Method of producing a composite fermented beverage using...

Details Method of producing a composite fermented beverage using... Method of producing a composite fermented beverage using...

1998-09-15

2001-12-04

Slobodyansky, Elizabeth (Department: 1761)

Chemistry: molecular biology and microbiology

Micro-organism, per se ; compositions thereof; proces of...

Fungi

C435S254100, C435S254210, C435S093000, C435S194000, C435S254110

Reexamination Certificate

active

06326184

ABSTRACT:

FIELD OF INVENTION
The invention relates to the field of yeast fermented beverages including beer, sake, wine and whisky and in particular the invention is concerned with the provision of customized composite yeast fermented beverages having a predetermined content of flavour compounds and/or flavour compound stabilizing compounds, which are made by combining two or more separate batches of yeast fermentates each having different contents of yeast-generated flavour compounds and/or compounds that are capable of stabilizing such flavour compounds. The invention also provides novel, genetically modified yeast strains that are useful in the invention.
Specifically, the invention has made it possible to make a composite yeast fermented beverage product such as lager beer having a predetermined content of one or more flavour compound(s) and/or compound(s) that stabilize(s) flavour compounds e.g. by providing at least one batch of fermented beverage which is produced by using a yeast strain which is modified to substantially not produce at least one flavour compound and/or compound stabilizing flavour compounds (base batch of fermentate) and combining such a batch with one or more batches of beverage fermentates that contain(s) a level of flavour compounds and/or stabilizing compounds which is normal for a fermentate produced by using a conventional yeast strain or which is produced by a modified yeast strain having, relative to conventionally used yeast strains, an enhanced production of at least one flavour compound or flavour compound stabilizing compound.
TECHNICAL BACKGROUND AND PRIOR ART
The sensory quality of a yeast fermented beverage such as beer depends largely on the particular brewing yeast strain which is used. During the beer brewing process, the yeast will in addition to ethanol produce a large variety of minor metabolites which affect the taste and flavour of the beer. Whereas the presence of certain of such minor metabolites may, at least up to certain levels, confer desirable sensory characteristics to the beer, several minor metabolites are undesired and may even, if present in higher amounts, result in spoilage of the beer.
Yeast fermented beverages including beer contain more than a hundred minor metabolites, of which some, at least when they are present above certain concentrations, will confer to the beverage an undesirable taste and flavour. A comprehensive review of beer flavouring compounds has been given by Meilgaard (1975) to which there is referred. In this connection, the most important metabolites include sulphur compounds such as dimethyl sulphide (DMS), hydrogen sulphide (H
2
S), thiols and thioesters which at certain levels will result in undesirable sulphur-containing flavours.
DMS is a thioether of great importance for the aroma and flavour of beer. The content of DMS in conventional lager beers regularly exceeds the taste threshold level of about 30 &mgr;g/L (Meilgaard, 1975) which explains the focus that has been on this compound. Above its threshold level but below about 100 &mgr;g/L DMS contributes to the distinctive taste of some lager beers. When present at concentrations above 100 &mgr;g/L DMS may, however, impart a generally undesired flavour described as “cooked sweet corn”. DMS in beer may be derived from thermal degradation of S-methyl-methionine during kiln-drying and wort preparation and it has been suggested that this is the only pathway of significance for the final DMS content in beer (Dickenson and Anderson, 1981; Dickenson, 1983). However, substantial evidence suggests that enzymatic conversion of dimethyl sulphoxide (DMSO) to DMS by the brewing yeast is of great importance and, under some circumstances, even the major source of the final DMS level in beer (Leemans et al., 1993).
