2011S- Yeast - Yeast Yeast Yeast Yeast Single cell,...

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Unformatted text preview: Yeast Yeast Yeast Yeast Single cell, reproduce by budding Classified as fungi Saccharomyces cerevisiae / uvarum uvarum (carlsburgensis) One species ?? different?? Goldammer suggests they Goldammer were recently reclassified were Yeast Strains Yeast C. cerevisiae top fermenting for ales, porters, stouts, Altbier, Kolsch, wheat beer porters, C. uvarum for lagers, Pilsner, Dortmunder, Marzen, Bock, malt liquors Marzen, Some disagreement on taxonomy Ale Yeast Ale Temperature optimum 10-25 °C (50-77 °F) Rise to surface during fermentation, Rise creating thick, rich yeast head creating Tend to have distinct levels of esters Tend (fruity character) due to warm fermentation (fruity Flocculation (how well they clump and Flocculation settle) settle) Mixed cultures may be more stable Lager Yeast Lager Best at temperatures from 7 °C (45 °F) to Best 15 °C (59 °F) 15 Growth rate is slower, settle out as Growth fermentation nears completion fermentation Temp and strain greatly affect flavor Powdery or flocculant Powdery have greater attenuation, not Powdery always clear always Lager Yeast Lager Pure culture (Emil Hanson Carlsburg Pure Denmark) Denmark) Sometimes combine flocculant and Sometimes powdery strains (European mainly) powdery LIFE CYCLE LIFE Begin as dormant cells – LAG – GROWTH – FERMENTATION – SEDEMENTATION LAG PHASE LAG Activated cells start building up energy Activated reserves reserves Use phosphate, oxygen, stored glycogen – Low glycogen leads to higher levels of vicinal Low diketones > diacetyl Lag should be as short as possible 8-24h Lag Lower lag by starting yeast in similar Lower medium medium GROWTH PHASE GROWTH Also called respiration phase Oxygen used, CO2 produced Acids formed, pH drops to 4.0-4.5 (ale), Acids 4.4-4.7 (lager) 4.4-4.7 Logarithmic growth Budding – 2 or 3 doublings in inoculum will Budding occur in commercial production occur FERMENTATION PHASE FERMENTATION When oxygen is used up, enter When fermentation phase fermentation Fermentation is anaerobic process Sugars used up, CO2, ethanol, flavor ethanol, compounds produced compounds Gravity drops Yeast is in suspension (3-7 days, longer in Yeast lagers) lagers) SEDIMENTATION PHASE SEDIMENTATION Flocculate and settle to bottom Build up glycogen stores for dormancy Flocculation and attenuation Low Flocculation - 73%-77% Medium Flocculation - 71%-75% High Flocculation - 67%-71% NUTRITIONAL REQUIREMENTS NUTRITIONAL Yeast need nutrients to grow and ferment Nitrogen, carbohydrates, minerals, Nitrogen, vitamins vitamins Inadequate nutrients can lead to poor beer Inadequate stability, yeast mutation, off flavors, slow or stuck fermentations or Stuck fermentation – yeast don’t complete Stuck conversion of fermentable carbohydrates conversion Especially problematic with high gravities Carbohydrates Carbohydrates Mono, di, and sometimes trisaccharides Mono, available for yeast growth available Glucose – maltose – maltotriose – Glucose maltotetrose (1, 2, 3, 4 glucose units) maltotetrose Concentration of carbos: maltose – Concentration maltotriose – glucose – sucrose – fructose maltotriose 75-85% of malt extract consists of these 75-85% simple sugars simple Utilization: sucrose, glucose, fructose, maltose, maltotriose maltose, Maltotetrose and dextrins go unfermented Glucose inhibits the permease enzyme Glucose required for maltose to enter cell required As glucose decreases, the enzymes As needed for maltose metabolism are synthesized synthesized Carbohydrates Carbohydrates Too much glucose or fructose can repress Too maltose metabolism and lead to stuck fermentations fermentations NITROGEN NITROGEN Nitrogenous compounds available for Nitrogenous yeast include amino acids, peptides, and ammonium salts ammonium Yeast prefer ammonium salts, but are Yeast present at low levels in normal worts present Amino acids (free amino nitrogen) Amino Optimum FAN depends on gravity of wort Optimum 110-140 mg/l 10 °P .. 300-325 mg /l 12 °P NITROGEN NITROGEN Nitrogenous compounds in wort depend Nitrogenous on choice of grains, malting process, adjuncts and mashing adjuncts More than 40% adjunct (think bud light) More can lead to nitrogen deficiency in wort can Add enzymes to increase FAN or add Add ammonium salts directly ammonium More than 70% wheat in grainbill can lead to N deficiency problems lead N deficiency can result in stuck/slow deficiency fermentations and abnormal metabolism, resulting in fusel alcohol production production Fusel alcohols are larger alcohols Fusel than ethanol (propanols, butanols, pentanols) pentanols) Fuseln = to bungle (Ger.) Fuseln Undermodified malts have low FAN (we remember they have less enzyme activation during germination) activation 4 main fusel alcohols, most pleasant is main phenylethyl alcohol phenylethyl Propanol, isobutanol, isoamyl & Pentanol Propanol, are the major contributors VITAMINS VITAMINS – Biotin, pantothenic