The two faces of microorganisms in traditional brewing and the implications for no- and low-alcohol beers.

The production of alcoholic beverages is intrinsically linked to microbial activity. This is because microbes such as yeast are associated with the production of ethanol and key sensorial compounds that produce desirable qualities in fermented products. However, the brewing industry and other related sectors face a step-change in practice, primarily due to the growth in sales of no- and low-alcohol (NoLo) alternatives to traditional alcoholic products. Here we review the involvement of microbes across the brewing process, including both their positive contributions and their negative (spoilage) effects. We also discuss the opportunities for exploiting microbes for NoLo beer production, as well as the spoilage risks associated with these products. For the latter, we highlight differences in composition and process conditions between traditional and NoLo beers and discuss how these may impact the microbial ecosystem of each product stream in relation to microbiological stability and final beer quality.

The role of inorganic-phosphate, potassium and magnesium in yeast-flavour formation.

Inorganic-phosphate, potassium, and magnesium are key-minerals required for yeast growth, metabolism, and survival, the present work investigated its impact in yeast-flavour formation using a multi-factor experimental design, which was used to generate a range of phosphorous-potassium-magnesium resulting in a 28-point D-optimal design. Samples were evaluated using HPLC (ethanol), GC-MS (aroma), and CountStar (total yeast cell). Results revealed that yeast requires a minimal amount of inorganic-phosphate, potassium, and magnesium (250, 500, and 70 mg/L, respectively) to support yeast-growth and ethanol/flavour formation. Inorganic-phosphate was important for fatty acid esters formation/short chain fatty acid (SCFA) reduction. Potassium was important in the formation of acetate esters/higher alcohols. Magnesium was the most important inorganic element for ester formation/SCFA reduction; furthermore, ethanol production is magnesium-dependent. In conclusion, inorganic phosphate, potassium and magnesium play an important role in yeast-growth, esters and higher alcohols formation; and SCFA reduction. Ethanol formation is Mg-dependent.

Influence of essential inorganic elements on flavour formation during yeast fermentation.

The relative concentration of available inorganic elements is critical for yeast growth and metabolism and has potential to be a tool leading to directed yeast flavour formation during fermentation. This study investigates the influence of essential inorganic elements during alcoholic fermentation of brewers wort, fermented using three independent yeast strains, Saccharomyces pastorianus W34/70, and Saccharomyces cerevisiae strains M2 and NCYC2592 under a range of conditions replicated for each yeast strain. 10 treatments were applied: 1 control and 9 inorganic supplementations: standard brewers wort, ammonia-nitrogen, inorganic phosphate, potassium, magnesium, copper, zinc, iron, manganese and a composite mixture, Twenty-five chemical markers were evaluated by HPLC (ethanol, glycerol), and GC-MS (aroma). There was a significant change in volatile aroma compounds during fermentation, which was more prominent when supplemented with ammonia nitrogen, inorganic phosphate, potassium or magnesium (P < 0.05). Heavy metal ions mostly had a negative effect on the flavour formation.

Physiological and transcriptomic response of Saccharomyces pastorianus to cold storage.

Removal of yeast biomass at the end of fermentation, followed by a period of storage before re-inoculation into a subsequent fermentation, is common in the brewing industry. Storage is typically conducted at cold temperatures to preserve yeast quality, a practice which has unfavourable cost and environmental implications. To determine the potential for alleviating these effects, the transcriptomic and physiological response of Saccharomyces pastorianus strain W34/70 to standard (4°C) and elevated (10°C) storage temperatures was explored. Higher temperatures resulted in increased expression of genes associated with the production and mobilisation of intracellular glycogen, trehalose, glycerol and fatty acids, although these observations were limited to early stages of storage. Intracellular trehalose and glycerol concentrations were higher at 4°C than at 10°C, as a consequence of the cellular response to cold stress. However, significant changes in glycogen degradation or cellular fatty acid composition did not occur between the two sets of populations, ensuring that cell viability remained consistent. It is anticipated that this data may lead to changes in standard practice for handling yeast cultures, without compromising yeast quality. This work has significance not only for the brewing industry, but also for food and biofuel sectors requiring short-term storage of liquid yeast.

Yeast responses to stresses associated with industrial brewery handling.

During brewery handling, production strains of yeast must respond to fluctuations in dissolved oxygen concentration, pH, osmolarity, ethanol concentration, nutrient supply and temperature. Fermentation performance of brewing yeast strains is dependent on their ability to adapt to these changes, particularly during batch brewery fermentation which involves the recycling (repitching) of a single yeast culture (slurry) over a number of fermentations (generations). Modern practices, such as the use of high-gravity worts and preparation of dried yeast for use as an inoculum, have increased the magnitude of the stresses to which the cell is subjected. The ability of yeast to respond effectively to these conditions is essential not only for beer production but also for maintaining the fermentation fitness of yeast for use in subsequent fermentations. During brewery handling, cells inhabit a complex environment and our understanding of stress responses under such conditions is limited. The advent of techniques capable of determining genomic and proteomic changes within the cell is likely vastly to improve our knowledge of yeast stress responses during industrial brewery handling.

Chitin scar breaks in aged Saccharomyces cerevisiae.

Ageing in budding yeast is not determined by chronological lifespan, but by the number of times an individual cell is capable of dividing, termed its replicative capacity. As cells age they are subject to characteristic cell surface changes. Saccharomyces cerevisiae reproduces asexually by budding and as a consequence of this process both mother and daughter cell retain chitinous scar tissue at the point of cytokinesis. Daughter cells exhibit a frail structure known as the birth scar, while mother cells display a more persistent bud scar. The number of bud scars present on the cell surface is directly related to the number of times a cell has divided and thus constitutes a biomarker for replicative cell age. It has been proposed that the birth scar may be subject to stretching caused by expansion of the daughter cell; however, no previous analysis of the effect of cell age on birth or bud scar size has been reported. This paper provides evidence that scar tissue expands with the cell during growth. It is postulated that symmetrically arranged breaks in the bud scar allow these rigid chitinous structures to expand without compromising cellular integrity.

The impact of brewing yeast cell age on fermentation performance, attenuation and flocculation.

Individual cells of the yeast Saccharomyces cerevisiae exhibit a finite replicative lifespan, which is widely believed to be a function of the number of divisions undertaken. As a consequence of ageing, yeast cells undergo constant modifications in terms of physiology, morphology and gene expression. Such characteristics play an important role in the performance of yeast during alcoholic beverage production, influencing sugar uptake, alcohol and flavour production and also the flocculation properties of the yeast strain. However, although yeast fermentation performance is strongly influenced by the condition of the yeast culture employed, until recently cell age has not been considered to be important to the process. In order to ascertain the effect of replicative cell age on fermentation performance, age synchronised populations of a lager strain were prepared using sedimentation through sucrose gradients. Each age fraction was analysed for the ability to utilise fermentable sugars and the capacity to flocculate. In addition cell wall properties associated with flocculation were determined for cells within each age fraction. Aged cells were observed to ferment more efficiently and at a higher rate than mixed aged or virgin cell cultures. Additionally, the flocculation potential and cell surface hydrophobicity of cells was observed to increase in conjunction with cell age. The mechanism of ageing and senescence in brewing yeast is a complex process, however here we demonstrate the impact of yeast cell ageing on fermentation performance.