <ref>[https://www.sciencedirect.com/science/article/abs/pii/S0963996923003770 Yan Li, Sevim Dalabasmaz, Sabrina Gensberger-Reigl, Marie-Louise Heymich, Karel Krofta, Monika Pischetsrieder. Identification of colupulone and lupulone as the main contributors to the antibacterial activity of hop extracts using activity-guided fractionation and metabolome analysis. Food Research International. 2023.]</ref>.

Multiple mechanisms have been proposed to explain why hops are antimicrobially active. One mechanism of the antimicrobial activity of hops is due to the role of iso-alpha alpha acids and possibly similar hop acids (such beta acids and iso-α-oxidized hop acids) as ionophores, or compounds which can transport ions across cell membranes. While their antimicrobial properties are strong, alpha and beta acids in beer and wort and their effects on brewing are generally disregarded because they do not solubilize <ref name="Fernandez and Simpson, 1993"> [http://onlinelibrary.wiley.com/doi/10.1111/j.1365-2672.1993.tb02782.x/full Fernandez and Simpson (1993)] </ref> <ref name="Sakamoto and Konings, 2003"> [http://www.sciencedirect.com/science/article/pii/S0168160503001533 Sakamoto and Konings (2003)] </ref>. The protonated iso-α-acid (the form of the acid with an associated H+ ion, an H+ ion is a proton) is the antimicrobially active form. This means that for a beer with a given iso-α-acid concentration, the antimicrobial effects will be stronger at lower pH values because a greater percentage of the acid will be protonated. Protonated iso-α-acids act against bacteria by crossing into the cell and dissociating (releasing H+ ions from the iso-α-acidand decreasing the pH within the cell <ref name="zhao_1027">[https://www.frontiersin.org/articles/10.3389/fmicb.2017.00239/full#B28 Heterogeneity between and within Strains of Lactobacillus brevis Exposed to Beer Compounds. Yu Zhao, Susanne Knøchel and Henrik Siegumfeldt. 2017. DOI: https://doi.org/10.3389/fmicb.2017.00239.]</ref>), therefore disrupting the cellular proton gradient which is necessary for cells to function, before binding an equal charge in metal ions and crossing back out of the cell. Cells with a resistance to hop bitter acids are better able to eject disassociated iso-α-acids from the cell and therefore preserve their proton gradients. The mechanism to expel iso-α-acids appears to be specific toward this type of compound rather than by a more general antimicrobial resistance mechanism such as multi-drug resistant bacteria possess<ref name="Sakamoto and Konings, 2003"/>. The anti-microbial power of iso-α-acids is pH dependent. At a higher pH (5.6) iso-α-acids begin to lose their anti-microbial properties, but at a typical beer pH (4.3) iso-α-acids inhibited a sample of 6 strains of ''L. brevis'' that exhibited a range of general hop tolerance in one study <ref name="zhao_1027" />. Hop resistant bacteria cultured in the absence of hop acids can lose their resistance if grown in an environment without antibacterial hop compounds<ref name="Fernandez and Simpson, 1993"/> and some hop resistant microbes need to be acclimated to hop acids by growth in sub-limiting levels of antibacterial acids before they are able to resist higher levels<ref name="Sakamoto and Konings, 2003"/>.

Another antimicrobial mechanism resulting from oxidative stress has been attributed to both iso-α-acids and humulinic acids<ref name="Schurr et al, 2015"> [http://www.sciencedirect.com/science/article/pii/S0740002014002470 Benjamin C. Schurr et al, Hannes Hahne, Bernhard Kuster, Jürgen Behr, Rudi F. Vogel. Molecular mechanisms behind the antimicrobial activity of hop iso-α-acids in Lactobacillus brevis.Food Microbiology, Volume 46, (2015), Pages 553-563, ISSN 0740-002., https://doi.org/10.1016/j.fm.2014.09.017.] </ref>. Humulinic acids are either not bitter tasting or much less bitter than iso-α-acids but are similar in structure to and are formed from the degradation of iso-α-acids as well as during the aging of hops<ref>[https://www.sciencedirect.com/science/article/abs/pii/S0040402001981992 The absolute configuration of the isohumulones and the humulinic acids. D.De Keukeleire, M.Verzele. 1971. https://doi.org/10.1016/S0040-4020(01)98199-2.]</ref>. Humulinic acids consist of the "cis-" and" trans-" forms of humulinic acid, cohumulinic acid and adhumulinic acid <ref>[https://www.sciencedirect.com/science/article/pii/B9781855734906500088 "8 - The chemistry of hop constituents," Editor(s): Dennis E. Briggs, Chris A. Boulton, Peter A. Brookes, Roger Stevens, In Woodhead Publishing Series in Food Science, Technology and Nutrition, Brewing. Woodhead Publishing, 2004. Pages 255-305. ISBN 9781855734906. https://doi.org/10.1533/9781855739062.255.]</ref>. This oxidative stress-driven antimicrobial activity is due to the potential for oxidation-reduction (redox) reactions within bacterial cells between Mn2+ ions and these specific hop acids. The redox potential is due to different conditions inside (higher pH, higher Mn2+) and outside (lower pH, lower Mn2+) of the bacterial cell<ref name="Behr and Vogel, 2010"> [http://aem.asm.org/content/76/1/142.short Behr and Vogel, (2010)] </ref><ref name="Schurr et al, 2015"/>. Iso-α-acids or humulinic acids passing into the cell, form complexes with Mn2+ and transfer electrons out of the cell<ref name="Behr and Vogel, 2010"/>. By targeted molecular modifications [http://www.sciencedirect.com/science/article/pii/S0740002014002470 Schurr et al. (2015)] determined that the Mn oxidative stress-driven antimicrobial effect of iso-α-acids was more important than the antimicrobial effect of the ionophore proton transfer discussed above in the overall antimicrobial activity of hops. Thus, the antimicrobial effects of humulinic acids have been found to be even stronger than iso-alpha acids, suggesting that aged hops retain at least some antimicrobial properties at least partially from humulinic acids <ref name="Schurr et al, 2015"/>. One study found that adding Mn2+ to a lager beer minimized this effect and slightly increased the cell count of 6 strains of ''L. brevis'' that had a range of hop tolerances <ref name="zhao_1027" />. The oxidized forms of hop acids have been shown to have a limited inhibitory effect on Gram-positive bacteria. This might explain the anecdotal experiences of some brewers that have tried using aged hops that were high alpha varieties and produced beer that wasn't sour. Stevens et al. (1961) reported that a strain of ''Lactobacillus'' that was cultured from infected beer was inhibited by alpha acids at 40 ppm, beta acids at 10 ppm, iso-alpha acids at 160 ppm, and oxidized beta acids (cohulupone) at 200 ppm. So, while the oxidized beta acids had the least inhibitory power, a high concentration was still inhibitory <ref name="Stevens_1961">[https://onlinelibrary.wiley.com/doi/pdf/10.1002/j.2050-0416.1961.tb01830.x R. Stevens, Ph.D., F.R.I.C., and D. Wright, Ph.D. 1961.]</ref>. Oxidized alpha acids (humulinones) have only been tested for antibacterial properties at a concentration of 50 ppm or less. At 50 ppm, oxidized alpha acids were not able to inhibit two strains of ''Lactobacillus'' that were isolated from infected beer, as reported by Hough et al. (1957) <ref name="Hough_1957" />. See [[Hops#Acids_2|oxidized hop acids]] for more information on oxidized hop acids.

* See also :*[[Lactobacillus#Hop_Tolerance|''Lactobacillus'' hop tolerance]].*[[Pediococcus#Hop_Resistance|''Pediococcus'' hop tolerance]]

Due to the multiple mechanisms for hop antimicrobial activity, multiple resistance mechanisms are necessary for a Gram-positive bacterial cell to successfully be hop-tolerant<ref name="Behr and Vogel, 2010"> [http://aem.asm.org/content/76/1/142.short Behr and Vogel, (2010)] </ref>. Hop resistance of bacteria will vary by species as well as within a species with individual strains. The environment in which strains are cultured and maintained may also influence their hop tolerance. It is possible for a small subpopulation of individual cells that have a higher tolerance to hops to eventually dominate the overall population <ref name="zhao_1027" />. The hop tolerance of lactic acid bacteria strains decreases when they are cultured in hop-free environments and strains cultured in media with increasing concentrations of hop compounds show an increase in hop tolerance<ref name="Sakamoto and Konings, 2003"/>. The stability of hop resistance, or the rate at which it is lost when bacteria are cultured in unhopped wort, varies by strain. It can take up to 1 year for maximum loss of hop resistance, suggesting that in some strains have a relatively stable hop resistance<ref name="Sakamoto and Konings, 2003"/>. Because of this intra-species variability and dependence on how the strains were cultured, it is difficult to give specific advice about the hop-tolerance of a wide range of strains offered from different sources. As a general rule, some common lactic acid bacteria species used in sour beer and found as beer spoilage organisms like ''Lactobacillus brevis'', ''Lactobacillus lindneri'' and ''Pediococcus delbrueckii'' have some resistance to hops<ref name="Sakamoto and Konings, 2003"/>. Brewers seeking to make acidic beers with higher doses of hops may wish to seek out one of these species. Some hop-tolerant species benefit from pre-culturing in media with below-limiting concentrations of compounds before being used in more highly hopped wort or beer<ref name="Simpson and Fernandez, 1992"> [http://onlinelibrary.wiley.com/doi/10.1111/j.1472-765X.1992.tb00636.x/abstract Simpson and Fernandez, 1992]</ref>.

