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Lactic Acid Bacteria – the Uninvited but Generally Welcome Participants In Lactic acid bacteria – the uninvited (a) but generally welcome participants in malt whisky fermentation Fergus G. Priest Coffey still. Like malt diversity resulting in Lactobacillus fermentum and LEFT: (b) whisky, it must be matured Lactobacillus paracasei as commonly dominant species by Fig. 1. Fluorescence photomicrographs of whisky for a minimum of 3 years. about 40 hours. During the final stages when the yeast is fermentation samples; green cells Grain whisky is the base of dying, a homofermentative bacterium related to Lacto- are viable, red cells are dead. (a) the standard blends such as bacillus acidophilus often proliferates and produces large Wort on entering the washback, (b) Famous Grouse, Teachers, amounts of lactic acid. after fermentation for 55 hours and and Johnnie Walker which (c) after fermentation for 95 hours. (a) AND (b) ARE REPRODUCED will contain between 15 Effects of lactobacilli on the flavour of malt WITH PERMISSION FROM VAN BEEK & and 30 % malt whisky. whisky PRIEST (2002, APPL ENVIRON Both products use a The bacteria can affect the flavour of the spirit in two MICROBIOL 68, 297–305); (c) COURTESY F. G. PRIEST similar process, but here ways. First, they will reduce the pH of the fermentation we will focus on malt (c) through the production of acetic and lactic acids. This whisky. Malted barley is will lead to a general increase in esters following milled, infused in water at distillation, a positive feature that has traditionally been about 64 °C for some 30 associated with the late lactic fermentation. This general minutes to 1 hour and the effect is apparent in the data presented in Fig. 3 in which How does malt Scotch whisky – a multimillion pound wort is drained off into a fermenter or washback. The the concentrations of various esters are increased in non- whisky acquire its industry grain bed is rinsed with a second water at a higher matured (new-make) spirits from laboratory-scale unique flavour? The production of Scotch malt whisky is governed temperature (typically about 70 °C) to remove residual fermentation/distillations with and without lactobacilli. Fergus Priest by The Scotch Whisky Act (1988) which limits the nutrients from the fermentation. Finally, it is rinsed with However, lactobacilli might also produce specific flavour explains how ingredients to water, malted barley, whole grains of a third water at about 80 °C which is used as the mashing compounds that contribute in a unique way to the lactobacilli can play other cereals, yeast and caramel for colour adjustment. water for the next mash. The spent grains are removed they do not require oxygen for growth and flourish at flavour of the spirit. We have investigated these effects Interestingly, no mention is made of bacteria, yet we are for cattle feed. You will note that the wort is not boiled relatively low pH (pH 6 to about 3.5). This fits them well a crucial role. beginning to think that lactic acid bacteria help refine as it is in a brewery. This is to permit the enzymes from for growth in alcoholic beverages in which they have LEFT: MB-stained Fig. 2. General progress of malt the flavour of malt whisky in important ways. But first, the malted barley to continue to operate during the both beneficial (e.g. the malolactic fermentation of wine) Sugar whisky fermentation. ‘MB-stained’ a little background to the Scotch whisky industry. fermentation and to ensure complete hydrolysis of starch and spoilage effects. Lactic acid cells refers to yeast cells that stain Yeast bacteria The production of Scotch whisky employs around into glucose, maltose and other fermentable sugars. Two While most of these bacteria from malted barley will with methylene blue and are thus 41,000 Scottish residents and 65,000 people throughout types of yeast are generally used: a pure-culture distiller’s be killed by the mashing process, some will survive to dying or dead. the UK, generating about £1.3 billion of income for UK yeast obtained from a commercial yeast supplier and enter the fermentation (Fig. 1a). Many will also colonize Lactic acid DATA COURTESY HISATO IKEMEOTO & (wash activity) TOSHIHIKO TAKATANI, SUNTORY LTD households. Sales of Scotch whisky topped £2.3 billion spent brewer’s yeast. The practice of adding brewer’s the pipework, heat exchangers and other parts of the in 2002 and reached more than 200 countries, making yeast is increasingly rare, but it is thought by some to distillery plant. If too many occur at the early stage BELOW: it the largest matured spirit market in the world. impart important positive flavour characteristics to the of the fermentation (generally more than 106 per ml of Fig. 3. Effects of Lactobacillus Apart from the quality of the product, two related spirit. Interestingly, Scotch whisky distillers are not so wash) they will grow strongly in the fermentation, in the fermentation on ester concentrations in new-make developments led to this phenomenal