CZ2017829A3 - The yeast strain of Saccharomyces cerevisiae CCM 8822 and its use in beer production - Google Patents

Abstract

The invention relates to a strain of CCM 8822 isolated from spontaneous fermentation of grape must which aerobically and anaerobically processes sucrose, raffinose, maltose, lactate and glucose, aerobically processes galactose, anaerobically treats trehalose and glycerol, and hydrolyses esulin, forming ellipsoidal to spherical cells. 6 - 8 μm x 4 - 6 μm, which does not form a mycelium. The invention further relates to beer production in which the main wort fermentation is carried out by the yeast strain CCM 8822.

Description

CZ 2017 - 826 A3

Yeast strain Saccharomyces cerevisiae CCM 8822 and its use in beer production

Technical field

The present invention relates to a novel yeast strain of Saccharomyces cerevisiae CCM 8822.

The invention further relates to the use of this yeast strain in the production of beer.

BACKGROUND OF THE INVENTION

Beer production consists of controlled fermentation of sugars contained in wort by brewer's yeast. Brewer's yeasts are cultural yeasts of several species and genera: Saccharomyces pastorianus is used to produce bottom-fermented beers, Saccharomyces cerevisiae is used to produce top-fermented beers, and specialty beers such as Brettanomyces or Torulaspora are produced (see for example T. Kochláňová et al .: "Yeast nonSaccharomyces and their importance in the brewing industry. Part I - Brettanomyces (Dekkera)" and "Yeast \ vm-Saccharomyces and their importance in the brewing industry. Part II", both Kvasný Průmysl 62/2016 ( 7-8)). In doing so, brewer's yeast converts carbohydrates into ethanol, carbon dioxide, and the energy required for yeast propagation, as well as about two thousand other substances, especially higher alcohols, esters, aldehydes, etc., which affect the resulting sensory properties of the finished beer. The overall ratio of these higher alcohols and esters is then very important for the overall sensory impression, since the individual compounds themselves are very often below the threshold concentration of sensory perception. For example, it is generally known that the strains of Saccharomyces cerevisiae produce a greater amount and a wider spectrum of sensory active substances than Saccharomyces pastorianus strains (see, e.g., Lodolo et al., "The Saccharomyces cerevisiae - the main character in beer brewing. FEMS Yeast Res 8 (7): 1018-36, 2008), and that the "lighter" taste of bottom-fermented beers causes, in addition to the lower main fermentation temperature, an altered expression of esters responsible for the production of esters in Saccharomyces pastorianus (see eg, Procopio et al., "Function and Regulation of Yeast Genes Involved in Higher Alcohol and Ester Metabolism during Beverage Fermentation", European Food Research and Technology 233 (5): 721729, November 2011).

Wine production then consists in fermentation of carbohydrates present in grape must with wine yeast. This fermentation results in the conversion of carbohydrates to ethanol and carbon dioxide, and the energy needed to propagate the yeast. Wine yeast is more resistant to alcohol than brewing yeast. They enter the must either from the surface of the berries (spontaneous fermentation) or by adding a pure culture of noble yeast (pure fermentation). Cultural wine yeasts belong to the species Saccharomyces cerevisiae and Saccharomyces bayanus.

SUMMARY OF THE INVENTION

The present invention provides a novel yeast strain of Saccharomyces cerevisiae CCM 8822, which is isolated from spontaneous fermentation of grape must. It was prepared for further use by selection and re-boiling on wort agar with tetracycline. Cultivation was then performed aerobically in a microbiological incubator at 26 ° C for 3-5 days. After the growth of the yeast culture on the agar surface, this culture was transferred with a sterilized inoculating loop to a new Petri agar dish and seeded with a so-called cross smear diluting the yeast concentration on the agar. This procedure was repeated until the yeast culture had grown to form visibly separated colonies. Subsequent isolation of the yeast strain Saccharomyces cerevisiae CCM 8822 was then performed by inoculating the selected colony onto a petri dish containing wort agar.