acid, thiamin, inositol – Deficiency causes metabolism problems Biotin: needed for carboxylation, nucleic Biotin: acid, fatty acid and protein synthesis, deficiency = high yeast death deficiency Pantothenic acid: carb & lipid metabolism, Pantothenic cell walls, deficiency = hydrogen sulfide accumulation (rotten egg smell) accumulation Thiamin: needed for decarboxylations Inositol: needed for cell division Inositol: Minerals Minerals Requirements for phosphate, potassium, Requirements calcium, magnesium, sulfur, & trace elements (copper, selenium, zinc, etc) elements Phosphate (P) needed for energy Phosphate conservation, cell growth conservation, Potassium (K) needed for P intake Calcium (Ca) affects flocculation and Calcium should be present at 50 mg/L or higher should Minerals Minerals Magnesium (Mg) needed for growth and Magnesium enzymes enzymes Sulfur (S) needed for sulfur amino acid Sulfur (methionine, cysteine - typo in book) synthesis synthesis Zinc (Zn) trace element needed for protein Zinc synthesis, etc synthesis, Zn deficience may be seen in use unmalted Zn cereals or switch from galvanized (Zn plated) pipes to SS - 0.3mg/l good, >1 maybe toxic pipes Metabolism Byproducts Metabolism Flavors in beer depend a lot on yeast Flavors metabolism byproducts metabolism Esters, alcohols, acids Nonflocculant strains often produce more Nonflocculant volatiles than flocculant volatiles llager years produce more sulfur ager compounds and fatty acids compounds Mutant strains often produce funky flavors ESTERS ESTERS most important aroma compounds in beer Impart fruity character, especially important in Impart ales ales Ester production enhanced by: – increasing fermentation temperature – restricting wort aeration – increasing attenuation limit and achieved apparent attenuation limit (AAL) of malt describes the apparent maximum percentage of fermentable sugars in a wort that can be metabolised by brewers yeast. can – increasing wort concentration to above 13°P ESTERS ESTERS FATTY ACID + ALCOHOL – ethyl acetate – isoamyl acetate – isobutyl acetate – phenyl acetate – ethyl caproate – ethyl caprylate ESTERS ESTERS ETHYL BUTYRATE – 0.4 mg/l threshold, 0.3 + in beer – apple flavors in Budweiser products ETHYL ACETATE – 25-35 mg/l threshold, levels in beers 5-30 mg/l – Fruity, solvent - like Fruity, – Some nail polish remover has EA ISOAMYL ACETATE – banana ester – 1-3 mg/l threshold – fruity, banana VICINAL DIKETONES (VDK) VICINAL Diacetyl and 2,3-pentanedione DIACETYL O H O H 2,3-PENTANEDIONE O H3C H H O CH3 VDK VDK levels decrease as beer is aged diacetyl more important of two buttery, butterscotch flavor diacetyl honey flavor pentanedione diacetyl flavor tends to be unstable defect in lager beers, desirable in British defect ales ales VDK VDK Decarboxylation of acetohydroxy acids Decarboxylation secreted by yeast secreted Yeast uses up diacetyl during active Yeast fermentation, but near end of fermentation, can end up with apparent diacetyl if yeast is overly flocculant, if you rack too much, if yeast is weak rack Dropping temperature too quickly can lead Dropping to VDK problems to VDK VDK Maturation results in lowering of levels Diacetyl rest, ruh storage, krausening Some bacteria (pediococcus, Some lactobacilllus) can result in VDK production production ALDEHYDES ALDEHYDES Many flavor active aldehydes in beer Acetaldehyde reduced to ethanol by end Acetaldehyde of fermentation but oxygen may result in oxidation back to aldehyde oxidation Aldehydes increased by rapid Aldehydes fermentation, temp. increase during fermentation, pressure during fermentation, low wort aeration, Zymomonas infection Zymomonas FLOCCULATION FLOCCULATION Clumping of yeast cells to one another Clumping and sedimentation up (with CO2) or down and Nonflocculant strains are also called Nonflocculant powdery powdery Premature flocculation results in Premature incomplete attenuation (may need to rouse up yeast several times) rouse Nonflocculant yeast may need fining or Nonflocculant centrifugation for clarification centrifugation FLOCCULATION FLOCCULATION Genetically and environmentally Genetically determined determined pH drop and ethanol increase enhance pH flocculation flocculation Overall negative charge on yeast cells Overall decreases, more likely to be attracted to one another one May accelerate flocculation by cooling Highly flocculant strains tend to result in sweeter, less attenuated beer with more diacetyl diacetyl Powdery strains are desirable for some Powdery types of beer types ATTENUATION: amount of sugar ATTENUATION: converted to alcohol as measured by sg converted Lager strains often better at using Lager maltotriose maltotriose Yeast Mutation Yeast Can lead to problems in fermentation as Can brewers yeast are often reused brewers Three main types of mutations: – change from flocculant to nonflocculant increase rate of reproduction Skimming or collecting bottom yeast will tend to Skimming eliminate nonflocculants eliminate – lose ability to ferment