The flavor and aroma compounds found in leaf/pellet hops is different than [[File:Svedlund 2022.jpg|thumb|400px|An overview of the hop derived flavor and aroma compounds found in finished beer (other than biotransformation reactions occurring in certain yeast with the case of dry hopping)required genetic/enzymatic capability. The brewing process (particularly boiling)Abbreviations: 3MH 3-mercaptohexanol, and fermentation greatly affect the composition of flavor and aroma compounds that are found in beer. For example3MHA 3-mercaptohexyl acetate, boiling wort and hops isomerizes non-bitter alpha acids into bitter iso-alpha acids. During boiling of the wortCys cysteine, many compounds found in hops are evaporatedGSH glutathione, such as many of the various sulfur compounds found in hopsTPA terpene alcohol. The terpene hydrocarbons which make up most of the hop oil content in hops (myrecene, humulene, and caryophyllene) are completely removed by fermentationCredit: Henrik Svedlund. It is believed that these terpene hydrocarbons stick to the yeast cells and fall out of solution during fermentation <ref name="Praet_2012"> Source: [httphttps://wwwlink.sciencedirectspringer.com/science/article/pii10.1007/S1373716311001636 Biotransformations of hops00253-022-12068-derived aroma compounds by Saccharomyces cerevisiae upon fermentationw Svedlund, N. Tatiana Praet, Filip Van OpstaeleEvering, Barbara Jaskula-GoirisS., Gibson, Guido AertsB. et al. Fruits of their labour: biotransformation reactions of yeasts during brewery fermentation. Appl Microbiol Biotechnol 106, Luc De Cooman4929–4944 (2022). 2012https://doi.]<org/ref>10. 1007/s00253-022-12068-w]]]

Some carbonyl The flavor and aroma compounds found in leaf/pellet hops are different than the hop-derived flavor and aroma compounds found in finished beer (citral, geranial, nerol, [https://enother than in the case of dry hopping).wikipedia.org/wiki/Citronellal citronellal] The brewing process (particularly boiling), and methyl ketones) can be used as a food source by yeast during fermentationgreatly affect the composition of flavor and aroma compounds that are found in beer. ''Cyclic ethers'' such as linalool oxides, karahana ether, hop etherFor example, boiling wort and rose oxide (aroma of roses <ref>[http://www.thegoodscentscompany.com/data/rw1035651.html "(Z)hops isomerizes non-bitter alpha acids into bitter iso-rose oxide "alpha acids. Good Scents Company. Retrieved 12/29/2016.]</ref>)During the boiling of the wort, many compounds found in hops are evaporated, increase after fermentation and have been identified such as secondary metabolites produced by yeast during metabolism from hop derived precursors. ''Esters'' many of the various sulfur compounds found in hops can be converted into ethyl esters by yeast during fermentation; for example geranyl esters found . The terpene hydrocarbons which make up most of the hop oil content in Cascade hops can be hydrolized into geraniol (flowerymyrecene, humulene, and caryophyllene)are completely removed by fermentation. The terpenoid [https://en.wikipedia.org/wiki/Citronellol citronellol] (citrus It is believed that these terpene hydrocarbons stick to the yeast cells and floral fall out of solution during fermentation <refname="Praet_2012">[httpshttp://eicwww.rscsciencedirect.org/magnificent-moleculescom/citronellolscience/2000020.article "There are no flies on Emma Stoye". Emma Stoye. Education in Chemistry website. 06/01pii/2016S1373716311001636 Biotransformations of hop-derived aroma compounds by Saccharomyces cerevisiae upon fermentation. Retrieved 01/10/2017.]</ref>) can be esterified by yeast fermentation into citronellyl acetate (freshTatiana Praet, rosyFilip Van Opstaele, fruity odor reminiscent of geranium oil <ref>[https://shop.perfumersapprentice.com/pBarbara Jaskula-6034-citronellyl-acetate.aspx "Citronellyl acetate"Goiris, Guido Aerts, Luc De Cooman. Perfumers Apprentice website. Retrieved 01/10/20172012.]</ref>). Yeast strains differ in their ability to convert these compounds. For example, one study found that lager yeast was able to form acetate esters of geraniol and citronellol, but ale yeast was not <ref name="Praet_2012" />.

Terpenes A "biotransformation" is any change in a chemical's structure that is initiated by a living organism <ref>[https://en.m.wikipedia.org/wiki/Biotransformation "Biotransformation". Wikipedia. Retrieved 05/10/2019.]</ref>. It has been hypothesized that biotransformations of some kind are taking place in beer during fermentation and explain changes to hop compounds during fermentation and terpenoids beer storage. Some carbonyl compounds found in hops (monoterpene alcoholscitral, geranial, nerol, [https://en.wikipedia.org/wiki/Citronellal citronellal], and methyl ketones) can also be transformed used as a food source by yeast during fermentation. Studies have found that geraniol and nerol can transform into linalool by a strain of ''S. cerevisiaeCyclic ethers''such as linalool oxides, karahana ether, hop ether, as well as nerol and linalool into alpharose oxide (aroma of roses <ref>[http://www.thegoodscentscompany.com/data/rw1035651.html "(Z)-terpineolrose oxide ". Good Scents Company. Retrieved 12/29/2016.]</ref>), which can then increase after fermentation and have been identified as secondary metabolites produced by further transformed to terpinyeast during metabolism from hop derived precursors. Geraniol ''Esters'' found in hops can also be converted into citronellol, and the content of geraniol and citronellol can be increased in finished beer ethyl esters by increasing the initial content of geraniolyeast during fermentation; for example, which is geranyl esters found in higher quantities in some varieties of Cascade hops can be hydrolyzed into geraniol (Citra, for exampleflowery). Linalool, nerol, The terpenoid [https://en.wikipedia.org/wiki/Citronellol citronellol] (citrus and alphafloral <ref>[https://eic.rsc.org/magnificent-terpineol gradually decrease during fermentation and aging (perhaps being transformed into ethers), while nerol and molecules/citronellol gradually increase/2000020.article "There are no flies on Emma Stoye". Emma Stoye. Education in Chemistry website. Geraniol also decreases during fermentation, but not as drastically as linalool06/01/2016. It has been hyptothesized that the bioconversion of geraniol into ctironellol could Retrieved 01/10/2017.]</ref>) can be esterified by means yeast fermentation into citronellyl acetate (fresh, rosy, fruity odor reminiscent of glycosidic activity (although another study found that glycosidic activity in ''Sgeranium oil <ref>[https://shop.perfumersapprentice.com/p-6034-citronellyl-acetate.aspx "Citronellyl acetate". Perfumers Apprentice website. Retrieved 01/10/2017. cerevisiae'' is not very strong]</ref>). Post-fermentation dry hopping preserves linalool and alpha-terpineolYeast strains differ in their ability to convert these compounds. For example, one study found that lager yeast was able to form acetate esters of geraniol and limits citronellol to trace levels , but ale yeast was not <ref name="Praet_2012" />.

In addition to biochemical changes to hop compounds, yeast derived flavor compounds that are independent of hops such as esters and phenols can affect the overall sensory characteristics of hop compounds in beer. Therefore, different strains of yeast can greatly impact the overall organoleptic experience of hop flavors and aromas in finished beers <ref>[https://www.mdpi.com/2304-8158/12/5/1064 Kumar, A.; Warburton, A.; Silcock, P.; Bremer, P.J.; Eyres, G.T. Yeast Strain Influences the Hop-Derived Sensory Properties and Volatile Composition of Beer. Foods 2023, 12, 1064. https://doi.org/10.3390/foods12051064.]</ref>.  ===Terpenes=== Terpenes and terpenoids (monoterpene alcohols) can also be transformed by fermentation. Studies have found that geraniol and nerol can transform into linalool by a strain of ''S. cerevisiae'', as well as nerol and linalool into alpha-terpineol, which can then be further transformed to terpin. Geraniol can also be converted into citronellol, and the content of geraniol and citronellol can be increased in finished beer by increasing the initial content of geraniol, which is found in higher quantities in some varieties of hops (Citra, for example). Linalool, nerol, and alpha-terpineol gradually decrease during fermentation and aging (perhaps being transformed into [https://en.wikipedia.org/wiki/Ether ethers], which is a class of organic compound that contains an oxygen atom connected to two alkyl or aryl groups), while nerol and citronellol gradually increase. Geraniol also decreases during fermentation, but not as drastically as linalool. It has been hypothesized that the bioconversion of geraniol into citronellol could be by means of glycosidic activity (although evidence for glycosidic activity in ''S. cerevisiae'' in regards to hop derived compounds is very weak; see [[Hops#Glycosides|Hop Glycosides]]). Post-fermentation dry hopping preserves linalool and alpha-terpineol, and limits citronellol to trace levels <ref name="Praet_2012" />.  [https://onlinelibrary.wiley.com/doi/abs/10.1002/j.2050-0416.2010.tb00428.x Takoi et al. (2012)] used Citra hops with a high content of geraniol added late in the boil, and reported a steep decline on geraniol during the first three days of fermentation with a lager yeaststrain. Linalool had a gradual decline, but ended up at higher levels than geraniol in the finished beer. Citronellol had a sharp increase during the first three days of fermentation, and then remained at a stable level until the end of fermentation. However, after storing the beer at 15°C (59°F) for 1 week, the amount of citronellol more than doubled. This indicated that active fermentation may not be required for the transformation of geraniol into citronellol (the yeast was filtered before packaging the finished beer, after a storage time of 6-8 days at 13–15°C and then at 0°C for 2–3 weeks). The enzyme NADPH dehydrogenase 2, encoded by the OYE₂ gene in some yeast strains, was proposed as the mechanism for this transformation. Interestingly, Takoi et al. (2012) also showed that coriander seeds, which also have high levels of linalool and geraniol, have a nearly exact same effect on beer, with a beer made with 0.5 g/L of coriander seed resulting in 20 ppb of citronellol and a beer made with 0.75 g/L of coriander seed resulting in 30 ppb of citronellol. The Citra beer had a citrus and "green" aroma, while the coriander beers had a very floral aroma with a slight citrus impression. They also conducted a sensory experiment with different levels of geraniol and citronellol added to linalool to see if small amounts of these would effect affect the flavor of a large dosage of linalool, and the results confirmed that small increases of geraniol and citronellol increased flowery and fruity flavors even in the presence of high dosages of linalool <ref>[https://onlinelibrary.wiley.com/doi/abs/10.1002/j.2050-0416.2010.tb00428.x The Contribution of Geraniol Metabolism to the Citrus Flavour of Beer: Synergy of Geraniol and β‐Citronellol Under Coexistence with Excess Linalool. Kiyoshi Takoi, Yutaka Itoga, Koichiro Koie, Takayuki Kosugi, Masayuki Shimase, Yuta Katayama, Yasuyuki Nakayama, Junji Watari. 2012. DOI: https://doi.org/10.1002/j.2050-0416.2010.tb00428.x.]</ref>. The data for the Citra beer is shown below:

[https://www.researchgate.net/publication/261475199_Screening_of_Geraniol-rich_Flavor_Hop_and_Interesting_Behavior_of_beta-Citronellol_During_Fermentation_under_Various_Hop-Addition_Timings Takoi et al. (2014)] continued their research into monoterpene biotransformations. They determined that some varieties of hops have higher concentrations of geraniol (floral flavor) than others, which when used in beer, can lead to higher citronellol levels (citrus flavor) in beer that wasn't present in the hops or wort. They found that while traditional German hops such as Saaz and New Zealand hops contain very little geraniol, American hops such as Bravo, Citra, Cascade, Mt. Hood, Mosaic, Chinook, Apollo, Amarillo, and others contain relatively large amounts of geraniol, with significant variations from different crop years. In this study, they measured the amount of linalool, geraniol, and citranellol citronellol in beers that were dry hopped at different time points: before yeast was added (labeled "pre-yeast" in the table below, and represents something similar to whirlpool hop additions), 3 days after yeast was added, and at the end of fermentation. For each of these timings, they tested three different hop varieties that contained high levels of geraniol: Cascade, Bravo, and Mosaic. Overall, the amount of linalool in the finished beers werenwasn't effected affected by the timing of the dry hop. The amount of citronellol was also not effected affected by the timing of the dry hop except for the Bravo hops where the post-fermentation hopping resulted in about half the amount of citronollel citronellol than it did for the pre-fermentation and 3-day fermentation dry hop timings (see the bar graph based on the data from Takoi et al. 2014 below). The timing of the dry hop had the largest effect on geraniol: the earlier the dry hop, the less geraniol was present in the finished beer for all three hop varieties, with hops added pre-fermentation producing the lowest amount of geraniol and hops added post-fermentation producing the most geraniol in the finished beers. As in their previous study, citronellol increased during the first three days of fermentation, remained relatively stable for the rest of fermentation, and then increased again during storage. Geraniol dropped significantly during the first three days of fermentation in the case of the pre-yeast and 3-day dry hop timing, and increased slightly during storage. This data indicates that while earlier dry hopping reduces geraniol, only certain varieties of hops have an increase in citronellol depending on the dry hop timing. It's been suggested that the transformation of geraniol to citronellol involves unknown mechanisms that are relatively complex, particularly because the rate of the disappearance of geraniol does not map onto the rate of increase in citronellol, and when post-fermentation dry hopping there is a high amount of free geraniol but not a corresponding increase in citronellol during storage <ref>[https://www.researchgate.net/publication/261475199_Screening_of_Geraniol-rich_Flavor_Hop_and_Interesting_Behavior_of_beta-Citronellol_During_Fermentation_under_Various_Hop-Addition_Timings Screening of Geraniol-rich Flavor Hop and Interesting Behavior of beta-Citronellol During Fermentation under Various Hop-Addition Timings. Takoi, Kiyoshi & Itoga, Yutaka & Takayanagi, Junji & Kosugi, Takayuki & Shioi, Toru & Nakamura, Takeshi & Watari, Junji. 2014. DOI: 10.1094/ASBCJ-2014-0116-01.]</ref>.

Other yeast species can also convert monoterpenes. For example, a strain of ''Kluyveromyces lactis'' was found to reduce geraniol to citronellol. This strain and a strain of ''Torulaspora delbrueckii'' produced linalool from both geraniol and nerol, and could also form geraniol from nerol <ref>[https://www.ncbi.nlm.nih.gov/pubmed/10790686 Biotransformation of monoterpene alcohols by Saccharomyces cerevisiae, Torulaspora delbrueckii and Kluyveromyces lactis. King A1, Richard Dickinson J. 2000.]</ref>. Many species of ''Debaryomyces'', ''Kluyveromyces'', and ''Pichia'' were found to transform geraniol into linalool, and nerol into linalool and alpha-terpineol <ref>[https://www.ncbi.nlm.nih.gov/pubmed/18357555 Biotransformation of acyclic monoterpenoids by Debaryomyces sp., Kluyveromyces sp., and Pichia sp. strains of environmental origin. Ponzoni C, Gasparetti C, Goretti M, Turchetti B, Pagnoni UM, Cramarossa MR, Forti L, Buzzini P. 2008.]</ref>. Sulfur based compounds known as Colomer et al. (2020) measured the monoterpenes in two experiments before and after inoculating with different strains of ''thiolsBrettanomyces'' have also been shown to be produced by yeast fermentation from hop derived precursors (suspected to be Sthat had varying degrees of beta-glutathione)glucosidase activity. So far, science has They found that these include the volatile thiols 3strains with the least beta-sulfanyl-4-methylpentan-1-ol (3S4MP; grapefruit) and 3-sulfanyl-4-methylpentyl acetate (3S4MPA; passionfruitglucosidase activity had the least impact on biotransformation, grapefruit). These thiols were found but the increase in beers dry hopped separately with Amarillo, Hallertau Blanc, and Mosaic hop varieties. The amounts of these two thiols were beta-citronellol was higher than expected based on the content of these thiols what has been reported in the hops alone biotransformation studies with ''Saccharomyces'' <ref name="Cibaka_2016" />. See also this [https://wwwonlinelibrary.facebookwiley.com/groupsdoi/MilkTheFunkfull/permalink10.1002/1373899592638251jib.610 Biotransformation of hop derived compounds by Brettanomyces yeast strains. Marc Serra Colomer, Birgitte Funch, Natalia Solodovnikova, Timothy John Hobley, Jochen Förster. 2020. DOI: https://doi.org/10.1002/ MTF thread speculating on how jib.610.]</ref>. See [[Brettanomyces#Hop_Biotransformation|''Brettanomyces'' might produce thiolshop biotransformation]]for more information.

In general, different yeast strains have a large impact on how hops are perceived in the final beer, including both perceived bitterness and flavors. For example, POF+ (phenolic positive) strains of ''[[Saccharomyces|Saccharomyces cerevisiae]]'' tends to mask the hop -derived aromas in dry hopped beers <ref name="Sharp_Presentation" />. A beer hopped with the Tradition hop variety produced fruit flavors when fermented with Abbaye ale yeast, and woody/spicy flavors when fermented with US-05. When the beer was brewed with Citra hops, with US-05 the beer had sweet fruits/citrus flavors and more bitterness, but when fermented with the Abbaye ale strain the beer had a more one dimensional sweet fruit/floral flavor and less bitterness <ref>"Influence of yeast strain on hop aroma development in dry hopped beers." Christina Schönberger, Elisabeth Wiesen, Benedikt Matsche, Barth Innovations Yves Gosselin, Stephan Meulemans, Fermentis. Presentation slides at 35th Congress EBC.</ref>.

:<youtube height="200" width="300">GCQ22HSDDUQ</youtube> <youtube height="200" width="300">r09eb46k97I</youtube>* [https://www.crowdcast.io/c/itcl8mhhsdsa Escarpment Labs presentation by Richard Preiss on biotransformation.]* [https://www.masterbrewerspodcast.com/282 MBAA Podcast #282: New Belgium's tests with mid-fermentation dry hopping.]

====Glycosides====Hops contain In addition to terpenes and monoterpene alcohols being found in hops in a free form, they are also present in the form of glycosides, which are flavor compounds that are bound to a sugar molecule. Plants use glycosides for a range of metabolic purposes. The amounts and types of glycosides in hops varies by hop variety, and can be affected by plant fertilization and harvest timing <ref name="Svedlund_2022">[https://link.springer.com/article/10.1007/s00253-022-12068-w Svedlund, N., Evering, S., Gibson, B. et al. Fruits of their labour: biotransformation reactions of yeasts during brewery fermentation. Appl Microbiol Biotechnol 106, 4929–4944 (2022). https://doi.org/10.1007/s00253-022-120.]</ref>. In their bound form, glycosides are flavorless. Studies on hop compounds elude to the possibility of compounds being produced by the glycosidic activity of ''S. cerevisiae'', however direct evidence of glucosidic activity in ''S. cerevisiae'' is lacking. Daenen (2008) reviewed the glycosidic activity of many strains of ''S. cerevisiae'', and found that only a few strains expressed any real glucosidic activity and none that exhibited exo-beta-glucosidase which would be required to break glycosidic bonds in the beer/wort. Daenen did find that enzymatic activity from some strains of ''Brettanomyces'' can efficiently release these bound compounds and release their flavor and aromatic potential <ref name="Praet_2012" />. Beta-glucosidase enzyme can also be added to beer to enhance the breakdown of glycosides and intensify hop -derived flavors and aromas. For example, one study showed an increase in citrus, orange, grapefruit, and tropical pineapple in a Cascade dry hopped beer that had beta-glucosidase enzymes added to it <ref>"Optimizing hop aroma in beer dry hopped with Cascade utilizing glycosidic enzymes (presentation slides)." Kaylyn Kirkpatrick from New Belgium Brewing Co. Young Scientist Symposium, Chico, CA 2016.</ref>. There is also some evidence to support that there is higher glucosidase activity in seeded hops, which are generally not used in the brewing industry <ref>"Seeded and "Unseeded Hops - a Quality Comparison (presentation slides)." Martin Zarnkow. EBC 2015.</ref>. Hops also contain polyhenols that are bound in glycosidic form that could contribute a small amount of bitterness <ref>[https://www.tandfonline.com/doi/abs/10.1080/03610470.2021.2024112?journalCode=ujbc20 Martin Biendl, Stefanie Ritter & Christina Schmidt (2022) Monitoring of Glycosidically Bound Polyphenols in Hops and Hop Products Using LC-MS/MS Technique, Journal of the American Society of Brewing Chemists, DOI: 10.1080/03610470.2021.2024112.]</ref> (see also [https://traffic.libsyn.com/secure/forcedn/thebrulab/056_Applying_the_Science_-_High_Hop_Loads_w_Jordan_Folks.mp3 Bru Lab Podcast Episode 055; Hop Bitterness And Polyphenols w/ Dr. Martin Biendl]).

