SOME EFFECTS OF WORT COMPOSITION ON THE RATE AND EXTENT OF FERMENTATION BY BREWING YEASTS

The time required to ferment worts of varied composition to a given extent is dependent upon the extent of exponential growth in the early stages of fermentation; in the worts studied this is determined by the concentration of assimilable nitrogen. When the concentration of all the non-carbohydrate nutrients in malt wort is halved by dilution with carbohydrate, the addition of appropriate quantities of serine or arginine restores the rate of fermentation to that of the malt wort. Minor nutrients, other than amino acids specifically required by the yeasts used, are thus present in at least two-fold excess in the malt wort. The yeast produced during exponential growth in malt wort (sp.gr. 1·040) is able to ferment rapidly much greater quantities of fermentable carbohydrate than are present in that wort. The majority of the strains of yeast examined ferment equally well when either glucose or maltose is added to malt wort and do so whether the sugar is added prior to fermentation or towards the end; however, one strain fails to ferment satisfactorily if a substantial quantity of glucose is added to wort prior to fermentation, because of the subsequent failure of the yeast to adapt to ferment maltose. It is suggested that most brewing strains do not require to adapt to maltose utilization during the fermentation of wort.     

FERMENTATION OF MINOR WORT CARBOHYDRATES BY BREWING YEASTS

Typical worts prepared from all-malt grists and from those using a proportion of adjuncts contain maltulose, maltotriulose, isomaltose, panose and isopanose at concentrations between 0·01 and 0·12%, and maltotetraose between 0·2 and 0·7%. Maltulose and maltotriulose are fermented by all brewing strains of Saccharomyces cerevisiae so far examined, soon after utilization of maltose and maltotriose respectively begins. Fast-fermenting strains absorb small amounts of panoses and certain of them remove some maltotetraose whereas slow fermenting strains fail to utilize these three sugars. The fast-fermenting strains utilize more isomaltose than do slow fermenting yeasts. Of the total (2–4 g/litre) minor sugars of wort, fast fermenting strains remove about 90% whilst slow fermenting strains absorb 70–80%.     

FERMENTATIVE CAPACITY OF YEASTS WHICH METABOLIZE MALTOTRIOSE RAPIDLY

A strain of Saccharomyces carlsbergensis which removes maltotriose particularly rapidly from wort exhibits a very high rate of fermentation of this sugar in isolation. This rate is quickly established after pitching and is maintained throughout the greater part of a batch fermentation despite a rapid decline in the ability of the cells to ferment glucose. This same strain and certain others of the same species which also remove maltotriose readily from wort are able to ferment the trisaccharide as fast as or even faster than maltose after growth on the disaccharide as sole carbon source; this property is usually accompanied by repression of glucose fermentation. Evidence is presented which indicates the presence, in these strains, of two or more α-glucosidases having different relative affinities for the oligosaccharides.     

PRODUCTION OF YEAST MASS DURING FERMENTATION OF WORT BY BREWERY YEASTS

The quantity of yeast dry matter present in fermenting wort increases throughout the period of sugar utilization. During the period of exponential production of new cells and yeast mass the yield of yeast dry matter per mol of hexose utilized (Y_hexose) does not vary significantly for the yeasts and worts examined. In the post-exponential phase, however, Y_hexose declines, to an extent which varies with strain and which is particularly marked if exponential growth has previously been restricted by deficiency of non-carbohydrate nutrients. The total yield is primarily determined by the concentration of the nutrient controlling the extent of exponential growth.     

THE INFLUENCE OF pH AND OF SELECTED CATIONS ON THE FERMENTATION OF MALTOSE AND MALTOTRIOSE

The fermentation of maltose and maltotriose in metal-free buffer solutions strongly depends on the pH value; the optimum pH is close to 5·0 and activity diminishes to zero above pH 7·2. Potassium stimulates utilization of these sugars at pH values of 5·0 and below, but in contrast to the situation with glucose, pH still influences fermentation markedly in the presence of potassium. Both pH and potassium influence sugar uptake rather than internal metabolism. Zinc, magnesium and ammonium ions stimulate utilization of both sugars by stimulating uptake mechanisms.     

