ENCYCLOPAEDIA OF BREWING ENCYCLOPAEDIA OF BREWING CHRIS BOULTON A John Wiley & Sons, Ltd., Publication This edition first published 2013 © 2013 Chris Boulton Registered office: John Wiley & Sons, Ltd, The Atrium, Southern Gate, Chichester, West Sussex, PO19 8SQ, UK Editorial offices: 9600 Garsington Road, Oxford, OX4 2DQ, UK The Atrium, Southern Gate, Chichester, West Sussex, PO19 8SQ, UK 111 River Street, Hoboken, NJ 07030-5774, USA For details of our global editorial offices, for customer services and for information about how to apply for permission to reuse the copyright material in this book please see our website at www.wiley.com/wiley-blackwell The right of the author to be identified as the author of this work has been asserted in accordance with the UK Copyright, Designs and Patents Act 1988 All rights reserved No part of this publication may be reproduced, stored in a retrieval system, or transmitted, in any form or by any means, electronic, mechanical, photocopying, recording or otherwise, except as permitted by the UK Copyright, Designs and Patents Act 1988, without the prior permission of the publisher Designations used by companies to distinguish their products are often claimed as trademarks All brand names and product names used in this book are trade names, service marks, trademarks or registered trademarks of their respective owners The publisher is not associated with any product or vendor mentioned in this book Limit of Liability/Disclaimer of Warranty: While the publisher and author(s) have used their best efforts in preparing this book, they make no representations or warranties with respect to the accuracy or completeness of the contents of this book and specifically disclaim any implied warranties of merchantability or fitness for a particular purpose It is sold on the understanding that the publisher is not engaged in rendering professional services and neither the publisher nor the author shall be liable for damages arising herefrom If professional advice or other expert assistance is required, the services of a competent professional should be sought Library of Congress Cataloging-in-Publication Data Boulton, Chris (Christopher M.) Encyclopaedia of brewing / Chris Boulton pages cm Includes bibliographical references and index ISBN 978-1-4051-6744-4 (cloth) Brewing–Encyclopedias I Title TP568.B68 2013 663'.3–dc23 2012050032 A catalogue record for this book is available from the British Library Wiley also publishes its books in a variety of electronic formats Some content that appears in print may not be available in electronic books Cover image: © iStockphoto/kedsanee Cover design by Meaden Creative Set in 10/13 pt Minion by Toppan Best-set Premedia Limited 2013 ACKNOWLEDGEMENTS I must first thank my publisher, Wiley, especially Andrew Harrison and Catriona Cooper for their patience and professionalism I owe much to colleagues past and present The brewing industry is unique in that sharing of knowledge and experience is seen as a virtue and not divulging secrets Long may this attitude continue Writing a book is most suited to solitary hermits and not those with responsibilities to family and friends This is particularly the case where work has to be fitted in the spaces that the day job doesn’t fill I am indebted to my wife, Wendy, to whom this book is dedicated, for her forbearance, not to mention many hours of sub-editorship in putting it together INTRODUCTION The Shorter Oxford Dictionary defines encylopaedia as ‘a work containing information on all branches of knowledge usually arranged alphabetically or a work containing exhaustive information on some one art or branch of knowledge arranged systematically’ An author who seeks to deliver a product that tries to fulfil these definitions knows that it will be a Sisyphean task This is especially the case with a subject such as brewing, with its long and rich history, its diversity of processes and products, not to mention the usually strong opinions of its practitioners In this respect I am well aware that this book will contain errors and omissions and probably an overemphasis on my own particular enthusiasms For all of these shortcomings I apologise and take full responsibility With regard to content, I have tried with each alphabetic entry to give a short initial definition which should provide the reader with all the essential information necessary for understanding such that further time need not be wasted The remainder of the entry is aimed at those who might wish to have further knowledge Hopefully, the system of cross referencing will provide greater context If there is a related entry the linking word is in bold Brewing and mainstream science have been inextricably intertwined for much of its history as an organised undertaking Indeed in its first industrial heyday many fundamental discoveries were made by brewers For this we should be justifiably proud, although it makes for some difficult decisions when deciding what should be included in a book such as this and what should be omitted This is all the more so when current scientific advances underpin many of the new processes and plants being introduced into brewing I have tried to steer a course which I am sure many will disagree with but one in which I hope that additional descriptions will serve to help with better understanding Finally, I have tried to encompass all parts of our industry, large and small, traditional and modern For this I not offer any apology I see no distinctions A Abbey beers Abbey beers are those produced commercially, largely in Belgium, and by statute solely within monasteries either directly or under the supervision of monks The popularity of Trappist beers in the period following the Second World War provided the impetus for arrangements under which commercial breweries produced beers that used the names of existing, or in some cases fictitious, abbeys as a marketing tool Commonly the use of a real abbey name involved a licensing agreement These products are collectively termed Belgian abbey beers Typically the beers ape the stronger dubbel and tripel true Trappist beers and in consequence are strong in alcohol, very flavoursome and made by top fermentation prior to bottling and a period