Much The effects of the work beta-glucosidase on hop derived glycosides has been done hops may be limited even in ideal conditions using hop oils, and might not apply to whole cones or pellet hopspure beta-glucosidase that is highly efficient in beer where beta-glucosidase activity by yeast is limited. Sharp et al. (2017) determined that hops contain such a small amount of glycosides that their release doesn't contribute much to hop flavor and aroma. While previous studies focused on hop extracts, they studied glycosides in whole leaf hops for the first time and found that non-significant levels of hop-derived monoterpenes from glycosides even when using pure beta-glucosidase extract extracted from almonds. For example, linalool was increased by 16.5 μg/L when using the highest hopping rate, but this amount has little impact on beer hopped with whole leaf hops that the amount overall aroma and flavor of increased monoterpenes such as linaloolthe beer. The terpenes citronellol, terpineol, citronellol, nerol, and geraniol is small and insignificant. The were also not increased in significant amounts, however, the fatty alcohol 1-octanol (waxy, green, citrus, orange, aldehydic, fruity <ref>[http://www.thegoodscentscompany.com/data/rw1021071.html Octanol. The Good Scents Company. Retrieved 03/31/2017.]</ref>) was the only measured flavor compound that was increased significantly <ref name="Sharp_2017">[http://onlinelibrary.wiley.com/doi/10.1002/jib.418/abstract The effect of hopping regime, cultivar and β-glucosidase activity on monoterpene alcohol concentrations in wort and beer. Daniel C. Sharp, Jan Steensels, Thomas H. Shellhammer. 2017. DOI: 10.1021/jf2042517.]</ref>. The alcohol octanol can be esterified into octyl acetate, which is a classically "citrusy" aroma, so perhaps certain yeasts can create this ester during mid-fermentation hopping <ref>[https://pubs.acs.org/doi/abs/10.1021/jf2042517 Eric G. Dennis, Robert A. Keyzers, Curtis M. Kalua, Suzanne M. Maffei, Emily L. Nicholson, and Paul K. Boss. 2012.]</ref>.

For techniques and usage amounts of aged hops, see [[Hops#Aged_Hops_in_LambicAged_Hops_in_Lambic_and_Other_Spontaneous_Fermentation_Beer|Aged Hops in Lambic]].

Hops Typically, only low alpha acid hops are used (high alpha acid hops may lead to more hop character and higher inhibition of lactic acid bacteria than desired possibly due to oxidized acids). The hops are typically bound in burlap sacks/paper bags or something similar that allows for exposure to oxygen, and then they are left to age in preferably low humidity conditions at room temperatures to warm temperatures (warmer temperatures will encourage faster aging). Changes in the environment such as temperature shifts are not a concern; for example [https://web.archive.org/web/20180204131402/http://jesterkingbrewery.com/home-for-our-aged-hops Jester King Brewery in Austin Texas ages hops in a horse barn]. The hops should are traditionally aged for 2+ years. Monitor for mold growth during this time, and discard any hops that show visible signs of mold growth. Some brewers prefer to age the hops until the cheesy character ([[Isovaleric Acid]]) is gone, while other brewers do not mind the presence of this cheesy character (for example, some [[Lambic|lambic]] beers display isovaleric acid character even after packaging). While hop leaves are generally preferred over pelletized hops, if aging pellets, it has been advised to break the pellets up so that the entirety of the hop material is exposed to the air.  Freshly harvested hops (also called "wet hops") should not be aged. Freshly harvested hops should be dried first , as is normal for hop processing, before aging in order to help prevent [[Mold|mold]] growth (see this [https://www.canr.msu.edu/news/drying_hops_on_a_small_scale article from Michigan State University] on measuring moisture levels and [https://www.homebrewersassociation.org/how-to-brew/how-to-harvest-prepare-and-store-homegrown-hops/ this AHA article on drying hops] for home growers)and this [https://www.masterbrewerspodcast. Typically, only low alpha acid hops are used (high alpha acid hops may lead to more com/117 MBAA podcast interview with Val Peacock on industrial hop character and higher inhibition of lactic acid bacteria than desireddrying]). The hops are typically bound in burlap sacks and left  See also:* [https://www.facebook.com/groups/MilkTheFunk/permalink/2696660217028842/ Joshua Martinez MTF thread on using a rotary composter to age in preferably low humidity conditions. The pulverize pellet hops should be aged for 2+ yearsaging.]* [https://www.facebook. Monitor for mold growth during this time, and discard any com/groups/MilkTheFunk/permalink/3042350479126479/ MTF thread on general tips on aging hops that show visible signs of mold growth.]

! Storage !! Oil !! Hallertau Magnum (Pellet, 14% AA) <ref name="Kishimoto_2021" /> !! Saaz (pelletPellet) <ref name="Mikyška_2012" /> !! Sládek (pelletPellet) <ref name="Mikyška_2012" /> !! Premiant (pelletPellet) <ref name="Mikyška_2012" /> !! Agnus (pelletPellet) <ref name="Mikyška_2012" /> !! Saaz (leafLeaf) <ref name="krofta_2013">[http://www.actahort.org/books/1010/1010_26.htm Stability of Hop Beta Acids and Their Decomposition Products during Natural Ageing. K. Krofta, S. Vrabcová, A. Mikyska, M. Jurková, T. Cajka , J. Hajslová. 2013.]</ref> !! Vital (leafLeaf) <ref name="krofta_2013" /> !! Pure Beta Acid <ref name="krofta_2013" /> |-| colspan="10" | '''Open air at 20°C for 12 months'''|- align="right"| || Alpha acids || || -80% || -88.3% || -64.3% || -78.2% || || |- align="right"| || Beta acids || || -60.5% || -83% || -53.7% || -51% || -50% || -77.5% || -99%|- | colspan="10" | '''Open air at 40°C for 3 months'''|- align="right"| || Alpha acids || -100% || || || || || || |||- | colspan="10" | '''Vacuum sealed at 20°C for 12 months'''|- align="right"| || Alpha acids || || -20.6% || -24.9% || -22.2% || -21.7% || || |||- align="right"| || Beta acids || || -2.7% || -1.7% || -2.1% || -1.2% || || ||

| colspan="910" | '''Open air Vacuum sealed at 20°C 2°C for 12 months'''|- align="right"| || Alpha acids || || -1.1% || -5.5% || -0.3% || -1.4% || || |||- align="right"| || Beta acids || || -1.7% || -2.3% || -0.4% || -0.5% || || ||

| || Alpha } It has been established that many factors influence the initial content of alpha and beta acids, such as cultivar (variety), cultivation area, climate and environmental conditions, harvest year, and harvest date. For example, early harvest dates result in hop cones that are not fully developed and contain less alpha/beta acids, and late harvest dates can produce hop cones with lower alpha/beta acids as these acids begin to degrade. Thus, optimal harvest dates occur when the hop cones reach their maximum size but before they become brittle and hop acid degradation begins. [https://www.pja.iung.pulawy.pl/stability-of-the-hop-bitter-acids || -80% || during-long-term-storage-of-cones/ Skomra and Koziara-88Ciupa (2020)] found that harvest date also affects how alpha and beta acids degrade over time during storage. They looked at alpha and beta acid degradation differences for 4 varieties of hops grown at the same farm, but harvest at 3% || -644 different dates (7-10 days between harvest dates) <ref name="Skomra_2020">[https://www.pja.iung.pulawy.pl/stability-of-the-hop-bitter-acids-during-long-term-storage-of-cones/ Stability of the hop bitter acids during long-term storage of cones with different maturity degree. Urszula Skomra, Marta Koziara-Ciupa. Polish Journal of Agronomy 2020, 40, 16–24. DOI: 10.26114/pja.iung.406.2020.40.03.]</ref>. The hops were stored at 5°C and 20°C in aerobic conditions for 12 months <ref>Private correspondence with Urszula Skomra by Dan Pixley. 07/21/2020.]</ref>. As expected, the lower storage temperature of 5°C preserved the alpha and beta acids better than the warmer 20°C storage temperature. In general, an earlier harvest date also resulted in greater degradation of alpha and beta acids, but there were some exceptions to this. For example, Sybilla Harvest Date II produced the most stable alpha acid content (harvesting too early or too late resulted in less stable alpha acid content), while for Magnet Harvest Date II resulted in the most degradation of alpha and beta acids and Harvest Date III was preferable for alpha acid stability. In general, the stability of beta acids depended more on the harvest date than the stability of alpha acids <ref name="Skomra_2020" />. See the table below. Skomra and Koziara-Ciupa (2020) also reported that for the first 3% || to 6 months, the alpha and beta acids were relatively stable. The cooler storage had a longer period of initial stability than the warmer storage. After the initial period of stability, the alpha and beta acids began a rapid drop around 9 months of total storage. From 9-7812 months, the rate of decline in alpha and beta acids was much slower and seemed to somewhat stabilize <ref name="Skomra_2020" />.2% || |  {| class="wikitable sortable"

