What to Post after a Long Hiatus? An Experiment (Yeast Aeration) of Course!

My first post was in 2010, a standard bitter in honor of close colleague and astounding scientist that passed. I was in my fourth year as a post-doc at Columbia University studying how retroviruses (HIV, MLV) interact with the cells they try to infect. Time was in abundance, allowing me to brew in a small Bronx apartment while I gathered many friends and scoured the city for rare-to-find craft beers. I was able to ranch yeast and create a bank, isolate unique Brettanomyces strains, and collaborate with a world renown master brewer. Most importantly, I learned as much as I could about the science behind brewing beer. My time in academic research came to an end, and I married the most amazing woman I have ever known and met another one:

photo 1

Natalie, my beautiful daughter, was a game changer for me. I assume that most father’s sense of their place in the world changes dramatically when your first-born looks straight into your eyes. Job hunting was next, and I was lucky enough to find a great job doing process development for vaccines at Merck. Moving and settling in, enjoying fatherhood, and trying to secure my family’s future has made me feel slightly old.

As a result of all of this stuff, my last post was back in May. I began to re-read some of my old posts and realized that this blog has been is linked to so many wonderful memories, both beer and non-beer events that I can’t possibly ignore it anymore. It’s posts serves as markers for my past. It is this for this reason I intend to start posting again.

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And an experiment may be the best way to start posting again. As a recap, I began brewing experiment beers for a yeast class that I taught at Brooklyn Homebrew. Ben and Danielle were kind enough to provide resources and space for my scientific curiosities that kept brewing. In contrast to my work life, my style of experiments are not rigorous and grounded in statistical significance. The goal has always been to pique the interest of the vast majority of non-scientific brewers, both professional and amateur. More importantly, I do not have the resources at home to measure key components of the brewing process. Instead, my conclusions rely on the most important data point – the beer drinker.

I am teaching again, this time at a great homebrew store in PA – Keystone Homebrew. Jason Harris, along with Lou (store manager) has been kind enough to provide support and space for the yeast class and the experiment beers. I’ve already taught two classes, and the most recent one was this past Saturday (November 9th).

Oxygen

Yeast are facultative anaerobes, meaning that they prefer to use oxygen to produce energy efficiently through oxidative phosphorylation. In the absence of oxygen, yeast still produce energy but with the inefficient process of alcoholic fermentation. Luckily for brewers, yeast will almost always undergo fermentation due to the Crabtree effect, where in the presence of excess sugars, even if there is oxygen, yeast will produce ethanol and CO2.

The main reason yeast need oxygen is for the production of unsaturated fatty acids (UFA) and sterols. Both molecules make up the plasma membrane of the yeast cell. I won’t go into how they are synthesized, but sterols, such as ergosterol pictured below, is present in the cell membrane at much lower concentrations. Sterols  provides fluidity, making the membrane easier to bend and twist in shape.

Ergosterol_structure

Without sterols, UFAs would form rigid micelles, or droplets of fats. Importantly, this membrane fluidity allows for other vital proteins to be imbedded in the membrane. Some of these proteins are responsible for importing not only complex sugars (maltose, maltotriose), but vitamins, minerals, and cofactors. With poor cell membranes yeast will not tolerate increasing amounts of ethanol (which is why oxygenating that strong ale is crucial). Lastly, if the cell membrane is not healthy enough, bud scars from cell budding will take a toll on yeast health and restrict future growth.

yeast_cell_membraneWhat are the consequences of introducing too little or too much oxygen? It is best to think of it biochemically – that is, oxygen acts as an accelerant to certain biochemical pathways. For example, what I did not mention before is that O2 drives the synthesis of UFAs and sterols through a cofactor called Acetyl CoA.

acetyl CoA

I talk about this molecule a bit in my post on esters and it sits at the crux of critical yeast biochemistry. In this case, oxygen drives acetyl CoA to form lipids (UFAs, sterols). As more acetyl CoA is used to make UFAs, less is used to make esters and fusel alcohols. Therefore, beers that have more oxygen tend are cleaner in esters and fusels (think IPAs). Beers that have less oxygen will have more esters and fusels and are more fruity (think English ales). Increasing oxygen will grow more yeast cells as their membranes are super healthy and ready for budding. The increase in yeast biomass will add a bump in attenuation and may even thin out the beer since the yeast will be so active.

How much Oxygen?