of lengthy secondary conditioning See Trappist beers ABD medium Microbiological growth medium (advanced beer-spoiler detection medium) designed by Asahi Brewers of Japan, for the cultivation of difficult-to-grow lactic acid bacteria The medium comprises MRS broth supplemented with beer (to inhibit non-beer spoilers) cycloheximide (to prevent the growth of yeast) and sodium acetate (shown to be stimulatory to many lactic acid bacteria) Aber yeast biomass monitor Apparatus used for the automatic determination of viable yeast concentration (http://www aber-instruments.co.uk; last accessed February 2013) The device depends on the dielectrical properties of microbial cells when suspended in fluids that are conducting because of the presence of charged species When the cells, in this case yeast, are subjected to electrical fields, the charged species in the suspending medium (wort or beer) and those which are intracellular migrate towards the electrode bearing the opposite charge Since the cell membrane is nonconductive the cells function as capacitors and the magnitude of this can be measured The total yeast cell membrane area, or biovolume, within the operating field of the electrode can be related to yeast biomass Providing the sample is well-mixed the derived value of capacitance measured by the instrument can be expressed in the usual units of yeast concentration Encyclopaedia of Brewing, First Edition Chris Boulton © 2013 Chris Boulton Published 2013 by John Wiley & Sons, Ltd A A ABRASION such as viable cells per millilitre or viable yeast mass per unit mass or volume Dead cells, which have a disrupted cell membrane, not function as capacitors and are therefore not detected In this respect the measured capacitance correlates strongly with the fraction of a yeast sample scored as viable by a conventional vital staining approach such as methylene blue Similarly gas bubbles and non-yeast solids not generate capacitance and are not detected A corollary is since dead cells are not detected it does not provide any indication of viability Calibration involves setting zero and then determining the relationship between derived capacitance and viable biomass concentration Strain-dependent differences in electrical properties require calibrations to be made for each individual strain Once these are entered into the memory of the machine they not need to be repeated The linear range of the instrument is approximately 1 × 105 to 1 × 109 cells per millilitre Since the calibration requires comparison of results with yeast concentrations measured using conventional yeast analyses such as methylene blue staining and microscopic cell counting, the absolute precision cannot be better than these relatively crude methods However, the machine provides excellent repeatability Versions of the instrument are sold that are suitable for both laboratory and in-line analyses The instrument comprises a probe bearing four electrodes, two of which generate the electrical signal and two of which measure the magnitude of the resultant capacitance due to viable cells All living cells respond in this way and the magnitude of the measured capacitance is frequency-dependent In the case of yeast cells a value of 0.3 MHz has been found to provide an appropriate response The probe is inert and resistant to all brewery cleaning regimes Via a system of electronics the signal can be used to generate a signal which can be integrated with output from a flow meter or load cell such that automatic systems for control of pitching and cropping can be used In complex in-line systems several probes can be multiplexed via a single controller allowing outputs to be taken from combinations of multiple pitching and cropping mains Integration of all outputs allows the concentrations of all yeast within the brewery at any given time to be monitored Apart from control of yeast pitching and cropping the device can be used to control other processes such as krausening, cask beer re-seeding, yeast propagation and continuous centrifuge operation The laboratory version makes use of exactly the same technology, but the electrode is placed within an attemperated stirred chamber Abrasion A treatment applied to barley grains in which the husk is damaged (but not totally disrupted) by the application of mild mechanical treatments; for example, the use of rotating wire brushes The treatment enhances rates of germination either by allowing the more rapid entry into the grain of additives such as gibberellic acid but more likely via the increased efficiency of wetting and oxygenation Abraded grains can be malted at relatively low moisture contents and thereby allow shorter steeping times and lower steeping temperatures See gibberellic acid Abscisic acid Abscisic acid is a plant hormone with the structure indicated in the following figure ACCELERATED BATCH FERMENTATION CH3 CH3 CH3 C CH-C-CH-C-C-OH H2 OH O H O CH3 It exerts global effects on plants; for example, it is implicated in stress tolerance, stomatal opening, response to pathogens, seed development, apoptosis and the maintenance of dormancy Its involvement in the latter process is of the most direct relevance to brewing via the control of dormancy in grains that require to be germinated during malting The mechanisms by which it exerts its effects are at present not fully characterised, although it appears to have short-term effects as an effector of various cellular processes In addition, it seems capable of exerting longer-term effects via the modulation of gene activity Gibberellic acid has an antagonistic effect to abscisic acid See dormancy and gibberellic acid ABV ABV is an acronym that stands for alcohol by volume It is the usual method of denoting the alcohol concentration of beers The value is provided on packaging as x% abv Most beers fall within the ranges of 3–10% abv with the vast majority being between and 6% There are outliers The Samuel Adams Brewery in the United States produces the beer Utopias, which boasts an alcohol concentration of 25% abv In most countries there are legal definitions, expressed in terms of ABV, for low- and zero-alcohol beers Most countries use the ABV of beers as the mechanism for collecting excise duty In this regard, it is usual