| colspan="9" | '''Vacuum sealed Open air at 20°C 5°C for 12 months'''(Harvest dates I, II, II, and only for Magnat/Magnum IV)

| || Alpha acids || -2010.68%, -4.5%, -7.9% || -2411.90%, -6.4%, -6.8% || -2225.8%, -40.8%, -2.3%, -9.3% || -2117.7% || || ||, -4.5%, -12.9%, -11.2%

| || Beta acids || -27.6%, -6.3%, -8.75% || -28.1%, -10.79%, -17% || -33.3%, -37.2%, -21.1%, -22.3% || -28.1%, -7.6%, -3.9%, -5.2% || || ||

| colspan="9" | '''Vacuum sealed Open air at 2°C 20°C for 12 months'''(Harvest dates I, II, II, and only for Magnat/Magnum IV)

| || Alpha acids || -128.5%, -11.3%, -27.1% || -518.56% || , -035.3%, -37.6% || -135.4%, -56.8%, -31.7%, -36% || || ||-38.3%, -25.7%, -28.9%, -24.4%

| || Beta acids || -126.4%, -38.6%, -49.75% || -243.3%, 45.7%, 54.1% || -046.3%, -66.46%, -59.8%, -59.2% || -053.7%, -28.51%, -16.8%, -19.9% || || ||

[[File:Lam Hop Oils'''Oxidized alpha acids''' (humulinones) are similar in taste perception to iso-α-acids, but have been described as less bitter (an average of about 66% as bitter on a 1 to 1 basis). The quality of the bitterness from oxidized alpha acids has been described in one study as "smoother and less lingering" than iso-alpha acids; this was attributed to humulinones being more polar than iso-alpha acids and therefore do not stick or linger on the tongue as long as iso-alpha acids <ref name="Shellhammer, Vollmer and Sharp, CBC 2015"/><ref name="Maye_2016" />. While the taste threshold of iso-alpha acids is 5-6 mg/L in light lager, the threshold for humulinones has been measured to be 8 mg/L in light lager (note that this is an average; tasters vary widely in how much bitterness they perceived from different bitter compounds) <ref name="Algazzali_2014" />. Humulinone content increases in hops after being pelletized (whole leaf hops have less humulinones). In fresh pellet hops that have a relatively low humulinone content, the humulinones contribute little to the bitterness of the beer when boiled, however when dry hopped they readily dissolve into the beer and have a significant impact on the beer's bitterness. With heavy dry hopping, the humulinones also decrease iso-alpha acid content of beer with more than about 25 IBU's, but not in beer with less than about 20 IBU.jpg|thumb|400| The decrease in iso-alpha acids and perceived bitterness/IBU is partially made up for the bitterness of the humulinones themselves (humulinones are picked up in IBU measurements with a [http://pubschem.acslibretexts.org/doiCore/Physical_and_Theoretical_Chemistry/Kinetics/Reaction_Rates/Experimental_Determination_of_Kinetcs/Spectrophotometry spectrophotometer] and as such it has been suggested that IBU's be [http:/pdf/10masterbrewerspodcast.1021com/jf00070a043 Data from "Aging of hops 004-dry-hopping-its-effects-on-bitterness-and their contribution -the-ibu-test-0 measured more accurately with HPLC]). In beers with less than 20 IBU, high dry hopping rates greatly increase the bitterness/IBU due to beer flavorthe bitter humulinones <ref name="Maye_2016" />. Humulinones also increase foam creation and stability by Kai Cbinding with the foam positive protein, Protein Z, via hydrogen bonding <ref>[https://www. Lamsciencedirect.com/science/article/abs/pii/S0308814623020678 Chen Xu, Xuanqi Zhang, Mingyang Sun, Robert THanhan Liu, Chenyan Lv. Foster Interactions between humulinone derived from aged hops and Max Lprotein Z enhance the foamability and foam stability. Food Chemistry, Volume 434, 2024, 137449, ISSN 0308-8146, https://doi.org/10.1016/j.foodchem.2023. Deinzer137449.] '''Aged I''': 2 weeks at 90°F; '''Aged II''': 60 additional days at 90°F</ref>.]]

'''Oxidized alpha acids''' (humulinones) are similar in taste perception to iso-α-acids, but have been described as less bitter (an average of about 66% as bitter on a 1 to 1 basis). The quality rate of the bitterness from oxidized alpha acids has been described in one study as "smoother and less lingering" than iso-alpha acids; this was attributed to humulinones being more polar than iso-alpha acids and therefore do not stick or linger on the tongue as long as iso-alpha acids <ref name="Shellhammer, Vollmer and Sharp, CBC 2015"/><ref name="Maye_2016" />. While the taste threshold of iso-alpha acids humulinone formation is 5-6 mg/L in light lager, the threshold for humulinones has been measured to be 8 mg/L in light lager (note that this is an average; tasters vary widely in how much bitterness they perceived from different bitter compounds) <ref name="Algazzali_2014" />limiting. Humulinone content increases in hops after being pelletized (whole leaf hops have less humulinones). In fresh pellet hops that have a relatively low humulinone content, the humulinones contribute little to the bitterness of the beer when boiled, however when dry hopped they readily dissolve into the beer and have a significant impact on the beer's bitterness. With heavy dry hopping, the humulinones also decrease iso-alpha acid content of beer with more than about 25 IBU's, but not in beer with less than about 20 IBUMaye et al. The decrease in iso-alpha acids and perceived bitterness/IBU is partially made up for the bitterness of the humulinones themselves (humulinones are picked up in IBU measurements with a [http://chem.libretexts.org/Core/Physical_and_Theoretical_Chemistry/Kinetics/Reaction_Rates/Experimental_Determination_of_Kinetcs/Spectrophotometry spectrophotometer] and as such it has been suggested that IBU's be [http://masterbrewerspodcast.com/004-dry-hopping-its-effects-on-bitterness-and-the-ibu-test-0 measured more accurately with HPLC]2016). In beers with less than 20 IBU, high dry hopping rates greatly increase the bitterness/IBU due to the bitter humulinones. The rate of humulinone formation is limiting, meaning showed that humulinone formation occurs occurred rapidly during hop pelletization, and the concentration peaks during this time (these researchers found that further exposure to air did not increase humulinone content). Scientists believe They speculated that this is because when whole leaf hops are baled, only 20% of lupulin glands are broken, whereas when they are pelletized 100% of the lupulin glands are broken. In contrast, Taniguchi et al. (2013) found that humulinone formed slowly over 40 weeks in pellets that were stored at 20°C, but at 40°C and 60°C storage temperature they formed quickly and then diminished to very low levels at 10 weeks and 2 weeks respective to the higher storage temperatures. This demonstrated that warm storage increases the rate of humulinone formation. In addition, the oxidized hop compound 4′-hydroxy-allohumulinone increased trailing behind the humulinone formation and at a similar concentration at 20°C over 40 weeks of storage, and were much more stable than humulinones when stored at 40-60°C for 40 weeks <ref>[https://pubs.acs.org/doi/10.1021/jf3047187 Identification and Quantification of the Oxidation Products Derived from α-Acids and β-Acids During Storage of Hops (Humulus lupulus L.). Yoshimasa Taniguchi, Yasuko Matsukura, Hiromi Ozaki, Koichi Nishimura, and Kazutoshi Shindo. Journal of Agricultural and Food Chemistry 2013 61 (12), 3121-3130. DOI: 10.1021/jf3047187.]</ref>. The exact mechanism by which alpha acids are converted to humulinones is not known . The amount of humulinone correlates strongly with the [http://methods.asbcnet.org/summaries/hops-12.aspx Hop Storage Index (HSI)] <ref name="Maye_2016" />. Humulinone content in long-aged hops (1+ years) has not been studied.

'''Oxidized beta acids''' produce some compounds that also contribute to the perception of bitterness, specifically hulupones. Unlike humulinones which form relatively quickly from the oxidation of alpha acids, hulupones form at a much slower rate <ref name="Dušek_2014" />. Also unlike humulinones, they survive boiling and fermentation. While some sensory analysis of beers containing oxidized beta acids describes the resulting bitterness as "harsh and clinging", another analysis by Krafta et al (2013) described the bitterness of oxidized beta acids in beer when added in their pure form at the beginning of the boil as "pleasant and not lingering". The more degradation of beta acids into oxidized beta acids that occurs in hops, the more bitter beers brewed with these hops will be <ref name="krofta_2013" />. Two other compounds other than hulupones have been identified as being produced by the oxidation of beta acids, epoxycohulupone and epoxyhulupone. Their effect on beer flavor is not yet known, ; however, it is thought believed that hulupones have the greatest a greater impact on beer flavor and bitterness than these compounds <ref name="Dušek_2014" />.

