100% Lactobacillus Fermentation
100% Lactobacillus Fermentation is the process of using a single pure strain of Lactobacillus to ferment with. This page will primary discuss the results of tests done by Lance Shaner from Omega Yeast Labs, as well as scientific explanations of why achieving full attenuation with Lactobacillus does not appear to be biologically possible. See the Lactobacillus page for more details on Lactobacillus in general.
Contents
Lance Shaner's Experiment
This experiment showed that fully attenuated (defined as >70% apparent attenuation) 100% Lactobacillus fermentations were not possible and those reported as such were in reality a result of unintentionally introducing yeast.
Procedures
Two Lacto strains from White Labs (delbrueckii and brevis) and Wyeast (buchneri and brevis), and the Omega L. plantarum strain were tested. All were grown on AOAC plates (a Lactobacillus medium) supplemented with cycloheximide. All Lacto cultures were cultivated in liquid AOAC at ~90°F (no stirring) to maximum cell density (achieved in about 3 days). 200 ml of each Lactobacilus was pitched into 2 liters of autoclave-sterilized unhopped wort with a starting gravity of 1.037. Each fermentation was incubated for 2 weeks at ~90°F, periodically checking gravity and pH.
Results
| Lacto Species/Strain | Final pH | Appt. Specific Gravity (Anton Paar Alcolyzer) | ABV (Anton Paar Alcolyzer) |
|---|---|---|---|
| White Labs L. brevis | 3.14 | 1.03386 | 0.10% |
| White Labs L. delbruekii | 3.72 | 1.03255 | 0.29% |
| Wyeast L. brevis | 3.72 | 1.03386 | 0.18% |
| Wyeast L. buchneri | 3.61 | 1.03279 | 0.24% |
| Omega Yeast Labs L. plantarum | 3.17 | 1.03394 | 0.10% |
None of the cultures, whether heterofermentative or homofermentative, produced a significant gravity drop. None produced a krausen or obvious CO2 evolution. All samples were sent off to be tested for alcohol levels using the Anton Paar Alcolyzer test. All strains showed some alcohol production, indicating heterolactic fermentation. The wort was fermented with foil covering the flasks, so it was not an anaerobic environment [1]. Note that the pH readings of this test correspond with the results of Matt Humbard's pH tests with Lacto.
Bryan Heit's Biology Explanation
"This is what I would have expected - those claims of complete attenuation always rubbed me the wrong way, and from a scientific point of view, appeared impossible. pH will (eventually) inhibit even the most acid-tolerant of bacteria. Even if the bug is capable of protecting itself from the degradative nature of acids (which many can), at some point the concentration of protons (acid "atoms") will block the bacteria's proton motive force, which is used to "pump" molecules into bacteria (and in the case of some bacteria, to generate energy). Once the proton motive force is gone, the bacteria is effectively dead.
Bacteria setup a proton motive force by pumping protons out of their cytosol, into the extracellular space. This acidifies the extracellular space and alkinalyzes the cytosol. The energy of this gradient can then be used to "force" other molecules into the cell - e.g. Lactobacillus uses this to collect amino acids and DNA for growth and replication. However, the more acidic the environment, the more energy is required to pump additional protons out of the cell. The ability of cells to do this is finite; the theoretical maximum "pump force" that can be exerted is equal to the energy of hydrolysis of one ATP (energy molecule); roughly -31 kJ/mol. The theoretical max acidity that will work against is 1.2, but in reality these pumps are generally less than 30% efficient, so for most species pumps fail at pH's above 3.0. Most Lactobacillus crap out somewhere around 3.2 to 3.0. The pH of a fully attenuated 1.040 wort would be between 1.9 and 2.4 (ignoring buffering effects), depending on whether the Lactobacillus was homo or heterofermentive. So the claim that Lactobacillus can completely ferment wort doesn't seem to be biologically plausible." - Comments by Bryan Heit of Sui Generis Blog [2]