to have bandings such that all beers falling within a certain range of concentrations will attract the same rate of excise duty This reflects the fact that for many brewers precise control of alcohol content is difficult, and therefore a degree of latitude is given Naturally, given this situation most brewers will seek to ensure that the actual mean alcohol concentration of any given beer is as close as possible to the middle point of the band This avoids paying excessive taxation but also ensures that on average the product satisfies the legal requirements Since most excise payments are based on self-assessment and, bearing in mind the pivotal role of ABV, the analytical methods used must have suitable precision and repeatability This has resulted in the adoption of so-called reference methods of analysis which have legal status Many other methods may be used for routine analyses, based on factors such as rapidity or ease of automation; however, at some stage analyses must be performed using a standard reference method Accelerated batch fermentation Accelerated batch fermentation is an umbrella term that covers a wide variety of approaches which have been developed with the aim of increasing the productivity of batch fermentations by shortening cycle times For any commercial brewer the capital costs of fermenters and associated plant represent a major investment This is particularly so in the case of the very-large-capacity vessels used by many of the major world brewers In addition to capital A YEAST STRAIN IMPROVEMENT 695 They are constructed from stainless steel, of cylindrical design with dished tops and bottoms and are fitted with wall-mounted cooling jackets with sufficient capacity to hold the temperature at approximately 3°C (±1°C) The vessels are designed to maintain the holding temperature and not to cool warm yeast Where yeast is cropped at relatively warm temperatures it is necessary to provide an external in-line chiller In order to promote efficient attemperation a system of gentle agitation is also provided This should be designed such that the agitation is sufficient to prevent yeast settling out but not too vigorous to generate excessive shear forces Paddle-type impellers fitted to a top-mounted electrical drive system meet this need A sample point mounted close to the base of the vessel allows removal of yeast for analysis Since good hygiene is essential for the correct operation of these vessels the sample tap should preferably be of the type which can be sterilised by steam prior to use In order to prevent ingress of microbes the vessels are top pressured with an inert gas such as nitrogen or CO2 It is important to exclude air in order to prevent changes of yeast physiology such as dissimilation of glycogen reserves coupled to sterol synthesis, which would have the potential to produce inconsistent performance when the yeast is pitched into a subsequent fermentation The gas exit point and top-pressurisation system must be fitted with appropriate sterile gas filters The majority of breweries have multiple YSVs, the individual sizes of which are chosen to meet the needs and capacity of the brewery It is usual to size the vessels such that they can accommodate the total crop, which is required to be retained from a single fermentation This arrangement allows individual yeast lines of varying generational ages to be segregated from each other Yeast strain Term used to denote a specific variety of yeast The term strain is used to denote the smallest taxonomic unit in which the possession of a distinct genotype can be demonstrated and is a subdivision of the species (see yeast for more details) In brewing terms many thousands of individual yeast strains are used and commonly they are the proprietary yeast types used, and jealously guarded, by individual brewers Although all have genotypes which are sufficiently similar to allow them to be classified into the larger species groups (either S cerevisiae, which include ale strains, or Saccharomyces pastorianus, lager strains), very small differences in genetic make-up can be detected, and when these are expressed, a phenotype is produced which provides a unique pattern of detectable and reproducible brewing properties Nomenclature at the strain level is unregulated and commonly either a name may be used which relates to the proprietary company, the beer quality for which they are used to produce, a description of a major brewing characteristic or simply a combination of letters and numbers Yeast strain improvement Techniques which allow the isolation or generation of yeast strains with desirable brewing properties The advent of the use of pure brewing yeast strains, pioneered by Hansen, provided a framework in which the properties of individual strains could be compared and contrasted, and this led to an appreciation that there was considerable diversity It has always been possible to simply choose a brewing strain which possesses desirable properties; however, directed strain improvement provides a more focussed route to this end Properties which may be looked for relate to fermentation performance and beer quality Examples include more efficient and/or more rapid conversion of extract into ethanol, the Y 696 Y YEAST STRAIN IMPROVEMENT ability to produce and survive in very high-gravity worts and to produce and tolerate the resultant high concentrations of ethanol, the ability to utilise a particular spectrum of carbohydrates, possession of flocculation characteristics which suit the type of fermentation and fermenter being used, the ability to outcompete potential contaminants and production of a particular spectrum of yeast-derived flavour compounds New strains can be identified by looking for natural variants within a population which have the desired property The search can be made less hit and miss by the use of mutagenic agents and screening for desired variants The development of genetic engineering extends the possibilities by providing a vehicle in which exogenous genes, from any source, can be introduced into brewing strains and thereby introducing characters that would not be normally be available Particular fermentation systems tend to be selective for certain yeast properties