====IBU==== The overall effect of oxidized compounds in aged hops has been shown by Val Peacock, a former scientist at Anheuser-Busch. Peacock stored hops at four different temperatures for 18 months. His data showed that although the alpha acid content in the hops and the iso-alpha acid content in the beers brewed with them decreased the older the hops were stored, the measured IBU of the different beers was about the same. This is because the oxidized acids in hops show up in the same spectrum as iso-alpha acids when using the ASBC method of measuring IBUs with a spectrophotometer <ref name="Aron">[https://www.homebrewersassociation.org/how-to-brew/resources/conference-seminars Dr. Patricia Aron. "Bitterness and the IBU: What’s It All About?" HomebrewCon 2017 Presentation. ~34 mins in. Retrieved 09/05/2017.]</ref>. This data is shown below. Caleb Buck's experiment [[Hops#Aged_Hops_in_Lambic|seen below]] supports this. Although it has not been shown that oxidized alpha and/or beta acids can inhibit lactic acid bacteria, if they do, then this might help explain reports <ref>[https://homebrewingfun.blogspot.com/2019/11/one-gallon-spontaneous-fermentation.html Adam Kielich. "One Gallon Spontaneous Fermentation Beer Batch 5 Recipe and Brewday". Brain Sparging on Brewing. 11/16/2019.]</ref> of using aged hops that originally had a high alpha acid content retaining a strong inhibitory effect towards lactic acid bacteria.

A recent 2017 study at the Shellhammer lab looked at how trained panelists and consumers perceived a lager beer dry hopped with slightly oxidized Hallertau Mittelfrüh hops (exposed to oxygen once, then stored at 38°C for two weeks) versus highly oxidized (daily exposure to oxygen and stored at 38°C for two weeks). They found that the trained panelists detected more characteristics that are associated with noble hops; e.g. more woody, earthy, and herbal characteristics in the lager beers dry hopped with oxidized hops. They also found the oxidized hopped beers to be more bitter (probably due to oxidized alpha and beta acids). Consumers were not statistically able to tell the difference. The study determined that oxidized hops might serve to provide nuanced increases in noble hop character <ref>[http://www.asbcnet.org/publications/journal/vol/2017/Pages/ASBCJ-2017-1287-01.aspx Aroma Properties of Lager Beer Dry-Hopped with Oxidized Hops. Daniel M. Vollmer, Victor Algazzali, and Thomas H. Shellhammer. 2017.]</ref>. In a similar study by Hengyuan et al. (2023), aging Saaz hops for a short amount of time (aged at 30 ℃ in a ventilated environment for 5 days) reduced the grassy and resinous character and improved the spicy and woody character of single hopped beers (hopped at three points in the boil and dry hopped). Consumers also preferred this beer over the other samples, including fresh Saaz. However, over-aging Saaz hops resulted in less preference with consumers (30 ℃ for 10 days, 40 ℃ for 5 days, or 40 ℃ for 10 days). This effect was not demonstrated by Simcoe hops, however, indicating that short aging is only beneficial for some hop varieties. Under the same aging condition with Simcoe hops, consumers preferred the fresh Simcoe, and any degree of aged Simcoe was less preferred. The beers with aged Simcoe hops were described as having less fruity and floral character and more dry, thin bitterness <ref>[https://www.bio-conferences.org/articles/bioconf/abs/2023/04/bioconf_icbb2023_01016/bioconf_icbb2023_01016.html Aging of Hops and Their Effects on India Pale Ale Flavor. Hengyuan Xu, Shaokang Sun, Xiaochen Wang, Haojun Zhang and Cong Nie. BIO Web Conf., 59 (2023) 01016. DOI: https://doi.org/10.1051/bioconf/20235901016.]</ref>.

* [https://www.mainiacalyeastyakimachief.com/commercial/hop-wire/introducing-the-ych-aged-hops/ Mainiacal Yeast -program YCH offers 1-2 oz bags for homebrewers and 44 lb bags of aged hops (small lots ; contact for homebrewers)more information.]

* <s>[https://hopsdirect.com/products/choice-debittered-pellets Hops Direct "Choice Debittered/Aged Hops" (Pellet - Columbus).]</s>* [https://www.freshops.com/shop/hop/aroma-hopproduct/lambic-hops/ Freshhops "Lambic Hops" (Leaf - Willamette)".]* [httphttps://www.yakimavalleyhops.com/Lambic2oz_pproducts/hopslambichops3.htm lambic-hop-pellets Yakima Valley Hops "Lambic / Aged Hops" (Pellet).]

* [httphttps://www.themaltmiller.co.uk/index.php?_a=viewProd&productId=592 The Malt Miller (UK).]* [http:/product/www.brewstore.co.uk/specially-aged-fuggles-hops-100-grams Brew Store UK / The Malt Miller (Leaf - Fuggles).]* [http://www.brewstore.co.uk/specially-aged-hallertau-hops-100-grams Brew Store UK (Leaf - Hallertau).]

* [https://www.yakimachiefbsgcraftbrewing.com/ YCH lambic-hop-blend BSG sometimes offers 44 lb bags of aged pellet hopsfor commercial brewers; contact for more informationdetails.] See [https://www.facebook.com/groups/MilkTheFunk/permalink/4326634927364688 this MTF thread]on experiences with these hops.* [https://www.ebay.com/itm/Lambic-Hops-pellet-or-whole-cone/172249202875?hash=item281ada40bb:m:mX2CSrKvWb0-3WIIuBh0aLA&var=471070249004 Ted from Hop Heaven on eBay sells 8 oz and 1 lb bags of aged pellet and leaf hops.] See this [https://www.facebook.com/groups/MilkTheFunk/?comment_tracking=%7B%22tn%22%3A%22R%22%7D reviews on permalink/4338527352842112/ MTFthread]on reviews of this product.* [https://shop.humle.se/humle/lambichumle/ Humlegårdens (Sweden); several varieties of aged whole leaf hops, 100 gram quantities.]

[https://beerandbrewing.com/dictionary/96jvGQTSdT/ Dry hopping] is the technique of adding hops to beer that has finished fermenting or is in the process of fermenting. Historically, dry hopping is associated with English pale ales of the 1800's, and it was re-discovered by Fritz Maytag of Anchor Brewing Company in 1975 when the company brewed the first American IPA, Liberty Ale <ref>[https://beerandbrewing.com/dictionary/96jvGQTSdT/ "The Oxford Companion to Beer definition of dry hopping," Garrett Oliver. Craft beer & Brewing Magazine. Retrieved 11/16/2023.]</ref><ref>[https://northamericanbrewers.org/liberty-ale/ "Liberty Ale". North American Brewers Association. April 28, 2022. Retrieved 11/16/2023.]</ref><ref>[https://www.youtube.com/watch?v=4ijHO5TFG3I Scott Ungermann, Brewmaster of Anchor Brewing Company. The Doug Piper Gourmet Brewing Podcast. July 14th, 2023.]</ref>(20 mins in). Brewers have had positive and interesting results dry -hopping sour and funky beer. Often fresh American or New Zealand varieties that compliment complement fruit flavors are chosen, however, other varieties have been used as well, including English and German hops. Just as in dry hopping normal beers, dry hopping sour/funky should be done after the beer has matured. Dry hopping for around 1-3 days before [[packaging]] the beer is adequate for extraction, depending on whether or not the beer is recirculated or agitated (agitation of the beer while on contact with the dry hops attains full extraction in 24 hours) <ref>[http://ir.library.oregonstate.edu/xmlui/bitstream/handle/1957/34093/Wolfe_thesis.pdf?sequence=1 A Study of Factors Affecting the Extraction of Flavor When Dry Hopping Beer (master thesis). Peter Harold Wolfe. 2012.]</ref>. Hopping rates generally range from 0.5-1 ounces per 5 gallons 1 gallon of beer (1-2 pounds per bblor 3.7-7.5 grams per liter) to achieve hop-forward flavors, although lesser rates can be used to achieve a more subtle character (see the threads below) <ref>[https://www.facebook.com/groups/MilkTheFunk/permalink/1693639013997639/?comment_id=1693660390662168&comment_tracking=%7B%22tn%22%3A%22R4%22%7D Nate Walter and Dan Pixley. Milk The Funk Facebook group. 05/21/2017.]</ref>.

Dry hopping can contribute to bitterness in beer through oxidized alpha acids and oxidized beta acids. Oxidized alpha acids can also reduce iso-alpha acids in beers that begin with more than 25 IBU from iso-alpha acids, potentially reducing percieved bitterness after dry hopping (see [[Hops#Acids_2|Oxidized Alpha Acids]] above). Some alpha acids will also dissolve into the beer, which are estimated as being 10% as bitter as iso-alpha acids. Dry hopping also has a linear impact on the pH of beer regardless of the starting IBU or pH: the pH rises by 0.14 per pound of hop pellets per barrel of beer in a beer that started with a pH of 4.2 (~0.5 ounces per gallonor 3.7 grams per liter) <ref name="Maye_2016" /><ref name="Shellhammer, Vollmer and Sharp, CBC 2015"/>. This rise in pH might be less in more acidic beers that are dry hopped since pH is a logarithmic scale. Dry hopping can also reduce head retention in beers, although this is variety dependent (one study found that dry hopping with Eureka and Apollo hops increased head retention, while dry hopping with Bravo, Centennial, and Cascade decreased head retention). Extended dry hopping times (after 3 days) can also reduce head retention <ref name="Maye_EBC2017" />.

