Industrial Lecture 3

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BIO 425Industrial Microbiology

Seyhun YURDUGÜL

Lecture 3Malolactic fermentation of wines

andother alcoholic beverages



C

ontent outline� Malolactic fermentation

� Beer making� Microorganisms of beer



Malolactic fermentation� After alcoholic fermentation

� Starts about 2-3 weeks after the alcoholicfermentation

� lasts about 2-4 weeks

� Lactic acid bacteria: Lactobacillus, Leuconostoc and Pediococcus species areresponsible





Main rxn of Malolactic Fermentation� L-malic acid L-lactic acid + CO2

ÇDecarboxylation rxn by Malate carboxylase(malolactic enzyme)

Ç

Provides a rise of pH between 0.3-0.5 units



Importance of acidity� Provides a very softer, good quality

drinkable and very good taste

� Completion of malolactic fermentationleads to greater microbiological stability &wines are less prone to spoilage by other

species of bacteria





Taste of wines are designed by

µmalolactic fermentation¶

� Malolactic bacteria produce off-flavors likediacetyl & acetoin to give a fruitful blend

� Increase the attitude through taste

perception� If microbial load increase Unwanted wines





Alternatives to deacidify wines� Yeast addition

� S chizosaccharomyces pombe

� S chizosaccharomyces malidevorans

� Completely metabolize malic acid-anaerobically

� Malic acid Pyruvate Acetaldehyde

Ethanol





Beer production� Fermented extract of malted cereal grains,

principally barley

� Has an ethanol content of 2-6% and adistinctive flavor

� Flavor arises from the constituents of malt,

extracts of hops and products of yeastmetabolism





Beer production� S accharomyces cerevisiae -formerly calledS accharomyces carlsbergensis andS accharomyces uvarum



Outline of processBarley

Malting (Steeping, Germination, Kilning)

Malt

Mashing

Wort

Boiling

Fermentation

Water Adjuncts, Enzymes

Spent grain

Spent hopsHopped wort

Pitching yeast





Malting & Steeping� Cause enzymatic degradation of raw

material into fermentable sugars

� First stage is steeping, where barley grainsare soaked in water at 10-15° C for 3 days

� Increase moisture to speed up germination

� Synthesis of enzymes e.g. amylases, proteases, peptidases



Kilning� Terminates germination

� Heat is applied for drying of grains up to85° C

� Browning Gives the color



Mashing� Extraction of fermentable sugars from the

grain

� Further enzymatic degradation of starch & protein



Fermentation tanks� Designed as

� Large

� Stainless steel, cylindroconical vessels

� Facilities for temperature control, stirringand cleaning in place(CIP)



Fermentation style of beer � Primary & secondary

� Primary ferm. consists separation of

sedimented yeast cells� Further clarification for packaging or

transferred to another tank for low T(3-5°C)

& secondary ferm. + maturation� Secondary ferm. is provided by residualsugar & yeast cells





Important points in fermentation

process where m/o have impact upon beer quality

� Preparation of barley & malt

� Management of the wort� Primary fermentation

� Secondary fermentation & maturation

� Retailing of bulk beer



M/o¶s of beer � For barley & malt

� Depends on climatic conditions,contamination of barley kernels duringharvesting, post harvest storage conditions,m/o growth on kernels during malting, m/o

survival after kilning of malted barley



M/o¶s of beer � Some of the important genera

� Bacteria: Bacillus, Brevibacterium,

Corynebacterium, Microbacterium,

S treptomyces, Pseudomonas spp.

� Yeast: Rhodotorula, S porobolomyces,

Hansenula, Candida, Cryptococcus spp.



M/o¶s of beer � Mold: Alternaria alternata, Cladosporium

spp.

� If moisture of barley (12.5%), A spergillus,Penicillum,Fusarium spp. maysignificantly grow



M/o¶s of beer � At wort phase

� If hygiene is delayed: Bacillus( B.

stearothermophilus, B. coagulans),Clostridium spp.

� Produce lactic & butyric acid, leads to

acidification & off-flavors



Fermentation� Dominated by S accharomyces cerevisiae

� Recombinant DNA & classical geneticmethods are applicable

� Leads µacceptable flavor¶

� Range of strains leads to different varietiesof beers (present in quality assurance

programs of breweries)



Fermentation� Initiated by inoculating(pitching) the wort

with desired S.cerevisiae (start pop. 10-15

X 106 cfu/mL)� Breweries produce own yeast for pitching-

start with a pure slant & propagate cells in

progressively increasing volumes of sterilized wort to give the amount neededfor pitching



Fermentation� On pitching the wort,

� yeasts commence growth with very little lag phase

� Active fermentation is visible by 12-24 hr.

� In 2-4 days, yeasts multiply up to 6-8 X 107

cfu / mL



Main factors affecting the rate of

fermentation� Yeast concentration & viability of yeast

cells inoculated

� Temperature of fermentation

� Dissolved oxygen in the wort at pitchingtime

� Fermentable carbohydrates and available N2

concentration in the wort



If improper hygiene!� Contamination

� Killer yeast strains of S.cerevisiae,

Hanseniaspora, Kluvyeromyces,

Torulaspora, Zygosaccharomyces spp

� Enterobacter spp. , Zymomonas mobilis for

bacterial contaminants.



LIT

ERAT

UREC

IT

ED� Forsythe, S.J. The Microbiology of Safe

Food, Blackwell Science, Cambridge, 2000.

� Wood, B. J. B., Microbiology of FermentedFoods, Vol.1 and 2, Blackie Academic andProfessional, London, Second edition, 1998.

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Industrial Lecture 3