maltotriose – respiration deficiency slower fermentation, increase diacetyl YEAST DEGENERATION YEAST Gradual performance degradation Results in slow fermentations, often cease Results too early, poor foam or yeast head too May be related to low oxygen levels or May shortages of Ca, Zn, P or too much Cu or Fe Fe Also, too much trub, high fermentation Also, temperatures, bacteria, washing problems, excessive storage time excessive YEAST DEGENERATION YEAST uvarum more susceptible to yeast uvarum degeneration, contamination, autolysis degeneration, AAAUTOLYSIS: Self degradation of yeast AAAUTOLYSIS: by yeast's own enzymes by Color changes (reddish), aromas change, Color strong sulfury strong Release of volatiles that are not Release associated with tasty beer associated PURE CULTURES PURE Isolated from a number of places, ATCC, Isolated commercial distributors, single cell growup, borrowing from other breweries growup, Single colony (pure culture) technique – Originated with Emil Hanson Carlsburg Originated brewery in CopenhagenPlate out dilute suspension of yeast - get single colonies formed formed – Grow up (slants, liquid media) Single Cell techniques are slightly more complex, where individual cells are isolated and then grown up isolated Lindner's hanging drop procedure, make Lindner's extremely fine pipette that has diameter in the range of cell, drops can have only one cell Never certain otherwise if several cells clumped together start colony clumped Subculturing Subculturing Agar Slants can be used to keep cultures Agar alive Use agar with MYGP (malt, yeast, glucose, peptone) medium peptone) Get growth visible at 20-30 °C in 2-3 days, Get then refrigerate at 4 °C for up to 6 months then Reculture after 3-6 months to keep the yeast Reculture strain from degrading strain More frequent reculturing risks contamination More or mutation Subculturing Subculturing Cover with sterile mineral oil and shelf life Cover can reach 2 years can Oil serves as oxygen barrier and keeps Oil sample from drying out sample Can also store at -70 °C or freeze dry FREEZE DRYING FREEZE Can freeze then dry under vacuum (get Can solid--> vapor) then store samples in liquid nitrogen (-196 °C) °C) Can last for decades (smallpox saved this Can way....keep or destroy) way....keep PROPAGATION OF PURE CULTURE CULTURE Want quick growth to provide enough Want yeast with known characteristics to make beer beer Can lead to less chance for yeast to Can provide beneficial mutations provide Grow up in larger and larger sizes until Grow have enough for brewing have A range of possibilities depending on Co. PROPAGATION OF PURE CULTURE CULTURE Differences between grow up and brewing Differences conditions are minimized by some brewers conditions 10ml - 100ml - 1L - 5L - 20L - 5000L 75-100kgs of yeast for brewery operations MAINTENANCE OF HARVESTED YEAST HARVESTED Yeast obtained from fermentation (barm) Yeast has consistency of thin cream has Don't often use this directly because Don't chance of infection, variable yeast cell concentration concentration Wash and concentrate B4 pitching Periodically wash the yeast or start from Periodically scratch to avoid contamination building up scratch YEAST WASHING YEAST Sterile water – cold, sterile, distilled water – yeast drop before contaminants (flocculate) – repeat Acid wash – llower pH enough with organic acid to kill weak ower yeast, bacteria, etc yeast, – kills most spoilage bacteria (pediococcus kills survives) survives) – pH 2.0-2.3, agitation, 1-2 hours YEAST WASHING YEAST Acid wash – Some acids may affect flocculation, phosphoric Some makes less flocculant, tartaric makes more makes – may negatively affect cell viability, reduce rates may of fermentation of – Maximize effect and minimize problems by Maximize keeping temp below 5 °C, agitate continuously, less than 2 hours, use food grade acid, pitch after washing, do not wash yeast used for high alcohol (>8% abv) fermentations alcohol YEAST WASHING YEAST Ammonium persulfate – Ammonium or sodium perfulfate (0.75%) Ammonium added to slurry then acidified added – Kills most contaminanys but has little effect on Kills yeast yeast Chlorine dioxide – can be used instead of water or acid – Short washing time Short YEAST STORAGE YEAST Yeast may be stored short time as slurry Longer time storage as press cake Have to be careful yeast metabolism is not Have altered altered Measure VIABILITY using SVM (slide Measure viability method), fermentation tests, and flocculation tests SVM SVM Count cells, plate out, grow colonies, Count Can use selective staining using buffered methylene blue, Rhodamille B or eosin methylene Dead cells show up as blue, red or pink Dead respectively respectively ASBC (Am. Soc. Brewing Chem.) use the ASBC methylene blue method methylene – 85% viability cutoff for good cultures ...
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This note was uploaded on 09/19/2011 for the course FST 3124 at Virginia Tech.

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