For example, early crops formed in deep cylindroconical fermenters tend to be the most flocculent Selection of this fraction of the crop can be used as a method of natural selection if a more flocculent strain is desired (and vice versa) Similar approaches can be used to select for other natural variants Standard mutagens such as UV radiation have been used successfully to increase the frequency of natural variation and thereby to generate increased numbers of variants Classical mating techniques are problematic with brewing yeasts because of their polyploid/ aneuploid genome, and the generation of hybrid diploids from the fusion of haploid spores is very difficult to achieve The technique of rare mating has been used with some success This method uses strong selective pressures to isolate rare hybrids Typically an excess of usually respiratory-deficient parental strains is mixed with respiratory-sufficient haploid auxotrophs Rare hybrids are selected as respiratory-sufficient prototrophs The technique of sphaeroplast fusion in which osmotically stabilised cells stripped of their cell walls are made to fuse by the application of a strong electric field has been used to form hybrids in which selected characters can be transmitted from donor to recipient cells Recombinant DNA genetic manipulations rely on the introduction of selected genes into recipient cells to generate individuals with altered genomes Typically the foreign DNA is attached to a plasmid which also bears a dominant marker which allows selection of target cells This approach is necessary because of the polyploid nature of the host brewing strains Commonly used dominant markers include resistance to copper, herbicides such as sulphometuron-methyl and various antibiotics such as chloramphenicol and gentomycin Multi-copies of the plasmids bearing genes of interest may be inserted into target cells and left in this form or they may be integrated into yeast chromosomes to provide more stable transformants Genetically manipulated brewing strains have been developed which are able to utilise dextrins (super-attenuating), possess killer factors, produce less than normal concentrations of diacetyl, possess β-glucanase activity, have constitutive maltose assimilating enzymes, produce reduced levels of H2S and increased production of SO2 The current public aversion to the use of a genetically engineered organism for food production has resulted in a situation in which many major brewers have such strains but are not currently exploiting them commercially YEAST SUPPLY 697 Yeast stress response A term applied to yeast strains which describe the response of the genome and phenotype to applied stresses The commercial brewing process is inherently stressful to yeast Examples include exposure to rapid changes in temperature (both upwards and downwards), low pH, high barometric pressure, low water activity, high osmotic pressure, exposure to high levels of ethanol shifts from anaerobiosis to aerobiosis (and the reverse) and periods of starvation Modern brewing practices such as the use of very large batch sizes, very high-gravity worts and elevated fermentation temperatures exacerbate many of these stresses Failure to manage these stresses can result in losses of yeast viability with concomitant undesirable effects on fermentation performance and beer quality Brewing yeast, in common with many other cells, are able to adapt their phenotypes in response to sudden changes in external conditions Part of these responses is the ability to adapt the phenotype so that the cells are better able to survive stressful conditions The ability to withstand stress is to some extent strain specific and in absolute terms there is considerable variation However, all strains exhibit a common stress response Application of a sub-lethal stress triggers changes which result in the cellular adaptation such that the cells exhibit greater resistance to that and other stresses The latter observation provides evidence that several different external triggers can elicit a common response When subject to an abrupt change in external conditions the cells typically move into a phase of apparent inactivity where growth ceases During this time the adaptive changes occur, which constitute the stress response The most studied response is that which occurs when cells are subject to a non-lethal heat shock During a period of growth arrest some 70 heat shock proteins (hsps) are synthesised Growth recommences and the hsps persist, and during this time the cells exhibit higher thermo-tolerance compared with the non-shocked cells The functions of the hsps are mainly unknown; however, some of them are enzymes that repair partially denatured proteins, the ubquitin system is activated, which degrades totally denatured proteins, and trehalose synthesis (a potent membrane stabiliser) is up-regulated A specific stress response element (STRE) has been identified in many genes which are involved in various stress-protecting duties Some of these are specific for individual stresses; others are part of a common stress response It appears that yeasts possess sensing systems which identify changes in external conditions Where these changes are stressful specific genetic responses are elicited, which via signal transduction pathways feed into a common stress response mechanism These responses are transmitted through the cell using mitogenactivated protein kinase (MAPK) cascades Yeast supply The processes used to ensure that breweries have a timely supply of pitching yeast of the correct strain, of guaranteed purity, in an appropriate physiological condition and in the correct quantity Several levels of sophistication are possible depending on the nature of the brewery: (1) perpetual serial fermentation, cropping, brewery storage and re-pitching (2) periodic supply of bulk pitching yeast slurry or cake from another brewer (3) third party supply of active dried yeast used in one-trip fermentation Y 698 YEAST SUPPLY (4) third party supply of bespoke bulk culture of pitching yeast slurry (5) third party supply of pure laboratory cultures for in-house propagation (6) in-house storage of one of more proprietary yeast strains supplied to parent or group breweries for