Food Biochemistry and Food Processing (2 edition)

BLBS102-c33 BLBS102-Simpson March 21, 2012 14:5 Trim: 276mm X 219mm Printer Name: Yet to Come

638 Part 5: Fruits, Vegetables, and Cereals

Threonine

Threonine

α-Ketobutyrate

α-Acetohydroxybutyrate

Isoleucine

Pyruvate

α-Acetolactate α-Acetolactate

Valine Leucine

α-Acetohydroxy-

butyrate

Diacetyl

Acetoin

2,3-Butanediol

2,3-Pentane-

dione

2,3-Pentanediol

Acetylethyl-

carbinol

CO 2 CO 2

Carbohydrate

metabolism

Bacterial

acetolactate

decarboxylase

ILV5 CO 2

Figure 33.3.The synthesis and reduction of vicinal diketones inSaccharomyces cerevisiae.

1975b), approximately 10 times lower than that of pentanedione

(Wainwright 1973).

The excretedα-acetohydroxy acids are overflow products of

the ILV pathway that are nonenzymatically degraded to the

corresponding vicinal diketones (Inoue et al. 1968). Tetraploid

gene dosage series for variousILVgenes have been constructed

and the obtained yeast strains were used to study the influence

of the copy number ofILVgenes on the production of vicinal

diketones (Debourg et al. 1990, Debourg 2002). It was shown

that theILV5activity is the rate-limiting step in the ILV path-

way and responsible for the overflow (Fig. 33.3). The nonen-

zymatic oxidative decarboxylation step is the rate-limiting step

and proceeds faster at a higher temperature and a lower pH (In-

oue and Yamamoto 1970, Haukeli and Lie 1978). The produced

amount ofα-acetolactate is very dependent on the used yeast

strain. The production increases with increasing yeast growth.

For a classical fermentation, 0.6 ppmα-acetolactate is formed

(Delvaux 1998). At high aeration, this value can be increased

to 0.9 ppm and in cylindro-conical fermentations tanks even

to 1.2–1.5 ppm.

It has been shown that valine inhibits the synthesis of

α-acetolactate through feedback inhibition (Magee and de

Robinson-Szulmajster 1968). This inhibition is directed to the

protein Ilv6p, which is the regulatory subunit of acetohydroxy

acid synthase (the catalytic subunit encoded byILV2)(Pang and

Duggleby 1999, 2001). Because the uptake of valine is delayed

in a normal beer fermentation, the suppressive effect of va-

line accounts for the postponed onset of total diacetyl (sum

of actual diacetyl andα-acetolactate), and this effect persists

longer in worts with high levels of FAN content. In contrast,

low FAN levels give two diacetyl peaks as a result of the re-

quirement for valine biosynthesis. Therefore, a minimum FAN

level above the critical value of 50 ppm (Nakatani et al. 1984)

or 140 ppm (Pugh et al. 1997) should be maintained dur-

ing the fermentation to ensure the presence of valine in the

fermenting wort.

Yeast cells posses the necessary enzymes (reductases) to re-

duce diacetyl to acetoin and further to 2,3-butanediol, and 2,3-

pentanedione to 2,3-pentanediol (Bamforth and Kanauchi 2004).

These reduced compounds have much higher taste thresholds

and have no impact on the beer flavor (Van Den Berg et al.

1983). The reduction reactions are yeast strain dependent. The

reduction occurs at the end of the main fermentation and during

the maturation. Sufficient yeast cells in suspension are necessary

to obtain an efficient reduction. Yeast strains that flocculate early

during the main fermentation needs a long maturation time to

reduce the vicinal diketones. Diacetyl can be complexed using

SO 2. These complexes cannot be reduced, but diacetyl can again

be liberated at a later stage by aldehydes. This situation is es-

pecially applicable to yeast strains, which produce a lot of SO 2.

Worts, which are produced using a high content of adjuncts, can

be low in free amino acid content. These worts can give rise to

a high diacetyl peak at the end of the fermentation.

There are several strategies, which can be chosen to reduce

the vicinal diketones amount during fermentation:

1. Since the temperature has a positive effect on the reduction
   efficiency of theα-acetohydroxy acids, a warm rest period
   at the end of the main fermentation and a warm maturation
   are applied in many breweries. In this case, temperature
   should be well controlled to avoid yeast autolysis.


2. Since the rapid removal of vicinal diketones requires yeast
   cells in an active metabolic condition, the addition of
   5–10% Krausen (containing active, growing yeast) is a
   procedure, which gives enhanced transformation of vic-
   inal diketones (NN 2000). This procedure can lead to