Aeration and Yeast Starters

Background

The pitching rate of yeast is generally accepted to be one of the most important factors in fermentation performance and the resultant beer character. The often quoted “optimal” pitching rate is 0.75 billion cells per liter of wort, per degree Plato for ale, and 1.5 billion/L-°P for lager. However, at the homebrew level, commercial yeast cultures are not available with cells counts adequate for fermentation of a typical (i.e., 20 L at 13°P) batch of ale, let alone a high-gravity lager, and multiple “smack packs” or vials of yeast can easily exceed the total cost of the remaining ingredients. The method most commonly proposed for increasing cell counts is to use a stir plate and flask, with a sponge, cotton, aluminum foil, etc. on or in the neck. There are three reasons typically given:

1. Keeping the yeast in suspension mechanically increases the attenuation of wort sugars.
2. Removing the toxic CO₂ from solution promotes the growth of healthy yeast.
3. Allowing O₂ to diffuse into the starter head space maximizes yeast reproduction.

Some sources also suggest that the reduced pressure in a starter without an airlock will have a beneficial effect, but I find this claim dubious at best. The static head associated with the water in an airlock – assuming a 5 cm column – is given by:

     P = ρgh = (1000 kg/m³)(9.8 m/s²)(0.05 m) = 490 Pa

which is roughly 0.5% of atmospheric pressure at sea level. This is well within the error of the experimental measurements, and so any effect present is not expected to be observable.

Experimental Method

A total of five starters were fermented out, and the volume of the resultant yeast slurry measured:

     A: A control starter, with airlock.
     B: A control starter, with foil wrapped over the neck in place of an airlock.
     C: An airlock starter, which was swirled frequently to simulate a stir plate.
     D: A foil starter, which was also swirled.
     E: A foil starter, which had air added via an aquarium pump and airstone, as often as foaming would allow.

For a starter medium, ordinary granulated table sugar, which is effectively pure sucrose, was used. Sucrose was chosen not only for its low cost, but also because using DME or wort from a mash would necessarily introduce some amount of trub (hot break and cold break), which would be measured along with the slurry and could introduce some non-systemic experimental error. The use of sucrose should also allow the yeast to consume 100% of the sugars in solution, eliminating the “fermentability” of the media as a variable. Finally, using sucrose means that a refractometer can be used to take gravity measurements directly, without applying a “wort correction factor” or removing a hydrometer sample with a statistically significant volume. The starters were each made by dissolving 200 grams of sugar into 2 liters of water, which was then boiled for 5 minutes. The resulting starter solutions averaged 10.7 Brix. In order to provide the nutrients that would otherwise be lacking in an all-sugar starter, two packets of yeast (14 g) were boiled for 5 minutes in water, then topped off to 250 mL total volume. Prior to pitching, this suspension was shaken, and 50 mL added to each starter.

Into each starter, one 7 gram packet of Red Star bread yeast was added dry. Bread yeast was chosen primarily for its low cost; as a strain of S. cerevisiae, its performance in a starter should be essentially identical to any ale yeast. No effort was undertaken to control the fermentation temperature in the starters, other than placing them in a room with a household thermostat set for 68°F (20°C). To accelerate fermentation as much as possible, the starters were placed directly in front of a heating vent, and observed air temperatures ranged from 20.4°C to 25.3°C. While not particularly well-regulated, the fermentation environment therefore emulates one which would be typical for a homebrewer. After fermentation was complete, as determined by identical refractometer readings on two consecutive days, the starters were placed in a cooler at 0°C for at least 24 hours, after which the majority of the liquid was poured off and a final gravity measured via hydrometer. The slurry was then resuspended and poured into a 250 mL graduated cylinder, and the jug rinsed once with tap water. The slurry was returned to the cooler and allowed to settle for another 72 hours before measurement. This does not provide perfect isolation of dense slurry, but again, in the absence of more sophisticated equipment (a centrifuge, e.g.) it emulates the methods available to a typical homebrewer.

Finally, the slurry volumes were converted into approximate cell counts by assuming 100% viability and a cell density of 3.8 billion/mL. This inevitably introduces a great deal of uncertainty, but true cell counts are not achievable without a cytometer. From a practical perspective, a variation in pitching rate of 20%, or even more, is probably of negligible importance in brewing.

Observations

As I had only two one-gallon glass jugs (my normal starter vessels) available, the experiment was conducted in three stages. The controls, Starters A and B, were fermented first. Some variations were immediately apparent. The krausen at the surface of Starter A consisted of a thick, uniform layer of large bubbles, whereas Starter B displayed a patchier covering of comparatively fine bubbles. Visible fermentation was completed more quickly in A, with krausen having dissipated completely after 44 hours, although the airlock continued to bubble until about eight days after pitching. Both A and B also had bubbles of what I assume is CO₂ coming out of solution on the glass, indicative of a supersaturated solution. In B these bubbles disappeared at roughly the same time as the krausen; in A they persisted for about six days.