propagation Ultimately, apart from the perpetual serial fermentation approach, all supply systems share common features These are (1) preservation of master cultures (2) recovery from storage (3) confirmation of strain purity and identity (4) preparation of working culture (5) laboratory propagation (6) brewery propagation Culture preservation and recovery The master culture is one of guaranteed purity and identity and which when used in brewing has desired brewing properties Master yeast cultures must be stored in a way that maintains viability and ensures freedom from contamination The latter is achieved using appropriate containment and conventional aseptic technique Viability is maintained by reducing metabolic activity This can be achieved by storage at low temperature and/or by removal of cellular water Several methods are used of varying efficacy Cultures may be stored, usually on nutrient media solidified with agar, at 2–4°C or better at −80°C with a layer of oil covering the growing yeast to exclude air Cultures may be freeze-dried (lyophilised) in which intracellular water is removed by sublimation by freezing under vacuum and the resultant powder is sealed in a glass vial Freeze-dried cultures can be stored at cool temperatures for several months and after recovery yield viable cells Although this method of preservation was very widely used, it is now known that overall viabilities may be very low, and in the case of brewing yeast strains the drying process may cause the generation of genetic variants The gold standard method of preservation relies on storage under liquid nitrogen Providing correct procedures are followed regarding the preparation of the culture and the method of freezing, cryopreservation in liquid nitrogen effectively provides an indefinite method of storage Cultures stored at refrigerator temperatures on solidified nutrient media not require any special procedure for recovery; however, for the purposes of preservation a portion of the culture should be used to seed fresh medium approximately every months Freeze-dried or liquid nitrogen cultures are transferred aseptically into a sterile liquid nutrient medium and are incubated to produce an intermediary culture Y Confirmation of purity and identity The purity of the intermediary culture is assessed using conventional microbiological techniques A variety of differential media are used to check for the presence of potential bacterial and wild yeast contaminants Typically streaking out onto a general yeast nutrient medium such as WLN medium, solidified with agar is used to check that the resultant colonies are uniform in appearance and have a colour and appearance characteristic for the particular strain Definitive proof of identity is guaranteed by genetic fingerprinting techniques, the results of which are compared with historical records of that of the master culture YEAST TAXONOMY 699 Working cultures The intermediate culture is used to inoculate working cultures which typically take the form of slopes (or slants) These are small glass tubes or bottles which contain a yeast nutrient medium solidified with agar Whilst the medium is still molten the tubes are inclined such that when solid the surface area for yeast growth is maximised After incubation for a few days at room temperature the slope cultures are stored at 2–4°C Propagation The working culture is used to supply the initial inoculum for a series of serial cultures of ever-increasing volume, first within the laboratory and then in the brewery, with the aim of generating sufficient yeast to pitch the first production-scale fermentation (see yeast propagation for more details) Documentation and management It is essential that the yeast cultures supplied to the brewery have the correct purity and identity Failure, particularly where multiple strains are used in one brewery, will have potentially severe consequences in terms of subsequent beer quality Once a culture has been supplied to the production environment there is little or no chance to remedy any error before significant financial losses become inevitable The design of the supply system must be sufficiently robust to avoid errors being made In order to minimise the risk of human error it is essential that yeast supply is underpinned by a properly documented and traceable system which uses colour or number codes to assist checking culture identity At critical steps the actions of the operator should be checked by an assistant At regular intervals the whole of the process should be subject to scrutiny by a suitably trained audit team and any weaknesses identified and eliminated Yeast taxonomy Yeast constitutes a diverse group of microorganisms which are classified within the fungi Representatives are found in three of the major fungal divisions, Ascomycota (to which brewing strains belong), Basidiomycota and Deuteromycota The discussion here is confined to brewing yeast The taxonomy of brewing yeast can be confusing Parallel systems of nomenclature have arisen which reflect, on the one hand, the behaviour of individual strains in the context of brewing and, on the other, based on more rigorous genetic analyses With respect to brewing properties yeasts are traditionally classified as being ale or lager types On the basis of their behaviour in the fermenter these are also referred to as either top cropping (ale types) or bottom cropping (lager types) Historically these yeasts have been classified as S cerevisiae (ale strains) and Saccharomyces uvarum or Saccharomyces carlsbergensis (lager strains) The term ‘carlsbergensis’, later changed to ‘uvarum’, reflects the fact that pure lager strains were isolated relatively recently at the Carlsberg Research Institute in Copenhagen and were shown to be distinct from the, in evolutionary terms, much older ale strains These names continue to be used; however, the taxonomy of yeasts has been and is still subject to a process of continual revision as a result of the greater precision of recently developed genetic analytical techniques These revisions have been accompanied by changes in nomenclature, Y 700 YEAST VIABILITY and unfortunately the older systems have remained in common use with the result that parallel naming