PRODUCTION OF DIACETYL, 2-ACETOLACTATE AND ACETOIN BY YEASTS DURING FERMENTATION

Results are given for the production of diacetyl, 2,3-pentandione- I-acetolactate, 2-acetohydroxy butyrate, acetoin and 3-hydroxy-2-pentanone for 13 yeasts belonging to the genus Saccharomyces. The data presented indicate that in general the yeasts do not produce significant amounts of vicinal diketones during unstirred laboratory fermentations. The observed maximum levels of diacetyl ranged from 0·02–0·32 ppm averaging 0·09 ppm (0·07 ppm if S. carlsbergensis (strain H) is omitted). During active fermentation all the yeasts tested produced 2-acetohydroxy acids. The 2-acetohydroxy acids and the 3-hydroxy-2-ketones are determined gas chromatographically by stepwise conversion of the four compounds to diacetyl and 2,3-pentandione. The maximum amounts of 2-acetolactate produced by the yeasts tested averaged 0·7 ppm, ranging from 0·08–1·65 ppm.     

STUDY OF THE ASSIMILATION OF PROLINE BY BREWING YEASTS

The problem of the incomplete assimilation of proline by brewing yeasts has been approached by studying growth on synthetic media under various conditions, by examining a catabolic reaction which could explain non-degradation under anaerobic conditions and by following the penetration of l -¹⁴C proline into the yeast cells. Demonstrations have been made that proline oxidase, the enzyme responsible for the first catabolic reaction of l -proline, requires the participation of cytochrome c to operate and therefore needs the presence of molecular oxygen, and that the permease responsible for the transport of proline is, under normal brewery conditions, repressed by other amino acids of wort. Permease derepression, which should normally occur after the complete consumption of the other amino acids, is made impossible because of the strong anaerobic conditions achieved at that stage.     

THE DIFFERENTIATION OF PRIMARY BREWING YEASTS BY IMMUNOFLUORESCENCE

The application of immunofluorescent methods to differentiate between primary brewery strains of Saccharomyces cerevisiae in current commercial use has been investigated. It was found possible to distinguish between strains which always sediment at the completion of fermentation and those which are capable of forming top crops. Attempts to differentiate between top-cropping strains were largely unsuccessful.     

QUANTITATIVE ASPECTS OF THE CONTROL BY OXYGENATION OF ACETATE ESTER CONCENTRATION IN BEER OBTAINED FROM HIGH-GRAVITY WORT

A single oxygenation of fermenting wort controls the otherwise excessive synthesis of ethyl acetate and iso-amyl acetate which may occur during the fermentation of wort of high specific gravity; ethyl acetate levels are more easily reduced than those of iso-amyl acetate. The effect of oxygen on acetate ester levels can be attributed to (a) the virtual suppression of ester synthesis during treatment and (b) the reduced rate of synthesis in the period following treatment. The latter effect is of greater mangitude if oxygenation is timed correctly. For an initial specific gravity of 1.080, acceptable levels of the acetate esters may be obtained by oxygenating for two hours whilst the specific gravity of the fermenting wort is in the range 1.063–1.044. The quantity of oxygen which must be supplied during this time has been defined in relation to the extent of attenuation which is desired and the effect of changing the rate of oxygen supply has been assessed. The rate of fermentation is greatly increased by oxygenation.     

THE VALUE OF LONG-CHAIN FATTY ACID COMPOSITION IN THE IDENTIFICATION OF SOME BREWERY YEASTS

The cellular long-chain fatty acids of 72 strains representing 29 species of brewery yeasts were extracted by saponification and analysed as methyl esters by gas chromatography. With this method, it was possible to distinguish between Saccharomyces cerevisiae and five other groups from the brewery industry, only four hours after they were obtained from 48 h old cultures under standard conditions. This compares favourably with the 7 to 10 days for conventional methods.     

ASPECTS OF FERMENTATION IN CONICAL VESSELS*

Cylindro-conical vessels are widely used for the fermentation of ales and lagers. These fermentors have the advantage of shorter fermentation times and they provide that measure of flexibility so necessary in breweries which produce a number of brands. It is usual to use a bottom-cropping strain of Saccharomyces cerevisiae as this results in easier yeast cropping, lower losses of bitterness and enhanced foam stabilities. Cylindro-conical vessels are not the cheapest of available fermentors but they allow the brewer to take maximum advantage of advances in technology. The fast rate of fermentation in cylindro-conical vessels can lead to the practical difficulty of controlling foam formation. However, by optimizing temperature, α-amino nitrogen concentration, yeast concentration, pressure and dissolved oxygen level the rate of fermentation and formation of foam can be controlled. The ability to control foam height allows vessel contents to be increased. The foregoing applies to both normal gravity and high gravity fermentations in which fermentation times and yeast growth values can be brought within the same ranges.