Starters C and D were fermented next. To simulate the effects of a stir plate, these starters were agitated in a vigorous circular motion for approximately 15 seconds, approximately every 15 minutes, 12-18 hours a day. Clearly this is not a perfect analog for a stir plate starter, but it was observed to be sufficient to keep yeast from collecting in the bottom of the glass jug between periods of agitation. Again, marked differences in the surface appearance of the starters were apparent. Within a few minutes of swirling, bubbles began to appear at the surface of D (the foil starter); C remained clear between periods of agitation, although airlock activity resumed quickly. When swirled, C also produced fewer and coarser bubbles than D. After being measured, the slurry from D was inadvertently left in the graduated cylinder for an additional 12 days; the value of 3.8 billion/mL was obtained by assuming this allowed for full compaction to 4.5 billion/mL.

Finally, starter E was aerated using an “Elite 800″ model aquarium air pump and a plastic aquarium air stone. The pump is rated for 2.0 W and 2.5 psi. Two 0.45 micron syringe-type filters were used in series to ensure sterility. The starter was aerated continuously for eight hours after pitching, and thereafter for approximately one minute in fifteen, 12-18 hours a day. This was the maximum duty cycle that was possible without the vessel overflowing. Qualitatively, this seems to be much less foam than would be expected from a wort starter of similar volume and gravity. This is sensible, given that the sucrose medium lacks most of the proteins associated with malt-based wort.

Results

Starter B produced approximately 8% more yeast than A. Although a rigorous calculation of the experimental error was not conducted, it is almost certainly within the error bar of the graduated cylinder measurements (±0.9 mL), and so the results for the control starters offer significant support for the hypothesis that access to atmospheric oxygen increases cell growth in a yeast culture. At least part of the increase, however, can be attributed to an overall more thorough fermentation in B, which exhibited approximately 2% greater attenuation than A.

Interestingly, although the “stir plate” starters did produce significantly larger volumes of slurry, they were not nearly as large as other sources suggest. For example, MB Raines observed a four-fold increase over a starter which was shaken. The Mr. Malty™ calculator seems to assume a factor of 1.27 (for shaking) or 1.53 (for a stir plate) versus a “simple starter”. The experimental results, however, show an increase of 17% for the foil starters, and only 8% when using an airlock. It is also worth noting that C and D exhibited identical attenuation, which was statistically equal to the attenuation of A.

Finally, starter E, the aerated sample, produced slightly less slurry than D (about 5% less), which is still 12% more than the “simple starter”. Again, however, the increase is not in line with others’ results; Raines and Zainasheff report increases of 50% and 35%, respectively. The final refractometer reading for E was 0.3 °Bx higher than any other starter; I hypothesize that this is due to the liquid being saturated with air, and the fact that the hydrometer-measured gravity was also 0.5 “points” higher than the other starters would seem to bear this out. The attenuation of E was, however, roughly in line with starters A, C, and D – meaning B exhibited significantly higher attenuation than the other four samples. The reason for this discrepancy is unknown.

Conclusions

1. All other things being equal, a starter covered in foil will grow more yeast than one with an airlock.
2. The primary reason to use a stir plate is not the mechanical mixing of the starter, but the introduction of oxygen. Using an airlock significantly reduces the effectiveness of a stir plate.
3. Contrary to what other sources indicate, a stir plate does not produce several times as much yeast per unit volume.
4. Given that it can also be used to aerate the main batch of wort, an aquarium pump is probably a more cost-effective investment for a homebrewer than a stir plate.
5. In the case of a pure sucrose fermentation, refractometer estimates of final gravity correlate well with hydrometer readings, with a maximum discrepancy of 1.5 “points”.
6. Bread yeast (at least this brand) tends not to flocculate, and on that basis alone would be a poor choice for beer.
7. I feel sorry for 17 year olds. This stuff isn’t terrible, but I wouldn’t want to drink it.

 
 
Update: 06 Feb 2010

To try to determine how much impact using sugar in place of malt would have on the behavior of a starter, I made up a 2 L starter, using 215 g of DME. In all other respects it was treated the same as starter D. Call it F. F produced 88 mL of slurry. Assuming 5% of the volume is non-yeast solids, this equates to 318 billion cells. Adjusted for attenuation, that’s 93% more yeast per unit of sugar compared to the sucrose starter. Significantly, it is also essentially identical to the figure (314 billion) given by the MrMalty calculator, when set to “intermittent shaking”. However, for a stir plate starter the calculator predicts a total of 379 billion cells. From this I draw a few additional conclusions:

1. Access to free amino nitrogen can be a limiting factor in fermentations which are largely non-malt based.
2. Increasing cellular access to oxygen can to a limited extent compensate for low levels of FAN.
3. The MrMalty calculator is sufficiently accurate at predicting cell counts for brewing, but may over-estimate the effect of a stir plate.

 
 
Update: 04 Mar 2010

I fermented out one additional DME starter, this time treating it identically to the aerated starter, E. The resulting slurry measured 116 mL. Again, assuming 5% non-yeast solids, that’s 419 billion cells. Not only is this significantly more yeast – 32% more – than the agitated starter, it’s more than the MrMalty calculator predicts for any starter handling technique, including “continuous aeration”. In fact, it’s roughly the quantity needed for a 5 gallon batch of average-gravity lager. Some final conclusions:

1. Given the choice between a stir plate and aeration stone, the stone will make more efficient use of starter wort.
2. By aerating, homebrewers can grow substantial amounts of yeast (such as those required for lagers) without having to make inconveniently large starters.

 
 
Download the full experimental data:
starter_experiment.ods | starter_experiment.xls