systems, alluded to already, have arisen Early systems of yeast taxonomy relied on traditional descriptors using morphological and biochemical markers These are useful since they describe features which are of practical relevance in commercial applications such as brewing However, from a purely scientific standpoint, they have been superseded by more precise genetic analyses which provide a more reliable basis for differentiation and indicate evolutionary relationships All Saccharomyces yeasts used in biotechnological applications are now classified within a subgroup termed Saccharomyces sensu stricto As the name suggests all the species in this group are closely related Strains from all member species can be crossed in any combination to produce hybrid ascospores However, only those made between members of the same species are viable Currently, six species are recognised within the sensu stricto group and these are S cerevisiae, Saccharomyces paradoxus, S bayanus, Saccharomyces cariocanus, Saccharomyces kudriavzevii and Saccharomyces mikatae, although further revisions are likely Of these, members of S cerevisiae are used for baking, brewing and winemaking Saccharomyces bayanus strains are associated with winemaking and S pastorianus includes those strains originally classified as S carlsbergensis and used as bottom-fermenting lager strains It has now been demonstrated that S pastorianus is a hybrid species derived from a fusion event involving parental strains of S cerevisiae and S bayanus Thus, the genome of the hybrids is larger than that of the parental types, typically 1.5–1.6× bigger, and is aneuploid Considerable genetic diversity occurs since chromosomes may be pure S cerevisiae or S bayanus or may be mosaics containing genes identical to both parental types Y Yeast viability Viability is defined as the percentage of live cells within a population Within the context of brewing yeast viability measurements are made to allow corrections to be made when determining pitching rates In addition, viability is used as a quantitative measure of the condition of pitching yeast Commonly breweries use a specified viability value (typically >90%) below which yeast would be deemed unsuitable for use in fermentation From a scientific standpoint the concept of viability is contentious It is usually defined as simply ‘the capacity for living’ In respect to yeast, it may be regarded as the ability to proliferate under appropriate conditions In the case of those stages in brewing which require the presence of yeast such as primary and secondary fermentation it is possible that some yeast cells are not capable of progressing through the cell cycle but may still make a (desirable or undesirable) contribution to fermentation The viability of microorganisms is typically determined using plate counts, or variants thereof Suspensions of microorganisms, with a known total cell concentration, are streaked onto a nutrient medium, solidified with agar, such that individual cells are separated spatially Following a period of incubation colonies form, and it is assumed that each of these must have arisen from a single viable cell With knowledge of the number of colonies and the original total cell count the viability can be calculated In order to differentiate between cells capable of proliferation and those which cannot but still exhibit metabolic activity the term ‘colonyforming unit (CFU)’ is often used Conversely, the term ‘viable but not culturable’, or simply, YEAST VITALITY 701 ‘non-culturable’ is used to describe cells which have detectable metabolic activity but which are not able to proliferate under the conditions where they are tested In order to use viability measurements to assess yeast quality and to determine pitching rates it is necessary to have a timely response Plate count methods require one to a few days in order to generate a result and therefore such approaches are impractical for routine use A result can be generated in several hours using a more rapid slide culture technique in which a layer of nutrient medium is layered onto a microscope slide and the development of microcolonies is observed Nevertheless, this is still too slow and other methods based on the use of so-called vital stains (dyes) are used routinely These are reagents which when mixed with suspensions of cells produce differential staining reactions such that sub-populations of viable and non-viable cells can be distinguished The dye methylene blue is most commonly used in the brewing industry to determine the viability of yeast This stain is taken up by dead cells such that they become stained blue Viable cells are able to reduce the dye to the colourless leuco form and remain unstained Although the method is the industry standard for yeast viability determination, justifiably it has some critics Compared with colony counts it gives an overestimation of viability at values less than approximately 90% The error increases with a decrease in the true viability This is a result of the presence of increased proportions of severely stressed cells which tend to stain pale blue which may or may not be counted depending on the preference of the operator For this reason alternative vital stains are used, which provide a more definite differentiation These include acridine orange, eosin Y, methylene violet 3-RAX, fluorescein diacetate, DiBAC4, propidium iodide, Chemchrome Y, MgANS and Rhodamine 123 The mechanisms which underpin these stains are various but generally involve exclusion by viable cells or uptake and modification by viable cells with an accompanying colour change Many of the dyes are fluorescent, a property which, it is claimed, provides better discrimination, albeit at a cost in terms of the generally higher costs of reagents and the need for a fluorescence microscope Many studies have been made in which correlations have been made between viability measured with vitality stains compared with other techniques such as plate counts In general the fluorescent techniques appear to offer the best discrimination and Mg-ANS appears to be very suitable for application with brewing yeast With regard to bright field stains, methlylene violet is probably easier to use compared with methylene