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The strain thus isolated was then characterized by biochemical assays and polymerase chain reaction (PCR).

The yeast strain Saccharomyces cerevisiae CCM 8822 produces ellipsoidal to spherical cells having dimensions of 6-8 µm x 4-6 µm that do not form mycelium and rarely form pseudomycelium.

Appearance and consistency of colonies - the coating is pasty, creamy, light brown, smooth, glossy. Character of growth in liquid medium - it forms sediment in liquids, wastes never form.

Saccharomyces cerevisiae strain CCM 8822 utilizes both aerobic (oxidized) and anaerobic (fermented) sucrose, raffinose, maltose, lactate and glucose. Aerobically uses galactose. Anaerobically uses trehalose and glycerol. Hydro skis esculin.

The yeast strain Saccharomyces cerevisiae CCM 8822 has some advantageous properties for use in the brewing process compared to the brewer's yeast Saccharomyces cerevisiae and Saccharomyces pastorianus. In particular, it involves the production of only a minimal amount of diacetyl, which gives the finished beer a taste for butter and is considered undesirable at higher concentrations. The optimum temperature for beer production using this strain is in the range of 18-22 ° C, where the yeast produces the optimal amount of sensory active substances; however, the strain can be used for fermentation at temperatures of 12-16 ° C, and even at these temperatures it is characterized by a high fermentation rate.

The yeast strain Saccharomyces cerevisiae CCM 8822 according to the invention is deposited in the collection of microorganisms of the Research Institute of Brewing and Malting, as, designated RIBM code RIBM 193 and in the Czech Collection of Microorganisms in Brno under CCM 8822.

DETAILED DESCRIPTION OF THE INVENTION

Example 1: Preparation of the yeast strain Saccharomyces cerevisiae CCM 8822 for laboratory fermentation tests

In the preparation of the yeast strain Saccharomyces cerevisiae CCM 8822 for practical use - e.g. for further testing or for the production of beer, a pure culture of this strain obtained by isolation from spontaneous grape must fermentation (see above) and used until of the preparation was stored in a refrigerator at 2 ± 1 ° C.

A yeast strain of Saccharomyces cerevisiae CCM 8822 was inoculated on sloping wort agar in cotton-stoppered tubes. From the yeast growth formed, a portion of the biomass was then sterile seeded into 10 ml of 10% sterile wort and cultivated at 25 ° C for days. The top fermentation layer was then discarded and the yeast sediment was transferred to 50 ml of 10% sterile wort, followed by cultivation at 20 ° C for 2 days. Subsequently, the strain was multiplied by converting it to an ever larger volume in a ratio of approximately 1: 4. The yeast sediment was first transferred to 250 ml of 10% sterile wort, where it was propagated at 17 ± 1 ° C for 5 days, then the yeast sediment formed was transferred. to 1000 ml of 10% wort, where cultivation was carried out at 17 ± 1 ° C for 3 days.

The yeast strain Saccharomyces cerevisiae CCM 8822 prepared in this way can be used directly for laboratory fermentation tests or for beer production. If necessary, it is washed again with water (repeated sedimentation and washing) to remove the culture medium.

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Example 2: Preparation of beer using the yeast strain Saccharomyces cerevisiae CCM 8822

A concentrated yeast concentrate of yeast Saccharomyces cerevisiae CCM 8822, prepared as described in Example 1, was added to the wort with the original extract of about 12% (but wort beer with another extract can be prepared by the same procedure). The fermentation dose was approximately 30 million cells / ml wort. Subsequent fermentation was carried out in a sterilized plastic vessel at 21 ° C ± 1 ° C for 5 days.

During fermentation, the course of cell proliferation and sedimentation was monitored by counting the cells by microscopic technique using a Bürker chamber. In addition, extract loss, fermentation, alcohol concentration and pH change were further measured. The measured values are given in Table 1.