Although See also:* [http://thebrulab.libsyn.com/episode-079-impact-dry-hopping-has-on-bitterness-ibu-and-ph-w-dr-john-paul-maye Bru Lab podcast Episode 079 - Impact Dry Hopping Has On Bitterness, IBU, and pH w/ Dr. John Paul Maye.]* [https://www.therarebarrel.com/index.cfm?method=blog.blogDrilldown&blogEntryID=8141D6FA-EE78-5BCE-E08E-704DB6EEA279&originalMarketingURL=blog/Dry-hopped--Changing-process-leads-to-bottling The Rare Barrel reports on an anecdote that dry hopping in a less sour beer extracts better hop aroma, and ''Brettanomyces'' preserves the character.]* [https://www.facebook.com/groups/MilkTheFunk/permalink/2358742877487246/ MTF thread by Dave Janssen on experiences with doing long-term dry hopping with noble hops in sour beers.] See also [http://www.horscategoriebrewing.com/2018/11/saison-from-1911-finishing-hops-and.html Dave's research on a 1911 saison that was dry hopped long term].* [https://www.facebook.com/groups/MilkTheFunk/permalink/2718954364799427/ MTF thread on the mechanisms are not fully understoodpossibility/mechanism of dry hopping contributing to the "funky" character of sour beers.]* [https://www.facebook.com/groups/MilkTheFunk/permalink/2828125150549014/ MTF thread on dry hopping with aged hops.]* [https://www.facebook.com/groups/MilkTheFunk/permalink/2994066257288235/ MTF thread from Benedikt Koch on using aged hops in a natural cider, dry inspired by Revel Cider.] ====Inhibiting Lactic Acid Bacteria====Dry hopping inhibits Gram-positive bacteria such as ''Lactobacillus''. Humulinic acids have been found to greatly inhibit bacteria (see [[Hops#Antimicrobial_Properties|Antimicrobial Properties]]). Other compounds such as non-isomerized alpha acids, other oxidized hop acids, or the small amount of isomerization of alpha acids that happens in beer at room temperature <ref>[http://scottjanish.com/zero-hot-side-hopped-neipa-hplc-testing-sensory-bitterness/ Janish, Scott. "Zero Hot-Side Hopped NEIPA | HPLC Testing for Sensory Bitterness". ScottJanish.com. Retrieved 03/09/2017.]</ref>, could contribute to inhibiting lactic acid bacteria. See the links [[Hops#Antimicrobial_Properties|antimicrobial properties]] above. See also reported data below.

* [https://www.facebookratebeer.com/groupsforums/MilkTheFunk/permalink/2358742877487246/ MTF thread by Dave Janssen on experiences with doing longlab-and-term dry hopping with noble hops in sour beershops_289071.] See also [http://www.horscategoriebrewinghtm "CLevar" on Ratebeer.com/2018/11/saison-from-1911-finishing-data point on ''Lactobacillus'' being inhibited by hops, but not as much by iso-andalpha acid hop extract.html Dave's research on a 1911 saison that was dry hopped long term].

Also known as "dry hop creep", it was first discovered in 1893 by Brown and Morris that dry hopping increases the ABV of beers and dries them out. Dry hop creep, along with the secondary fermentation of ''Brettanomyces'' in long aged British ales of the late 19th and early 20th centuries, were once characterized as the potential mechanisms by which long aged ales could retain a high level of carbonation in casks <ref>[https://archive.org/details/principlespracti00syke "The principles and practice of brewing" Sykes, Walter John. London, C. Griffin and Company, limited, 1907. Pgs 384-388.]</ref><ref>[https://www.facebook.com/groups/MilkTheFunk/permalink/4709953772366133 Gareth Young. Milk The Funk Facebook group thread about English brewers historically relying on Brettanomyces and dry hop creep for carbonation in long aged ales. 06/17/2021.]</ref>. It was proposed that the likely cause is the release of glycolytic enzymes that break down starches into sugars which viable yeast can then ferment. Brewers normally aim to control the final alcohol percentage in a beer through brewhouse operations rather than postfermentation dilutions with lower/higher alcohol beers or water. This approach to brewing is called "brewing to final gravity." Due to the need to have a predictable ABV for government regulatory reasons, unexpected fermentation is, therefore, a concern for many breweries <ref name="Kirkendall_2018">[https://www.tandfonline.com/doi/abs/10.1080/03610470.2018.1469081?journalCode=ujbc20 The Freshening Power of Centennial Hops. Jacob A. Kirkendall, Carter A. Mitchell & Lucas R. Chadwick. 2018. DOI: https://doi.org/10.1080/03610470.2018.1469081.]</ref>. Hop creep can also result in additional attenuation and higher carbonation after packaging, as well as diacetyl production.

More recent studies have shown that there is a difference in this enzymatic power between different hop varieties. [https://www.asbcnet.org/publications/journal/vol/2017/Pages/ASBCJ-2017-2257-01.aspx Cibaka et al. (2017)] reported an increase in ABV when dry hopping with Amarillo and Sorachi Ace hops, but not when dry hopping with Citra or Hallertau Blanc. Interestingly, they also found that Mosaic hops resulted in the opposite effect and it ; the Mosaic dry hopped beer dropped from 4% ABV to 3.6% ABV. It was hypothesized (probably possibly incorrectly) that Mosaic hops might release some sort of unidentified molecule that inhibits yeast fermentation/growth or viability . Cibaka et al. (2017) also demonstrated that late kettle additions might work to combat dry hop creep; the beer dry hopped with 2 g/L of Sorachi Ace finished at 5.1% ABV, while a beer that received a late kettle addition of 2 g/L of Sorachi Ace in addition to 2 g/L of Sorachi Ace dry hop finished at 4.3% ABV (the control with no dry hop finished at 4.0% ABV) <ref name="Kirkendall_2018" />.

Kirkpatrick and Shellhammer (2018) found that the enzymes responsible for the conversion of dextrins into sugars include amyloglucosidase (removes glucose from non-reducing ends of α-1,4 and branching α-1,6 linkages, with a preference for α-1,4 linkages and longer chain oligosaccharides), α-amylase (hydrolyzes randomly along glucopolysaccharides to produce maltose, maltotriose, maltopentaose, and maltohexaose products from amylose as well as maltose, glucose, and branched dextrins from amylopectin), β-amylase (saccharifiying enzyme, cleaving maltose in small amounts from nonreducing ends of glucopolysaccharides, and to a minor extent, maltotriose), and limit dextrinase (debranches limit dextrins at α-1,6 linkages, producing linear α-1,4 chains which can further be degraded by the combined action of amylases). They were able to successfully extract them from Cascade pellet hops using commercially available assays (enzyme specific para-nitrophenyl blocked oligosaccharide substrates). The amount of α and β-amylase found in Cascade hops was well below that of malted barley, but within the range reported in other plant leaves. These enzymes are denatured by high temperatures, and as such would be denatured when boiling hops. They reported a similar increase in ABV of 1.3% after 40 days when dry hopping a beer with Cascade hops (and a decrease of 1.9°P) at a rate of 10 g/L. They also found that the hops contained glucose and a small amount of fructose, which accounted for a sugar increase of 0.02−0.03 °P per gram of hops. More studies on whether or not the amount of dry hopping has a large effect needs to be done, and whether or not warmer temperatures speed up the enzymatic breakdown of dextrins, and the authors hypothesized that the rate of dextrin break down could be slowed by dry hopping at lower temperatures <ref name="Kirkpatrick_2018">[https://pubs.acs.org/doi/pdf/10.1021/acs.jafc.8b03563 Evidence of Dextrin Hydrolyzing Enzymes in Cascade Hops (Humulus lupulus). Kaylyn R. Kirkpatrick and Thomas H. Shellhammer. 2018. DOI: DOI: 10.1021/acs.jafc.8b03563.]</ref>. Rubottom and Shellhamer (2023) later demonstrated that drying hops at 150°F reduced the amount of enzymes in dried pellitized hops versus drying them at 130°F <ref>[https://www.tandfonline.com/doi/abs/10.1080/03610470.2023.2194838 Evaluating the Impact of High and Low Kilning Temperatures on Popular American Aroma Hops. Lindsey N. Rubottom, Thomas H. Shellhammer. Received 07 Feb 2023, Accepted 21 Mar 2023, Published online: 28 Apr 2023. https://doi.org/10.1080/03610470.2023.2194838.]</ref>.

