Yeast
Yeast Management
Numerous production steps in the brewing process are affected by the yeast management or, in turn, affect it. A steadily high beer quality and a high brewery productivity greatly depend on the optimum handling of yeast. Brewing yeast management includes strain storage (in a culture collection), propagation, cropping, culture storage and acid washing, and this leads to wort fermentation itself.
Counting Yeast Cells
Pitching rate can be determined by measuring the yeast quantity by several methods, such as dry weight, turbidity, and cell counting (either visually using a hemocytometer or electronically). Especially important is also to assess viability (the percentage of live cells in a sample) and vitality (a function of total cell viability and the physiological state of the viable cell population).ntroduced back into the process, the ethanol is oxidized back into acetaldehyde.
Hemacytometer
A hemacytometer is a graduated counting chamber that can be viewed under a microscope to deter-mine the concentration of yeast cells in suspension. It consists of a heavy glass slide with two counting chambers, each of which is divided into nine large 1 mm squares, on an etched and silvered surface separated by a trough. The engraved grid on the surface of the counting chamber ensures that the number of yeasts in a defined volume of liquid is counted. The hemacytometer is placed on the microscope stage and the cell suspension is counted.
Automated Methods
The most common method for determining yeast cell number and viability is manual counting on a standard microscope using a hemacytometer and a viability dye. One advantage of this method is that visual inspection of each sample allows the operator to check for contamination, presence of interfering debris, and obvious dilution errors. A major disadvantage is that the manual method is laborious, error-prone, and the data acquired is not easily traceable.
Fluorescence Microscopic Imaging. Image-based cell counters utilize bright-field or fluorescent microscopes coupled with digital cameras to obtain images that are then analyzed with image analysis software. These automated cell counters are either self-contained (internal PC) or connected to an external computer.
Flow Cytometers. Flow cytometers are used to measure the characteristics of individual cells and particles that are delivered in a flow system past a point of measurement where light is focused on one cell or one particle at a time.
Radio-Frequency Impedance Spectroscopy. Radio-frequency impedance spectroscopy is based on applying an electric field to a cell suspension and creating a charge separation within each cell. The charged ions within each cell cannot move past an intact plasma membrane.
Pitching Rates
The brewer’s ability to pitch the correct number of yeast cells to initiate fermentation is crucial to consistently producing a product of superior and constant quality (Section 12.1). Pitching rates are governed by a number of factors, including yeast strain, fermentation capacity of the yeast, yeast viability, flocculation characteristics, previous history of the yeast, and desired beer flavor characteristics. Other considerations when choosing pitching rates include wort gravity, wort constituents, fermentation temperature, and the degree of wort aeration.
Preservation of Stock Yeast Culture
Once yeast has been selected, accepted, and fully proven for use in brewing, it is essential that a pure culture (working master culture) is maintained in the laboratory yeast bank for prolonged periods. Some small breweries do not maintain a master culture but rather purchase fresh slants for each in-house propagation cycle or hold stock cultures at independent third-party institutions.
Sub-Culturing
This traditional and popular method involves the use of two vials—one for transfer and one for laboratory use, for inoculation to scale up the culture for plant use. The cultures are maintained on a yeast growth medium (such as MYGP, PYN), incubated between 20 and 30 degrees C (68–86°F) to stationary phase (about 72 hours), and then stored for up to 6 months at 1 to 4 degrees C (34–39°F).
Desiccation
This method uses purified silica gel as a desiccant, or squares of Whatman No. 4 filter paper (Kirsop, 1991).
Lyophilization
Lyophilization or freeze-drying is another popular technique among research laboratories and culture collections, but less so for brewers. Cultures are rapidly frozen followed by drying under vacuum.
Freezing in Liquid Nitrogen
With this method pure cultures are kept in vials and submerged in liquid nitrogen (‒196°C, -320°F), thereby maintaining viability and genetic integrity for tens of years.
Yeast Cropping
Aside from the need to remove most of the yeast from the beer prior to conditioning, yeast recovery for reuse in subsequent fermentations is an important process in the brewery (Section 12.9).
Traditional Lager Fermentation System
The traditional method of collecting lager yeast is to decant the “green” (unconditioned) beer from the settled yeast on the bottom of the open fermenters.
Traditional Ale Fermentation System
Traditionally, top-cropped ale yeast was harvested by skimming the head of yeast/foam that formed on top of the beer in the shallow, flat-bottomed fermentation vessel.
Cylindroconical Fermenters
Today, with the advent of cylindroconical fermenters, the differentiation on the basis of bottom and top cropping has become less distinct. Cylindroconical tanks allow improved yeast separation and collection strategies for both lager and ale yeast (Figure 4.4). The angle at the bottom of the tank allows the yeast to settle into the base of the vessel at the completion of primary fermentation.
Culture Contamination
It frequently happens that brewing yeasts carry a persistent low level of contaminants such as Obesumbacterium proteus, acetic acid bacteria, and slow-growing Torula-type yeasts. These organisms are generally regarded as harmless because their numbers never reach a point where they are likely to have adverse effects on the beer. On the other hand, L. pastorianus, Z. anaerobia, and S. carlsbergensis are strains considered harmful at low levels.
Microscopic Examination
Microscopic examination of the yeast culture can be useful in assessing the overall health of the population. Abnormal-looking or irregularly shaped yeast cells are signs of cell stress, possibly indicating potential problems with wort composition, aeration, poor yeast handling, or fermentation conditions.
Yeast Washing
Pitching yeasts collected from brewery fermentations are never absolutely free of microbiological infection. In spite of whatever care and sanitary precautions are taken, some bacteria will contaminate the pitching yeast. It is important to remember that washing will not remove wild yeast—only bacteria are removed—as bacteria are more susceptible to a lower pH than yeast. The pitching yeast can contain healthy yeast cells and trub (dead yeast cells and organic residues) and may contain 5 to 15 percent dry solids (Lewis et al., 2001).
Distilled or Sterile Water Wash
In the first method, the yeast slurry and cold, distilled or sterile water are mixed thoroughly in a decantation tank. The yeast is allowed to settle and the supernatant water is decanted, taking with it dead cells, trub, grain, and hop particles. The flocculent nature of the yeast makes yeast flocs denser than contaminants. The process may be repeated several times.
Acid Wash
The second method is to wash the yeast with acids, e.g., tartaric, citric, sodium metabisulfite, sulfuric, or phosphoric acid which is typically the most commonly used acid. Tartaric acid may tend to make yeast more flocculent, while phosphoric acid—though very popular with craft brewers—may tend to make it powderier, with corresponding effects on degree of attenuation. Tartaric acid is the safest of the acids to use, but all of these acids are equally effective. Acid washing lowers the pH of the yeast slurry to the point at which bacteria and weak yeast cells are killed off, but it does not harm the healthy yeast cells.
Acid Wash with Ammonium Persulfate
Some brewers use an acid-persulfate combination rather than just acid claiming that it is a more effective treatment than treatment with acid alone. Briggs recommended the addition of a strong oxidizing agent as ammonium persulphate (0.75% w/v) with phosphoric acid (Briggs et al., 2004).
Chlorine Dioxide
Chlorine dioxide, an alternative to distilled water or acid washing, is relatively new to the brewing industry and is gaining acceptance as a method for washing yeast. It kills via microbes by reacting chemically with sulfur-containing amino acids, the building blocks of protein which are used to form cell membranes.
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