blue; however, it seems likely that many brewers will continue to favour the latter based on conservatism Measurement of viability can be automated using electronic cell counters, such as the flow cytometer (see flow cytometry), although at a cost at least an order of magnitude greater See also yeast vitality Yeast vitality The term used to describe the basis of numerous tests that have been developed with the aim of probing yeast physiological condition and how this relates to fermentation performance Traditionally the condition of yeast is assessed based on the determination of the proportion of living cells in the population (see yeast viability) using the reasoning that if the viability is low, it is likely that the viable fraction is not fit for purpose Vitality tests extend this concept by assessing the physiological status of the viable fraction of yeast populations Y 702 YEAST WASHING Ideal vitality tests are rapid, inexpensive, simple to perform and provide a result which can be used either as the basis of a decision on the fitness to pitch of a particular batch of yeast but preferably one which is predictive of subsequent fermentation performance such that an optimum pitching rate and/or wort oxygenation regime can be deduced It might be supposed that the rigorous application of quality assurance principles should preclude the need for such testing; however, the development of intensive fermentation techniques such as high- and ultra-high-gravity brewing, together with very large batch sizes, which undoubtedly increase the stresses to which yeast is exposed, has fuelled the perception that testing of pitching yeast condition should use techniques which provide more information than simple viability tests A plethora of tests has been suggested These include simple microscope-based staining techniques (assessment of membrane integrity or visible changes brought about by intracellular processing of dyes), chemical analysis of cellular composition (ATP, NADH, sterols, glycogen, trehalose), ability to proliferate, assessment of the electrokinetic properties of cells and measurements of biochemical activities under defined conditions The latter includes rapid and small-scale assessment of characters related to fermentation performance such as uptake of sugars and oxygen or the formation of heat, CO2 or ethanol Other physiological assessments rely on detection of intracellular pH, or the ability to acidify the medium or the degree to which cells can withstand an applied stress In the case of the biochemical tests, a few specific pieces of bespoke equipment have been developed to make conducting the test less reliant on the skills of the operator and therefore possibly more suitable for use in a production environment Should costs fall to levels within the reach of a typical brewery laboratory it is likely that flow cytometry will offer a powerful means of assessing the condition of yeast populations There is little consensus as to which tests should be adopted Many brewers are content to use viability testing based on the pragmatic assumption that if this is low it may be assumed that the viable fraction of the population is compromised The value of vitality tests is that they provide added value This could be an automatic method for viability detection, or a related parameter, which is rapid and avoids the errors associated with manual microscopic counting techniques Other tests may give additional information This can be particularly useful where it is not just necessary to identify yeast with compromised physiology but to bring focus to the areas of yeast handling where problems are being introduced In this regard many brewers will choose a single vitality test (the acidification power test and its variants seems to be particularly popular) for routine use Providing this is applied in a consistent manner it is undoubtedly useful A broader range of tests may be used for troubleshooting exercises Y Yeast washing A generic term for the variety of treatments which are used to disinfect pitching yeast slurries The most common is acid washing in which bacterial contaminants are killed based on their relative intolerance of low pH compared with brewing yeast Several other selective biocides have been used Antibiotics were used until this practice was precluded by the realisation of the risks associated with the selection of resistant strains Most recently it has been suggested that chlorine dioxide could also find utility in this role YSV 703 YM medium Yeast and mould medium (see MYGP medium) Yorkshire square Yorkshire squares are fermentation vessels traditionally made from slate or stone, which are variants of the more usual open squares Traditional types are of modest capacity, usually >50 hL; more modern types are bigger, up to 900 hL, and are made from stainless steel They were developed specifically to be used with flocculent ale strains The vessels comprise rectangular enclosures the top of which takes the form of an inclined deck located towards the top of the vertical sides but leaving a lip around the periphery of the vessel The deck is pierced by a central manhole which has a rim on its upper surface about 15 cm in height The upper and lower compartments, separated by the deck, are also connected by a number of circular pipes, termed organ pipes Control of temperature is via external cooling jackets or submerged attemperators The deck is also provided with an aperture for dipping to measure wort volume and an inlet to a drain for yeast removal At the start of fermentation pitched and aerated wort is pumped into the vessel to a height close to that of the deck During primary fermentation the yeast rises through the central manhole where it is retained by the deck Beer separates from the yeast and re-enters the lower compartment of the vessel via the organ pipes Where very flocculent yeast strains are used a pumped recirculation system is provided, which takes wort from a drain point at the base of the vessel and returns it to the top of the deck When the primary fermentation is judged complete the recirculation loop is switched off and the yeast crop is allowed to settle on the surface of the deck from where it is collected by pushing into the exit