PRODUCTION OF BEER USING IMMOBILIZED YEAST ENCODING Α-Acetolactate DECARBOXYLASE

Abstract

Genetically modified brewer's yeast encoding a-acetolactate decarboxylase (a-ALDC) was tested in immobilized yeast bioreactors for main fermentation of beer. The a-ALDC enzyme produced by the transformant catalyzes the direct conversion of a-acetolactate to acetoin without formation of diacetyl. The long lagering period required for beer maturation in conventional brewing can thus be shortened or even omitted.Three different packed bed bioreactors were employed, with volumes of 1.6 dm3, 5 dm3 and 25 dm3.The 5 dm3 column had a slightly conical geometry in contrast to the others which had cylindrical shapes. Sintered glass beads were chosen as the carrier material on the basis of experiments with the parent strain.The brewing performance of the transformant compared well with that of the parent strain in the immobilized system. Fermentation, utilization of amino acids (including isoleucine, valine and leucine) and flavour formation were practically identical with both strains, the only difference being a marked decrease in the formation of diacetyl by the transformant. Small differences were, however, observed in the long-term biochemical stability.By using yeast encoding a-ALDC in the immobilized yeast system the total (primary and secondary) fermentation time could be reduced to approximately 2-6 days, compared with 3-6 weeks in a conven tional batch process.

Characteristics of High Cell Density Fermentations with Different Lager Yeast Strains

“…However, several strategies can be followed to decrease the diacetyl content in beer, such as accelerated maturation using immobilised yeast 27 , supplementation of α-acetolactate decarboxylase to the wort 19 or the use of genetically manipulated yeast strains 20,32 . Next to this, the implementation of TFL-resistant mutants, which produce higher amounts of isoamyl alcohol and isoamyl acetate, in HD fermentations, seems pointless, because of the higher production rate of α-acetolactate during fermentation, resulting in a higher diacetyl content at the end of fermentation.…”

Primary beer fermentation by immobilised yeast—a review on flavour formation and control strategies

“…92 -95 The optimisation of oxygen availability appears critical for the control of flavour-active compound formation in immobilised systems. 96 Kronlöf and Linko 97 successfully ensured a constant dissolved oxygen concentration in their immobilised yeast reactor by sparging with a mixture of air and nitrogen as an inert gas. In a gas-lift bioreactor system with yeast cells immobilised in alginate microbeads and on spent grains, an optimum flavour and aroma profile of the continuous produced green beer was obtained at zero airflow in the gas feed (mixing induced by pure N 2 and CO 2 , respectively) and temperatures between 13 and 17…”

A Review of Flavour Formation in Continuous Beer Fermentations*

“…The viability and fermentation capacity (vitality) of immobilized brewing yeast in continuous fermentations have already been reported to decrease 10,37,45,46,72 . However, there is little known about the impact of senescence and aging of immobilized yeast in continuous fermentation on product quality.…”

“…Addition of bacterial α-acetolactate decarboxylase, to the fermenting beer, thus converting α-acetolactate directly to acetoin 30 and/or the use of genetically modified brewer's yeast e.g. encoding an α-acetolactate decarboxylase would lead to lower final diacetyl level in continuously fermented beer 46,109,118 . However, the legal regulation regarding enzyme supplementation and the lack of consumer acceptance towards genetically modified yeast favour other methods of diacetyl control.…”

“…Since acetaldehyde is mostly formed during the active growth phase of yeast, its content can be controlled by proper oxygen supply 10 . As well, it can be lowered by prolonged maturation 46 .…”