Homebrewers have endless methods of adding oxygen such as shaking and splashing the wort, sloshing the wort from bucket to bucket, aeration with aquarium pump, and adding pure oxygen. I have heard lots questions since many homebrewers practice different techniques:

  • How long do I shake?
  • Should I use a drill with a whirl attachment?
  • How long do you aerate with a pump?
  • How long do you pump in pure O2? At what flow rate?

Let me answer all of these questions by saying that whatever works for your system that gives you the best beer is what you should do. Experimenting here is key. Brew that same pale ale three times, one with splashing, one with pure O2 (one minute), and one with pure O2 (five minutes).

In reality, as homebrewers it is very difficult not to introduce O2 once the wort is cooled. Simply transferring the cold wort into a carboy (without splashing) will add some amount of oxygen for the yeast to use, however this is far from ideal. Yeast need greater than 8 parts per million (ppm) to adequately ferment a batch of beer. Here are some numbers on different methods (keep in mind that O2 dissolves less in higher gravity wort) on an average batch of homebrew (5 gallons, 1.050 – 1.070):

  • Shaking: 2 ppm
  • Aquarium pump: maximum of 8 ppm
  • 30 seconds pure O2: 5 ppm
  • 60 seconds pure O2: 8-10 ppm
  • 2 minutes pure O2: greater than 14 ppm
    • The above pure O2 is assumed to be one liter per minute.

IMG_9929

Splashing the wort is a very ineffective method of introducing oxygen and may even introduce contamination. Notice that with an aquarium pump, air can only give you a max of 8 ppm, whether you pump for five or thirty minutes. Oxygen makes up only 21% of dry air so there is only so much O2 you can force into beer with this method. Moreover, 30 minutes of aquarium pumping may drastically reduce head formation since you will create so much foam. The only way to get enough oxygen into your wort is by using pure oxygen with a sintered stone.

Exactly how much O2 to add to your beer as this depends on many factors:

  • Yeast strain (different O2 requirements for different strains)
  • Wort gravity (need more O2 for higher gravity worts)
  • Beer style

Beer style and flavor profiles generated from oxygenation is the most important factor to me and is the basis of my experiment. For example, will adding less oxygen enhance fruity characters of some english ales? Would my IPAs be cleaner with higher amounts of O2? Would there be any changes in fusel alcohols? Would the body be thinner or fuller? Textbooks and the internet can tell you the answers, but I feel it is better to experiment and find out for yourself.

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For the experiment beer, I brewed an ESB in a ten gallon batch at Keystone Homebrew (OG: 1.056 with 40 IBUs):

  • 1 lbs Crystal Light – 45L (Thomas Fawcett)
  • 1 lbs Crystal Malt – 90L (Thomas Fawcett)
  • 8.0 oz Pale Chocolate Malt (Thomas Fawcett)
  • 12 lbs DME Golden Light (Briess)
  • 2.00 oz Target [11.00 %] – Boil 60.0 min
  • 2.00 oz Goldings, East Kent [5.00 %] – Boil 0.0
  • 1.0 pkg British Ale II (Wyeast Labs #1335)

DSC_0483

From this wort I split the batch four ways and provided different amounts of oxygen:

  1. 30 seconds of splashing the carboy
  2. 45 seconds O2
  3. 5 minutes O2
  4. 10 mgs of Olive oil

Sample (1) represents what a new homebrewer might do. I did not dump the wort between two buckets but rather sloshed the wort around in the carboy. This sample should have little dissolved O2. Sample (2) is my default oxygenation regimen at home – I usually add pure O2 for 1-2 minutes in a 5 gallon batch. Sample (3) was to observe any changes in the other direction with large amounts of oxygen added to the beer. When I did oxygenate this batch, the foam generated was huge, almost a 3 gallon tick mark. Unfortunately I did not have access to a medical grade O2 regulator and was unable to determine flow rate. I opened my regulator, what you typically find at homebrew stores, to its max setting.

DSC_0421Sample (4) may be the most interesting of all. I decided to mimic what Grady Hull, brewmaster at New Belgium, did for his masters thesis at Heriot-Watt University. The thesis can be found here and is generously hosted by brewcrazy.com. In a nutshell, he added dissolved olive oil to the yeast starter of a commercial batch of Fat Tire as a source of linoleic acid, a precursor to UFA synthesis and did not aerate the wort. The reasoning being that the oil provides all the UFA and sterol needs of the cell and you don’t need to add O2. The goal of the experiment was  to prolong beer half-life since oxygen has a severe staling effect. What they found was that the beer took a bit longer to ferment and was slightly more fruity. Independent tasters actually preferred this beer over the control beer.