The Kirkpatrick and Shellhammer (2018) also reported that the exposure time of the beer to the dry hop material played a significant role in the breakdown of dextrins. Most of the breakdown of dextrins occurs within 5 days (+0.7% ABV), but continued slowly up until at least 40 days (+1.3%). They also tested removing the hops via centrifuge and storing the beer at 10°C or 20°C. Their results suggested that the effect of the enzymatic breakdown of dextrins by hops appears to only be active when in contact with the hops and that once the beer is removed then this breakdown of dextrins stops. The authors suggest that to avoid as much breakdown of starches and over-attenuation from dry hops as possible, brewers can limit the amount of time sits on the hops and reduce the temperature, however, it is also important to consider how this might impact the product's flavor and careful measures should be taken to balance the over-attenuation problem and overall beer quality <ref name="Kirkpatrick_2018" />. After removing the beer from the hops, a second diacetyl rest has been suggested as a way to clean up any diacetyl or off-flavors that the yeast produces from the additional fermentation during dry hopping <ref>[https://www.rockstarbrewer.com/how-dry-hop-creep-causes-diacetyl-in-beer-and-how-brewers-can-minimise-the-risk/ STEVE 'HENDO' HENDERSON. How “Dry Hop Creep” Causes Diacetyl In Beer and How Brewers Can Minimise The Risk. Rockstar Brewer Academy website. 09/03/2018. Retrieved 10/05/2018.]</ref>. Other recommended solutions to avoiding hop creep is pasteurizing, filtering, or cold crashing out the yeast before dry hopping, storing the beer cold so that the yeast remains inactive, reducing dry hopping amounts, and dry hopping before fermentation is finished <ref>[http://beersmith.com/blog/2019/03/31/dry-hop-creep-over-carbonation-and-diacetyl-in-beer/ Brad Smith. BeerSmith blog. 03/13/2019. Retrieved 07/23/2019.]</ref>. [https://www.mbaa.com/publications/tq/tqPastIssues/2021/Pages/TQ-58-3-0705-01.aspx Teraoka et al. (2021)] argued that enzymes present in dry hops are not derived from the hop plants themselves, but microbes living on the hops and that the reported variability of starch degrading enzymes present in hops is due to the varying environmental conditions created by drying hops that encourage or discourage microbial growth. It has been reported that proteobacteria are the dominant type of bacteria that grow on hops, with the majority of them being Pseudomonas and Sphingomonas, with Pseudomonas having being previously reported to produce alpha-amylase. However, the authors recommend that further research is needed to identify which microbe species might be producing these enzymes on hops. They found that hops treated with the anti-microbial chemical sodium azide resulted in hops with much lower starch degrading enzymes. They also reported that alpha-amylase decreased in dried leaf and pellet hops over 3 months of anaerobic storage (storage temperature did play a role in the degradation rate of the enzymes). Alpha-amylase stayed relative consistent in fresh hops during storage. Beta-amylase remained stable over the three month storage time regardless of hop type. The authors also acknowledge another possible variable being that hop seed material might contain starch; however, seeded hops are generally not used in the brewing industry <ref>[https://www.mbaa.com/publications/tq/tqPastIssues/2021/Pages/TQ-58-3-0705-01.aspx "Do Starch-Degrading Enzymes in Hop Samples Originate in Microorganisms?" Ryohei Teraoka (1), Makoto Kanauchi (1), and Charles W. Bamforth (2). 1. Department of Food Management, Miyagi University, Hatatate Taihaku-ku Sendai Miyagi, 982-0215, Japan. 2. Sierra Nevada Brewing Company, Chico, CA 95928, U.S.A.]</ref>. See also [https://www.masterbrewerspodcast.com/229 MBAA Podcast episode 229 "Is Hop Creep Caused by Microorganisms?" with Dr. Charlie Bamforth]. [https://www.mdpi.com/2311-5637/7/2/66/html Bruner et al (2021)] investigated whether or not yeast strain selection has an impact on hop creep with the goal of finding if any strains would reduce the potential for dry-hop creep. They examined 30 different ''Saccharomyces'' yeasts from different yeast labs, and measured the [https://beerandbrewing.com/dictionary/gaKDNn0yxE/ real degrees of fermentation (RDF)] that occurred after dry-hopping post fermentation. With the exception of two strains, all of the yeasts re-fermented the beer after dry hopping, indicating that yeast strain selection won't reduce the chances for dry hop creep. There were two exceptions to this. The first was a diastatic strain of ''Saccharomyces cerevisiae'' (SafAle™ BE-134), which was able to break down starches in the beer prior to dry hopping due to the [[Saccharomyces#Diastatic_strains_of_Saccharomyces_cerevisiae|diastatic properties]] of this yeast strain. The other exception was a strain of ''Saccharomyces mikatae'' that is used as a co-fermenter in wine fermentations and is a poor attenuater of wort. The study also found no correlation between flocculation and attenuation from dry-hop creep, but suggested that further research be done to investigate the belief that higher flocculating strains could reduce the negative effects of dry hop creep <ref>[https://www.mdpi.com/2311-5637/7/2/66/html Bruner J, Marcus A, Fox G. Dry-Hop Creep Potential of Various Saccharomyces Yeast Species and Strains. Fermentation. 2021; 7(2):66. https://doi.org/10.3390/fermentation7020066.]</ref>.

===Aged Hops in [[Lambic]] and Other [[Spontaneous Fermentation|Spontaneous Fermentation Beer]]===

Modern lambic traditionally uses aged hops at a moderate rate to help limit and select for microbes and regulate acid production. Modern Lambic brewers cite rates in the range of roughly 450 grams of hops per Hl hectoliter of finished beer (0.6 ounces per gallon) <ref name="Jean Van Roy on Basic Brewing Radio"> [http://hwcdn.libsyn.com/p/e/a/2/ea26e00136fe1638/bbr05-30-13cantillon.mp3?c_id=5723890&expiration=1443888327&hwt=8dd886677defabdd73669cdc262ef446 Jean van Roy on Basic Brewing Radio] </ref> (~43 min in) (see also the notes pertaining hopping rates on the [[Cantillon]] page), with some brewers possibly going above this range. The age of hops used depends on the producer and their preferences/stock. Cantillon uses hops that are roughly 3 years old<ref>D. Janssen personal communication with Jean Van Roy, 9-Nov-2013</ref>, while 3 Fonteinen reports using hops that are over 10 years old<ref name="Drie Fonteinen on Belgian Smaak"> [http://www.belgiansmaak.com/armand-debelder-michael-blancquaert-drie-fonteinen/ Drie Fonteinen on Belgian Smaak] </ref> (~48 minutes in). Jester King reported using 0.66 - 0.75 pounds of whole leaf aged hops per BBL (0.34-0.39 ounces per gallon) in their spontaneously fermented ales <ref>Averie Swanson. "Sour Power! A Pro Brewer Spontaneous Fermentation Roundtable". HomebrewCon seminar. 2018.</ref> (~31:00 mins in). Lambic brewers either add their hops while still collecting wort, sometime before the wort comes to a boil<ref>[https://www.facebook.com/groups/Lambic.Info/permalink/1831338433787524/ Video of Cantillon wort reaching a boil from Bill on Lambic.info]</ref> (also known as "first wort hopping"), or shortly after boil is reached<ref name="Drie Fonteinen on Belgian Smaak"/> (~48 min in). The hops are then boiled with the wort for essentially the full length of the boil <ref>[https://www.facebook.com/groups/MilkTheFunk/permalink/1593059604055581/?comment_id=1593928187302056&reply_comment_id=1593938693967672&comment_tracking=%7B%22tn%22%3A%22R9%22%7D Conversation with Dave Janssen on MTF. 02/24/2017.]</ref><ref>[http://www.lambic.info/Brewing_Lambic#Hopping "Brewing Lambic", section "Hopping". Lambic.info website. Retrieved 02/24/2017.]</ref>. The resulting lambic beers are often surprisingly bitter, especially when young. Historically, there is [http://www.horscategoriebrewing.com/2016/04/hops-in-spontaneous-fermentation.html some evidence] that lambic brewers used a combination of aged hops and fresh dried hops. Not all aged hops are the same; different varieties/sources result in different levels of residual alpha/beta acids (probably not zero), oxidized acids, IBU's, perceived bitterness, and inhibition of lactic acid bacteria. Varieties with high acids and hop oils probably have more residual acids and oils, and aging times/conditions may not be ideal enough to completely age high alpha/beta/oil hop varieties. Therefore, it is impossible to give a blanket statement on how much aged hops to use given a specific lot of aged hops. Andrew Holzhauer from Funk Factory Geuzeria suggests tasting aged hops for bitterness and adjusting the amount of hops depending on how bitter they taste <ref>[https://www.facebook.com/groups/MilkTheFunk/permalink/2723907944304069/?comment_id=2723956477632549&reply_comment_id=2724104977617699&comment_tracking=%7B%22tn%22%3A%22R%22%7D Andrew Holzhauer. Milk The Funk Facebook group on how much aged hops to use. 06/13/2019.]</ref>, while James Howat from Black Project suggests making a small batch and having the wort/beer analyzed for IBU's and adjusting accordingly.

Homebrewer For example, homebrewer Caleb Buck performed an experiment comparing two different hopping rates for spontaneously fermented beer at home using whole leaf aged hops that were independently tested to have 0.5% alpha acids and 0.2% beta acids and were obtained from Hops Direct in Junuary 2016 <ref>[https://www.facebook.com/groups/MilkTheFunk/permalink/2302073339820867/?comment_trackingcomment_id=%7B%22tn%22%3A%22R%22%7D 2302314029796798&reply_comment_id=2303737466321121 Caleb Buck. Milk the Funk Facebook group thread on Caleb's aged hop experiment. 10/01/2018.]</ref>. The two rates tested were 0.3 ounces of aged hops per gallon and 0.6 ounces per gallon, both added at the beginning of the boil <ref>[https://www.facebook.com/groups/MilkTheFunk/permalink/3970005893027595/?comment_id=3970763046285213 Caleb Buck. Milk The Funk Facebook group. Addendum to when hops were added to the boil for his associated hop experiment. 10/14/2020.] </ref>. Samples of the two worts were sent to Sweetwater Science Labs to perform IBU analysis using the ASBC standard IBU test. Interestingly, the results were 72 IBU and 127 IBU respectively. The unexpectedly high IBU might be due to the variety of aged hop, as well as oxidized hop acids showing up in the standard IBU test (see Peacock's data [[Hops#Acids_2|here that showed that aged and fresh dried hops produce a similar IBU]]). After about 7 months, one of the 0.3oz/gal batches got down to a pH of 3.6, a second batch at 0.3oz/gal got to a pH of 4, while the 0.6oz/gal batches remained within a pH of 4.2 - 4.3. From this experiment, Caleb will attempt using only 0.15 oz/gal of aged hops which should be closer to 30 IBU and so that more acidity can be achieved. James Howat from Black Project Spontaneous Ales suggests making sample wort with the hops that will be used for a larger batch and sending that sample off for IBU testing in order to more easily achieve the desired IBU's. More detail can be found on [http://www.archaicpursuit.com/2018/08/2017-coolship-experiment-hopping-rate.html?m=1 Caleb Buck's collected data on cooling rates, acidity from hopping rates, and other collected data over a multi-year, multi-batch experiment] and [http://hwcdn.libsyn.com/p/b/d/2/bd2703ec2214a5c7/bbr09-27-18souribus.mp3 Caleb's interview on this experiment on BasicBrewing Radio].