drain provided for that purpose The green beer is then removed from the bottom drain Proponents of these vessels claim that their disadvantages, namely, complexity and high beer losses, are outweighed by the quality of the beer produced This has resulted in the installation of modern versions which have provision for automatic CIP and CO2 collection In addition, the removal of yeast crops and tank bottoms has been automated by the incorporation of an arrangement of sequentially operated water jets which force the solid products towards exit points attached to suction pumps YSV See yeast storage vessel Y Z Žatec Žatec is a town in what was Bohemia and now the Czech Republic It has a long history, probably more than 700 years, of cultivating hops It is the site of origin of the famous noble aroma hop cultivar, Saaz, which gives the Czech pilsner-type beers their characteristic delicate low bitterness, floral hoppy aromas and taste Zatecky Chmel Zatecky Chmel, literally hops from Žatec, is the preferred name for Saaz hops grown in the Czech Republic In some ways it reflects the wish by Czech nationals to dispense with older German nomenclature (as in ‘Saaz’) and to replace it with the native ‘Žatec’ The term is protected by laws of the European Union in which Zatecky Chmel is designated as a Protected Designation of Origin such that by statute (EU regulation 2081/92), from May 8, 2007, hops may only bear the mark Zatecky Chmel if they are aroma types of the Saaz variety or designated clones The crops of 2007–2008 from this region were the first to be so protected Named clones of Žatec (Saaz) aroma hops grown in this region, together with the dates of their registration and covered by the legislation, are Lucan (1941), Blato (1952), Osvald’s Clones 31, 72, and 114 (all registered in 1952), Sirem (1969), Zlatan (1976), Podlesak (1989) and Blsanka (1993) These types derive from testing local landraces of the Saaz variety and as well as having desirable aroma properties were selected largely on the basis of yield See Saaz hop Zbyszko Zbyszko is a high alpha bittering hop variety of Polish origin Analysis is 8.5% total α-acids of which 25.6% is cohumulone Total oil content is 1.9% of which 9.7% is caryophyllene, 23.9% is humulene and 53.3% is myrcene Zein Zein is a prolamin protein obtained from the grains of maize (Zea mays) It is the equivalent to the hordeins in barley Pure zein is water insoluble and edible but also hard, colourless and Z Encyclopaedia of Brewing, First Edition Chris Boulton © 2013 Chris Boulton Published 2013 by John Wiley & Sons, Ltd ZETA POTENTIAL 705 odourless These properties have favoured industrial applications such as use as various coatings However, since maize grits and flours are also used as adjuncts in brewing, particularly in the case of sorghum beers, zeins must contribute to the nitrogenous contents of worts and the resultant beers Zeiss–Pulfrich nephelometer An early beer haze meter based on the measurement of light scattering at 45° It was used as a standard reference instrument in many breweries before the advent of modern apparatus Zenter A measure of weight used in Europe for the quantification of hop yields Numerically, Zenter is equal 50 kg Zero-alcohol beers See reduced-alcohol beer Zeta potential Zeta potential is a parameter which is used to describe the electrostatic properties of colloidal particles suspended in a medium which contains charged ionic groups It is of relevance to brewing in several ways The zeta potential of haze particles suspended in beer and the ionic composition of the latter influence the rates at which these particles sediment Process aids such as fining agents influence zeta potential and thereby accelerate the rates at which particles form sediments in tanks The colloidal particles suspended in beer or wort may include yeast cells The zeta potential of yeast cells influences their behaviour in such media in that it affects rates of sedimentation, as described, and also the ability of cells to come together and form flocs Yeast cells bear a net negative charge owing to the presence of phosphate and/or carboxyl groups on the cell surface These tend to make the cells mutually repulsive This repulsive force must be overcome in order for cells to come together and form flocs It has been observed that in the later stages of fermentation the zeta potential of yeast cells decreases and this in part explains why flocculation occurs at this time (see yeast flocculation for more details) It is suggested by some that the zeta potential of yeast cells should be monitored as a routine test of condition Zeta potential is of importance in depth filtration Many artificial filtration media, particularly those made from synthetic polymers, are made from fibres which carry a net positive zeta potential This allows the filtration medium to bind and retain oppositely charged particles even though they may be smaller than the average pore size Zeta potential differs from simple surface charge Charged particles suspended in an ionic medium become surrounded by a layer of articles of opposite charge This is termed the fixed layer Outside of this is a diffuse layer which forms a cloud-like region made up of ions of opposite polarity such that the whole is electrically neutral The potential difference between the particles and surrounding medium is a function of the nature of the surface charge of the particle and the ionic composition of the medium Zeta potential is measured by assessing the mobility of particles when they are placed within an electrical field In these circumstances the charged particles will migrate towards the Z 706 ZEUS electrode of opposite charge The particles migrate with the fixed layer and the inner part of the diffuse layer, termed the sliding surface At the point of electrostatic neutrality the particles will form aggregates The zeta potential is a measure of the potential difference of this sliding layer and that of the bulk suspending liquid Zeus Zeus is a US super alpha hop variety developed in the Yakima Valley It is very similar to Columbus (Tomahawk) It contains 12.0–16.5% α-acids, of which 27.0–35.0% is cohumulone, and 4.0–6.0% β-acids Total oils are 1.0–2.0% (5.0–15.0% caryophyllene,