For my experiment I added olive oil directly into the wort (no starter were made for each sample). Importantly, the amount of olive oil I added followed close to what Gary Hull did (1 mg per 25 billion yeast cells), which was exceedingly small (about a tenth the size of a pinhead). Moreover, olive oil will not dissolve in wort and must first be dissolved in 100% ethanol. For any homebrewers reading this – do not add a drop of olive oil to your beer. I had to weigh 1 gram of oil, dissolve it, and make a serial dilution until I had close to 0.1 ug/ml. Of this solution, I added 100 uls directly to the wort.

The beer is already fermented, bottled and was sampled at a yeast class this past Saturday (11/9/2013) at Keystone Homebrew. Similar to my past experiments, I will post flavor profiles from the students as data points in a future post.

Cheers!

17 Comments

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17 responses to “What to Post after a Long Hiatus? An Experiment (Yeast Aeration) of Course!

  1. Great writeup. I also use a homebrew regulator without a gauge to display how much oxygen I’m actually adding. I never fully open it up because of how much foam would be produced and how much oxygen I’d burn through.

    Oxygenation is an area that I’m interested in studying more as I prepare to brew a high ABV imperial stout. What are your thoughts on oxygenating at multiple times during primary fermentation on a high gravity (north of 1.100) batch?

    • Open it all the way up!!! ;)

      The oxygen canisters I use are around 8$ at Home Depot and last me a long time (greater than 1/2 year with about 1-2 batches per month).

      As per foam, it is an issue and there will be some head loss. I would rather attenuate my beer well though and take it as a price to pay. I find that my head, although not 4 inches of rocking foam, is still quite respectable with O2 that I add.

      As for higher gravity beers, you definitely can add O2 during fermentation to give an extra boost. I have done it before. Importantly, you want to oxygenate with 24 hours of pitching your yeast, or at the height of yeast activity. The reason is you do not want to oxidize your beer and the yeast needs to be super healthy to take up all the O2. If you add O2 in the tail end of fermentation (less that 1.040-1.045) yeast are sluggish, already beginning to flocculate, and will respond slower to utilizing O2.

      If you really want to boost your ABV, try adding some sugar and oxygenate 12-24 hours after pitching. Make sure you have a blow off setup!

      Best,

      J

  2. What is the source for your data on air pump aeration? White Labs measured and found that they were able to reach the same wort dissolved oxygen as with pure oxygen, which makes sense as the wort will eventually saturate, even with “only” ~20% atmospheric oxygen http://www.youtube.com/watch?v=2vELwUsBmWQ

    • Hi Roy!

      Thanks for commenting! I based my numbers from Chris White’s yeast book and use the same numbers that she shows in her presentation.

      Interestingly she does say that you can get 14 ppm using air, but you need to have a way to pressurize the wort to keep the oxygen from coming out of solution while you aerate. I would assume that the partial pressure of O2 in air is less than that of dissolved wort. In any case, I think it would be very hard to get carboy pressurized to the point where the partial pressure is higher than O2 dissolved in the wort.

      By using pure O2 you saturate the wort with O2. Sure you lose O2 over time, but not as fast as the yeast can consume it.

      • Roy

        Thanks Jason. It looks like the equilibrium level of dissolved oxygen in water is around 8 ppm. It wonder what the kinetics are of it dissipating vs. being consumed by yeast or reactions in the wort – probably pretty complicated.

  3. I’ve read the Grady Hull thesis (as well as discussed its finer points elsewhere on the web), and I think its important to point out the following:

    The goal of GH’s thesis project was not to establish olive oil as an alternative method to wort aeration. He assumed that is was a acceptable method of wort aeration based on previous studies with linoleic acid addition, which is a major constituent of olive oil’s lipid profile. One could argue that is a bit of a stretch in scientific thinking, but I won’t do that here.

    Instead, his work instead was asking the question: Using this alternative wort aeration scheme in a production brewery, do we encounter any deleterious effects from it (i.e. lagging fermentation time, off-flavors, etc.). His results show that olive oil did not have any such effects. However, it should be noted that this conclusion is MUCH different from saying that olive oil is an effective method of wort aeration. I think his study is often misstated to conclude the latter point.