Table 1

Measured parameter Oh 24 h 48 h 72 h 96 h 120 h Alcohol (% vol) 0 1.96 3.95 3.96 3.97 3.97 Apparent fermentation (%) 0 31.75 64.82 65.46 66.50 66.64 Apparent extract (% by weight) 11.87 7.83 4.02 3.82 3.78 3.74 pH 5.74 4.34 4.08 4.12 4.18 4.20 Number of cells (mil / ml) 30.8 77.35 120 59.5 46.2 34.5

After fermentation, the beer thus prepared was transferred to lager vessels and stored at a temperature of 2 ± 1 ° C in a lager cellar. After 6 weeks of lying, it was measured for pH, fermentation, extract, alcohol content, diacetyl, pentanedione, acetaldehyde, and dimethylsulfide (DMS) concentrations, the measured values are shown in Table 2.

As can be seen, the strain of Saccharomyces cerevisiae CCM 8822 produces only low concentrations of diacetyl, which is responsible for the undesirable buttery flavor of beer, compared to conventional brewer's yeast.

Table 2

Measured parameter Value Apparent extract (% wt.) 3.72 Extract real (% wt.) 5.18 Alcohol (% vol) 4.00 Apparent fermentation (%) 66.9 Diacetyl (mg / l) 0.011 Pentandione (mg / l) <0.005 Acetaldehyde (mg / l) 40.75 DMS (pg / l) 19 Dec

In addition, the content of higher alcohols and esters, which significantly contribute to the sensory profile of the finished beverage, was also measured with this beer. Table 3 lists selected higher alcohols and esters and the ratio of total higher alcohols to esters (A / E ratio). The value of this ratio indicates that the yeast Saccharomyces cerevisiae CCM 8822 behaves

A3 CZ 2017 - 829 A3, similar to the top brewer's yeast, but it produces higher amounts of betaphenyl alcohol than most of their strains (usually up to 4) and at the same time very low amounts of diacetyl and pentanedione. This is very advantageous from the point of view of beer production technology, since it is not necessary to include a long delay in the production process for the degradation of diacetyl.

Table 3

Monitored substance Content ethyl formate (mg / 1) 0.04 ethyl acetate (mg / l) 9.52 propyl acetate (mg / l) 0.06 isobutyl acetate (mg / l) 0,103 ethylbutyrate (mg / l) 0.157 Propanol (mg / l) 24.25 isobutanol (mg / l) 11.39 isoamyl acetate (mg / l) 0.8 2 and 3-methyl-butanol (mg / l) 45.24 ethyl caproate (mg / 1) 0.154 ethylcaprylate (mg / 1) 0.174 ethyl hexanol (mg / l) 0.012 furfuryl alcohol (mg / l) 0,054 Phenylethyl acetate (mg / 1) 0.314 β-phenyl alcohol (mg / l) 16.99 sum of alcohols (mg / 1) 97.94 sum of esters (mg / 1) 11.39 A / E ratio 8.60

In addition, the production of phenolic substances responsible for the medicinal or clove aromas of beer was also analyzed. The values obtained are shown in Table 4.

Table 4

Monitored substance Content 4-vinylguaiacol (mg / l) 0,654 Tyrosol (mg / l) 9.99

A sensory evaluation of beer was also carried out by the tasting committee. The beverage prepared using the yeast Saccharomyces cerevisiae CCM 8822 showed no sensory defects, was pleasantly saturated, with a pleasant scent of coriander and clove and tasted like top-fermented beer. Bitterness was evaluated as slightly adhering. The overall subjective impression was assessed as above average.

PATENT CLAIMS

Claims (2)

1. The yeast strain Saccharomyces cerevisiae deposited with the Czech Collection of Microorganisms under number 8822 isolated from spontaneous fermentation of grape must, which aerobically and anaerobically treats sucrose, raffinose, maltose, lactate and glucose, aerobically processes galactose, hydrolyzes and anaerobes, and which produces ellipsoidal to spherical cells with dimensions of 6-8 µm x 4-6 µm that do not form mycelium. A method for producing beer, characterized in that the main wort fermentation is carried out by the yeast strain Saccharomyces cerevisiae CCM 8822 according to claim 1.