BLBS102-c33 BLBS102-Simpson March 21, 2012 14:5 Trim: 276mm X 219mm Printer Name: Yet to Come
33 Biochemistry of Beer Fermentation 637and elevated wort aeration or wort lipid concentration (Ver-
strepen et al. 2003b). Enhancing ester production is slightly more
complicated.
If it is possible, overpressure or wort aeration can be reduced.
Otherwise, worts rich in glucose and nitrogen combined with
higher fermentation temperatures and lower pitching rates or
application of the drauflassen technique may prove helpful.Biosynthesis of Organic AcidsOver hundred different organic acids have been reported in
beer (Meilgaard 1975b). Important organic acids detected in
beer include pyruvate, acetate, lactate, succinate, pyroglutamate,
malate, citrate,α-ketoglutarate, andα-hydroxyglutarate; and the
medium-chain length fatty acids, caproic (C6), caprylic (C8),
and capric (C10) acid (Coote and Kirsop 1974, Meilgaard 1975b,
Klopper et al. 1986). They influence flavor directly when present
above their taste threshold and by their influence on beer pH.
These components have their origin in raw materials (malt, hops)
and are produced during the beer fermentation. Organic acids,
which are excreted by yeast cells, are synthesized via amino
acid biosynthesis pathways and carbohydrate metabolism. Es-
pecially, they are overflow products of the incomplete Krebs
cycle during beer fermentation. Excretion of organic acids is
influenced by yeast strain and fermentation vigor. Sluggish fer-
mentations lead to lower levels of excretion. Pyruvate excretion
follows the yeast growth: maximal concentration is reached just
before the maximal yeast growth and is next taken up by the
yeast and converted to acetate. Acetate is synthesized quickly
during early fermentation and is later partially reused by the
yeast during yeast growth. At the end of the fermentation, ac-
etate is accumulated. The reduction of pyruvate results in the
production ofd-lactate ofl-lactate (most yeast strains produce
preferentiallyd-lactate). The highest amount of lactate is pro-
duced during the most active fermentation period.
The change in organic acid productivity by disruption of the
gene encoding fumarase (FUM1) has been investigated and it
has been suggested that malate and succinate are produced via
the oxidative pathway of the TCA cycle under static and sake
brewing conditions (Magarifuchi et al. 1995). Using a NAD+-
dependent isocitrate dehydrogenase gene (IDH1,IDH2) disrup-
tant, approximately half of the succinate in sake mash was found
to be synthesized via the oxidative pathway of the TCA cycle in
sake yeast (Asano et al. 1999).
Sake yeast strains possessing various organic acid productivi-
ties were isolated by gene disruption (Arikawa et al. 1999). Sake
fermented using the aconitase gene (ACO1) disruptant contained
a twofold higher concentration of malate and a twofold lower
concentration of succinate than that made using the wild-type
strain. The fumarate reductase gene (OSM1) disruptant produced
sake containing a 1.5-fold higher concentration of succinate,
whereas theα-ketoglutarate dehydrogenase gene (KGD1)and
fumarase gene (FUM1) disruptants gave lower succinate con-
centrations. InS. cerevisiae, there are two isoenzymes of fu-
marate reductase (FRDS1 and FDRS2), encoded by theFRDS
andOSM1genes, respectively (Arikawa et al. 1998). Recent
results suggest that these isoenzymes are required for the reox-idation of intracellular NADH under anaerobic conditions, but
not under aerobic conditions (Enomoto et al. 2002).
Succinate dehydrogenase is an enzyme of the TCA cycle
and thus essential for respiration. InS. cerevisiae, this enzyme
is composed of four nonidentical subunits, that is, the flavo-
protein, the iron–sulfur protein, the cytochromeb 560 , and the
ubiquinone reduction protein encoded by theSDH1,SDH2,
SDH3,andSDH4genes, respectively (Lombardo et al. 1990,
Chapman et al. 1992, Bullis and Lamire 1994, Daignan-Fournier
et al. 1994). Sdh1p and Sdh2p comprise the catalytic domain in-
volved in succinate oxidation. These proteins are anchored to
the inner mitochondrial membrane by Sdh3p and Sdh4p, which
are necessary for electron transfer and ubiquinone reduction,
and constitute the succinate:ubiquinone oxidoreductase (com-
plex II) of the electron transport chain. Single or double disrup-
tants of theSDH1,SDH1b(which is a homologue of theSDH1
gene),SDH2,SDH3,andSDH4genes have been constructed and
shown that the succinate dehydrogenase activity was retained in
theSDH2disruptant and that double disruption ofSDH1and
SDH2orSDH1bgenes is necessary to cause deficiency of suc-
cinate dehydrogenase activity in sake yeast (Kubo et al. 2000).
The role of each subunit in succinate dehydrogenase activity and
the effect of succinate dehydrogenase on succinate production
using strains that were deficient in succinate dehydrogenase,
have also been determined. The results suggested that succinate
dehydrogenase activity contributes to succinate production un-
der shaking conditions, but not under static and sake brewing
conditions.
The medium-chain fatty acids account for 85–90% of the
fatty acids in beer and impart an undesirable goaty, sweaty, and
yeasty flavor (Chen 1980). These fatty acids are produced de
novo by yeast during anaerobic fermentation and are not the
result ofβ-oxydation of wort or yeast long-chain fatty acids. As
a result, any change in fermentation conditions that promote the
extent of yeast growth also favor increased levels of medium-
chain fatty acids in beer. Higher temperature, increased wort
oxygenation, and possibly elevated pitching rates are all effective
in this respect (Boulton and Quain 2006). The presence of these
medium-chain fatty acids in beer is also related to yeast autolysis
(Masschelein 1981). Yeast autolytic off-flavors are stimulated
at high temperatures, high yeast concentrations, and prolonged
contact times at the end of primary fermentation and during
secondary fermentation.Biosynthesis of Vicinal DiketonesVicinal diketones are ketones with two adjacent carbonyl groups.
During fermentations, these flavor-active compounds are pro-
duced as by-products of the synthesis pathway of isoleucine,
leucine, and valine (ILV pathway) (see Fig. 33.3) and thus
also linked to amino acid metabolism (Nakatani et al. 1984)
and the synthesis of higher alcohols. They impart a “buttery”,
“butterscotch” aroma to alcoholic drinks. Two of these com-
pounds are important in beer, that is, diacetyl (2,3-butanedione)
and 2,3-pentanedione. Diacetyl is quantitatively more impor-
tant than 2,3-pentanedione. It has a taste threshold in lager
beer from 17μg/L (Saison et al. 2009) to 150μg/L (Meilgaard