    More to this point, in order to demonstrate that olive oil addition is an effective method of wort aeration, GH would have had to have a different negative control. In his study, he used New Belgium current aeration method as a negative control (i.e. in-line diffusion stone + pure O2). He showed that there was no difference in between this control and olive oil addition, and these data are often extrapolated to demonstrate that olive oil is a good replacement for conventional O2 injection. The assumption here is that NB’s aeration system actually has a measurable effect in the first place, and the results show that olive oil mimicks this effect perfectly. However it is possible that NEITHER method has a measureable effect, that those batches would ferment out just fine if non-oxygenated at all. The way the experiments were done, you can’t rule out this possibility.

    The only way to conclude that olive oil addition is an effective wort aeration method is to compare its effects to wort that receives no other treatment whatsoever. Granted, it is possible for the wort to pick up oxygen from other brewery processes (pumping from kettle to fermenter, for example), thus increasing the “background” to see an effect, but it is a better experiment than extrapolating from GH’s work. You could also look at a sample beer wort that is purged of most/all O2I am unaware of a method to do this (vacuum maybe?)

    FYI: here is an experiment by a homebrewer than specifically looked at the Olive oil addition to beer wort compared to a non-treated control. No differences observed in either fermentation or in taste by blind panel.

    http://hbd.org/discus/messages/43688/45581.html

    It is often overlooked by people who bang the drum of Olive oil addition for homebrewing, probably because the research wasn’t done through a university system and is somehow flawed. You and I both know that flawed results are certainly possible in the university system as well!!!

    Sorry for the long comment…

    • Hello BroadBill,

      First of all, thanks for the great comment. Lots of insight here.

      I’ll start off by saying that I worded my post to reflect no conclusions. I agree with you that the point of the thesis was not to suggest aeration could be supplanted by olive oil and even mention this in the post. In my view, they were interested in the shelf life of beer since oxidative potential of the beer is much higher than without aeration. My goal of doing this was to try and detect any changes in flavor profile compared to the different oxygen additions. However, when I post the results of the experiment in the coming weeks I will make sure not to suggest that homebrewers should use olive oil over oxygen, but rather let them come to their own conclusions. I will also point out the the added benefits to oxygen versus olive oil.

      For scientific discussion, here is how I would have done it:

      1) No aeration of any kind (negative control)
      2) Inline aeration only (postive control)
      3) Olive oil only in yeast storage tanks
      4) Olive oil only in yeast storage tanks with different olive oil concentrations
      5) Olive oil only in primary fermentor
      6) Olive oil only in primary fermentor with different olive oil concentrations
      7-10) Repeat samples 3-6 but with the addition of inline aeration
      11-18) repeat samples 3 thorugh 10 but use pure linoleic or oleic acid

      Obviously, the capacity to do all of these conditions might be too much for a commercial brewery and would have to be replicated in a scaled down model. Really we are dealing with two independent variables that need to be addressed. I would be curious to know if both olive oil and aeration had a synergistic effect, as in better fermentation profiles compared to any condition. Moreover, the olive oil was added to the yeast storage tanks and not the actual fermentation vessel. My small scale experiment is different in this respect. and would try both conditions. It would also be nice to pinpoint whichi fatty acid is contributing the desired effects as well.

      The conclusion and future suggestion work also brings up another interesting point, which I would like throw out there for debate if anyone wants to comment. It was suggested that the yeast storage tanks be aerated to bring glycogen and trehalose levels back up. As homebrewers we do this routinely with stirred yeast starters. So the question is if we are providing the starter with all the oxygen it needs to ferment a batch of beer, do we even need to aerate the wort?

      Cheers and thanks for commenting,

      J

  4. Great post. I am very interested in oxygenation and have used a range of techniques, including stainless airstone. I have now switched to using a “lance” to introduce O2 to the chilled wort as it exits a silicone tube from the chiller into the FV. I find that this works as well as the stainless airstone and has a number of advantages:

    1. It develops positive pressure in the FV so that pure O2 exits under pressure when the wort enters, thereby reducing potential airborne contamination
    2. It is a simple design
    3. It is easy to clean and sanitise
    4. It removes the extra step of oxygenating with the airstone as it is done during the chilling process

    I made a youtube video showing the technique:

    cheers
    steve

    • Steve,

      Love this and I might use this as I upgrade my brew system.