There is no doubt, that Saccharomyces strains do contain an enzymatic activity that can reduce DMSO to DMS in a NADPH-dependent manner (Zinder and Brock, 1978a; Anness et al., 1979; Anness, 1980). A multicomponent methionine sulphoxide (MetSO) reductase (EC 1.8.4.5) has been isolated from yeast (Black et al., 1960; Porqué et al., 1970), and it has been suggested that this activity is identical to the DMSO reductase activity (Anness et al., 1979; Anness, 1980; Bamforth, 1980; Bamforth and Anness, 1979; 1981). The normal function of MetSO reductase seems to be reduction of oxidized methionines of cellular proteins which is in agreement with observations showing that the enzyme has a higher affinity for MetSO than for DMSO (Bamforth and Anness, 1979; 1981) and that MetSO inhibits DMSO reduction (Anness et al., 1979; Anness, 1980; Bamforth and Anness, 1981). The consequence hereof is that the degree of MetSO formation during kiln-drying of the malt will affect the degree of DMSO reduction.
The nitrogen content of the growth medium also seems to affect DMS formation by yeast. Thus, high amounts of highly assimilable nitrogen keep DMSO reductase activity at a low level, whereas enzyme activity is induced under nitrogen-limiting conditions (Gibson et al., 1985). The high nitrogen content of most worts would appear to keep DMSO reduction at the base level during fermentation. See Anness and Bamforth (1982) for a review on DMS formation in beer production. So-called DMSO reductases from some prokaryotes have been characterized, as well as the genes encoding them (Satoh and Kurihara, 1987; Bilous et al., 1988; Weiner et al., 1992; Yamamoto et al., 1995). While such enzymes, in some cases, have MetSO reducing capabilities, their main purpose in the bacterial cell is probably to act as the terminal step in DMSO respiration (Zinder and Brock, 1978b). Peptide methionine sulphoxide reductases (PMSRs) probably fulfil the role of repairing oxidised methionine species in proteins, thereby restoring their biological activity.
Sulphite is another sulphur-containing metabolite being formed during yeast fermentation. Sulphite is a versatile food additive used for preservation of foodstuffs. In beer sulphite has a dual activity in that it acts both as an antioxidant and as an agent that masks certain off-flavours. Due to its great importance, considerable work has been carried out to elucidate the physiology of sulphur metabolism in Saccharomyces species in relation to sulphite production.
Sulphite is produced by the yeast during beer fermentation and is present in the final beer. The compound is produced via the sulphur assimilation pathway. Inorganic sulphate is taken up through two sulphate permeases encoded by SUL1 and SUL2, respectively (Cherest et al., 1997). Intracellularly, sulphate is converted to adenylylsulphate (APS) by the action of ATP sulphurylase (EC 2.7.7.4) encoded by MET3. In a subsequent step, the MET14 encoded APS kinase (EC 2.7.1.25) catalyses the formation of phosphoadenylylsulphate (PAPS) which in turn is reduced by PAPS reductase (EC 1.8.99.4) (MET16 encoded) to sulphite.
Hydrogen sulphide results from reduction of sulphite and in this form inorganic sulphur is incorporated into organic compounds by fusion with O-acetyl homoserine leading to the formation of homocysteine. The latter compound is the precursor for biosynthesis of cysteine, methionine and S-adenosylmethionine (SAM). SAM represses transcriptionally all MET-genes (Cherest et al., 1985; Sangsoda et al., 1985; Langin et al., 1986; Thomas et al., 1989; Thomas et al., 1990; Korch et al., 1991; Mountain et al., 1991; Hansen et al., 1994).
Hydrogen sulphide is, as it is mentioned above, an intermediate in the sulphur assimilatory pathway of Saccharomyces spp. It is also the point of entry into the methionine biosynthetic pathway of carbon backbones derived from the threonine biosynthetic pathway. It is derived in four enzymatic steps from inorganic sulphate ions obtained from the growth medium of the yeast.
Besides being an intermediate in the yeast sulphur metabolism, hydrogen sulphide is an important flavour compound in beer and its distinct taste of “putrefied eggs” generally renders this compound undesired in beers, except at very low concentrations, where it may aid in disguising the taste

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