dominance of possessive of their yeasts as brewers and do not develop inhibit the yeast and reduce the alcohol yield. This spirit as detected by GC-MS. Scotch whisky. By the end of the 19th century the and maintain their own yeast strains with particular early lactic fermentation is to be avoided since it reduces continuous still, patented by Aenas Coffey in 1827, fermentation and flavour properties. Instead, they rely the distillery efficiency; these are unwelcome partici- enabled the prodigious production of grain whisky on a on the distillation process to govern the flavour profile pants in the fermentation. Attention to plant cleanliness Alcohol fermentation Late lactic fermentation scale that could never be emulated in traditional pot of the finished product. The fermentation is conducted is generally sufficient to avoid the early lactic fermen- (Fermentation time – 3 days) stills. Second, this grain whisky provided the lighter base in wooden or stainless steel washbacks and is not tation. In a well-maintained distillery, the numbers for blending with malt whisky to provide the consistency attemperated. Consequently, the temperature can rise of bacteria entering the fermentation are relatively few 3.5×107 and quality of blended whisky. Blended whisky, which to over 30 °C and the fermentation is complete within 2 and bacterial growth is hardly evident during the initial today often involves 30–40 individual whiskies, is the days, reaching a little under 10 % alcohol by volume ethanol fermentation stage (Fig. 2). Then, as the yeast 3.0×107 major component of the modern Scotch whisky market. (abv). The wash, as it is known, is first distilled in the dies, after about 36 hours, the bacteria proliferate, Malt whisky is prepared from a mash of malted barley wash still to 21 % abv. It is then distilled a second time growing at the expense of yeast autolysis products, 2.5×107 which is fermented, distilled in traditional copper pot in the spirit still to over 70 % abv. Finally, it is reduced to malto-oligosaccharides and pentose sugars (Fig. 1b). stills and matured in oak casks for not less than 3 years 60 % abv for maturation in used (generally ex sherry or Finally, after about 80 hours even the bacteria begin to 2.0×107 and generally a lot longer. Malt whisky is increasing in bourbon) oak casks. die (Fig. 1c). The wash is normally distilled after about market share with a buoyant demand for ‘boutique’ 60–80 hours, and everything is discharged to the still, 1.5×107 products. However, despite increased export volume Lactic acid bacteria wash, yeast and bacteria. 7 Peak area . × last year (by 9.3 % to £268 million), malt whisky still Malted barley carries a varied microbial load with a For many years it was thought that the lactic acid 1 0 10 represents only a little over 10 % of the export market. predominance of lactic acid bacteria. These Gram- bacteria grew homogeneously throughout the 0.5×107 About 85 malt distilleries operate today in Scotland, but positive bacteria are strictly fermentative organisms fermentation. However, recent studies using denaturing ABOVE: their production is dwarfed by the dozen or so grain that cannot respire using an exogenous electron acceptor. gradient gel electrophoresis (DGGE) have revealed 07 distilleries. Grain spirit is prepared from unmalted They produce either lactate (homofermentative) or a changes in the population as the fermentation proceeds. Ethyl Isoamyl The Suntory Distillery, Osaka, Ethyl Butyl Isoamyl Japan. cereals as a source of starch, saccharified with malt mixture of lactate, acetate and carbon dioxide (hetero- Initially, a mixed flora of various cocci and rods enters hexanoate hexanoate octanoate × octanoate octanoate COURTESY SUNTORY DISTILLERY enzymes, fermented and distilled in a continuous or fermentative) from glucose catabolism. Consequently, the fermentation. This is followed by a reduction in the ( 10) (×10) 16 MICROBIOLOGYTODAY VOL 31/FEB04 MICROBIOLOGY TODAY VOL 31/FEB04 17 Table 1. Effects of some Lactobacillus strains on the organoleptic qualities of new-make spirit evaluated by sensory analysis Species Strain Sensory evaluation Estery Fatty Fruity Leafy Other L. acidophilus 1 + Green L. brevis 1 + Strong character L. casei 1 + Cereal 2 + Cereal 3 Diacetyl-like L. fermentum 1+++ 2+ L. pentosus / 1 + + Husky L. plantarum 2+ + PHOTO IAN ATHERTON, SGM Further reading by preparing laboratory-scale fermentations with Acknowledgements Russell, I. (editor) (2003). different bacterial strains present and analysing the new- Much of the work from my laboratory was supported by Suntory Whisky: Technology, Production make spirit by gas chromatography-mass spectrometry Ltd of Japan. I am particularly grateful to Hisato Ikemoto and and Marketing. London, San (GC-MS). The presence of bacteria increased the Toshihiko Takatani of Suntory for many valuable discussions Diego: Academic Press. concentrations of damascenone in the new-make spirit and for the material reproduced in Table 1 and Fig.
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