      Nicely done!

      J

    • Hey Steve,
      Pretty neat setup. I was curious though why you wouldn’t put the lance closer to the exit of the plate chiller. Seems to me the oxygen would be forced to take ride with the wort farther through the tube, and would easily double (or more with longer tubing) the oxygen contact time. You could then use a lower O2 flow rate to achieve the same results. Also, any sort of twists/turns/kinks would help create turbulence and act as a diffuser of sorts.
      – Dennis, Life Fermented Blog

      • Hi Dennis
        I considered it but was worried that introducing the O2 near the plate chiller further upstream would create some back pressure which would impede the flow of wort. However I take your point about it increasing the contact time. I think I feel an experiment coming on…
        cheers
        steve

  5. Jason,
    It is my quick and dirty understanding that oxygen is needed for membrane health, and that membrane health is adversely affected by the budding and splitting process as the yeast multiplies. Lacking the ability to properly oxygenate the wort, then, couldn’t one simply over-pitch? With today’s increasingly high quality and relatively cheap dry yeasts, its pretty easy to do. Even for a 1.100 beer, the Mr. Malty yeast calculator only recommends about 1.7 packets. It seems like if you just pitched 3 packets and shook the carboy a few times you would be good to go.

    Can you think of any reason this method wouldn’t yield similar results as properly oxygenating?

    – Dennis, Life Fermented Blog

    Sorry if this is a repost- my browser was being wonky.

    • Great point and the answer is both yes and no.

      Yes, I have read you could do this, in particular for high gravity beers. The effect would be similar to oxygenating your wort in that you would keep esters low. Interestingly, fusel alcohols increase as you oxygenate but not with high pitching rates, so you might be at a better advantage to not super-oxygenate your high gravity brew. At minimum, overpitching could potentially make up for lack of aerating. This is all from observation from others brewers and I would like to try something like for myself to really find out.

      Having said that, oxygenating your wort (even for high-gravity brews) is always a good thing. By overpitching your yeast you are essentially “setting” the amount of yeast that will ferment that batch of beer. The yeast will have less time to create biomass and adapt to the wort. As a result, attenuation might not be as high as you would expect. It has been shown that underpitching (even slightly) may increase attenuation since the yeast need to grow and adapt to the wort. My experiment on pitching rates reflects this and commercial breweries have seen this too.

      http://sciencebrewer.com/2012/03/02/pitching-rate-experiment-part-deux-results/

      Another factor is resistance to alcohol toxicity. At high levels of alcohol, yeast will stress and either die or go dormant. Providing adequate oxygen strengthens cell membranes to they can survive and ferment that much longer.

      To sum up (sorry for the long winded response), if you can’t oxygenate I would definitely overpitch. However I think of it terms of attenuation and oxygenating will definitely help, and would do this in addition to overpitching. This would keep esters down very low and the overpitch rate would tamp down fusels.

      Sorry if this was confusing (there is no right answer here),

      Jason

      • Awesome, exactly what I was looking for. I had listened to a few talks (Neva Parker and others) on the effects of pitch rate for achieving different results, and the effects of properly oxygenating wort, but none really touched on whether you could make up for one with the other.

        I finally broke down and purchased a regulator and 0.5um stone yesterday in preparation for an imperial russian stout. It’ll sure be a far cry from my first ever brew when I pitched one WLP004 vial into a 1.108 wort. Tasted like bubble gum for months!

        – Dennis, Life Fermented Blog

      • Glad to help. Be sure when you overpitch and oxygenate that you also keep fermentation temps down. Actually I would ferment cool for the majority of the time (65F) and slowly ramp up to 68-69F to get a bump in attenuation.

        Cheers,

        J

  6. chris bushnell

    I have used olive oil in starters, and grapeseed oil as well- pro-brewer in Asia has suggsted that grapeseed oil provides the same building blocks as EVO but is less likely to contaminate.
    that being said, I still use O2 with an airstone to oxygenate wort.

  7. Jeff

    Jason,
    Is the whole oxgenation issue over thought? While I definately have a bad habit of grossly oversimplifying issues, my beers turn out very drinkable (usually). Furthermore, we have been making beers for thousands of years and many of our beer heroes had tonsures for crying out loud. All that being said, I love to find ways to improve my beer and I look forward to seeing your results

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