JPH0746982B2 - Mutant yeast culture method - Google Patents

Description

The present invention relates to a method for producing an alcoholic beverage or the like having a high scent.

More specifically, the present invention produces various alcoholic beverages, foods, and fragrances by using a mutant yeast that has produced a large amount of caproic acid and / or ethyl caproate among aroma components by mutation. It is about how to do it.

In general, aroma is an important factor that determines the quality of alcoholic beverages and foods, and aroma ester ethyl caproate is one of the preferable aroma components similar to the aroma of apple. This ethyl caproate is produced by yeast from caproic acid and ethanol, but its production amount is generally too small.

That is, as a result of the studies by the present inventors, it was clarified that it is caproic acid that is the rate-determining agent for the production of ethyl caproate in yeast. And, this caproic acid is produced in the middle of the fatty acid biosynthesis system of yeast,
The amount of ethyl caproate derived from caproic acid is negligible because it is converted to the next compound without being accumulated as caproic acid.

Therefore, the present inventors have a mutation in fatty acid synthase,
After intensively researching for a mutant yeast having a large amount of caproic acid, a method for selectively obtaining a mutant yeast that produces a large amount of this was found, and using this mutant yeast, a method for producing a fragrant alcoholic beverage, etc. It was developed.

In order to obtain a mutant yeast in the present invention, any method may be used as a mutation method. Examples of physical methods of mutation include ultraviolet irradiation and irradiation, and chemical methods include mutagenizing agents such as ethyl methane sulfonate and
N-methyl-N-nitro-N-nitrosoguanidine, nitrous acid,
There is a mutation method of suspending in a solution such as an acridine dye.

In the present invention, these mutation methods can be appropriately used, but they have a feature in selection of mutant yeast. That is, the growth medium of the mutant strain must contain cerulenin.

Cerulenin is known as an antibiotic that inhibits fatty acid synthase, and acquiring resistance to cerulenin means that the fatty acid synthase or its function has changed.

Therefore, when the properties of this cerulenin-resistant strain were examined, it was found that there were many strains in which the intermediate fatty acid containing caproic acid was increased compared to the parent strain.

Cerulenin may be added in either a solid medium or a liquid medium, and is used after adding about 25 μM.

As treated yeast, sake yeast, shochu yeast, brewer's yeast,
A cerulenin-resistant strain can be obtained from any of yeasts such as wine yeast and baker's yeast.

After mutating various yeasts, transfer to cerulenin-containing medium, isolate the strains grown by culturing at 30 ° C for about 1 week,
From this, yeast that produces a large amount of caproic acid may be adopted.

The mutant yeasts obtained here are capable of producing caproic acid well in any of sake yeast, shochu yeast, brewer's yeast, wine yeast, and baker's yeast, so using these, sake, shochu, beer, wine , Bread, etc., can be used to produce products containing a large amount of ethyl caproate.

Example 1 A haploid yeast (hereinafter referred to as K-
7-H is abbreviated as YPD medium (yeast extract 1%, peptone 2).
%, Glucose 2%) and incubating for 1 day, the cells were collected and washed. To the washed cells, 5 ml of 0.2 M phosphate buffer (pH 8.0), 0.25 ml of 40% glucose solution and 0.25 ml of ethyl methane sulphonate were added, and the mixture was subjected to mutagenesis at 30 ° C. for 1 hour with slow stirring. After the treatment, the cells were washed with sterile water and spread on YPD agar medium (YPD medium plus 2% agar) containing cerulenin (final concentration 25 μM). A large number of cerulenin-resistant strains grown on this medium were inoculated into YNBC medium (Daifuco yeast nitrogen base 0.67%, glucose 2%, casamino acid 1%) and cultured at 30 ° C for 3 days to produce caproic acid in the medium. The concentration was measured. Here, strain 7-C-8 having improved caproic acid-producing ability as compared with the parent strain was obtained.
This strain has been deposited with MIC institute as FERM-P-8452. Table 1 shows the concentration of caproic acid in the medium when 7-C-8 and the parent strain K-7-H were cultured in the YNBC medium at 30 ° C. for 3 days.

As shown in Table 1, it was found that the 7-C-8 strain produced about 6 times as much caproic acid as the parent strain.

Example 2 Sake was produced using the mutant yeast 7-C-8 (FERM P-8452) and the No. 7 yeast of the Association with the preparation shown in Table 2 below.

The yeast was fermented at 15 ° C for 15 days by adding 2 g of the cultured yeast in wet cell weight. The ingredients of the sake thus obtained are shown in Table 3.

As a result, among the aroma components of sake, ethyl caproate, which is one of the most important aroma components, increased about 5 times as much as the parent strain, and a sensory test showed a strong apple-like aroma.

In addition, caproic acid, which is one of the substrates for ethyl caproate, increased about 6 times compared to the parent strain. Cerulenin
Since it is an inhibitor of fatty acid synthase and the biosynthesis of caproic acid in yeast is carried out by fatty acid synthase, the resistant strain mutates in the fatty acid synthase and the amount of caproic acid produced increases. It is thought that it was done.

Example 3 Mutant yeast 7-C-8 (FERM P-8452) and wine yeast Saccha
Wine was brewed with romyces cerevisiae (IAM 4274).

Fresh Koshu grape juice is supplemented with glucose to add sugar 24
Adjusted to%. 2 g of each yeast was added to this fruit juice 1 as a wet cell weight and fermented at 18 ° C. for 7 days. The ingredients of the wine obtained here are shown in Table 4.

For wines made with 7-C-8, ethyl caproate is IAM 427.
It was five times as high as four, and it was a new type that was sensually different from the wines of the past.

Example 4 Mutant yeast 7-C-8 (FERM P-8452) and wine yeast Saccha
Brandy was produced using romyces cerevisiae (IAM 4274).

Fresh Koshu grape juice was supplemented with glucose to adjust the sugar content to 24%. 2 g of each yeast was added to this fruit juice 1 as a wet cell weight, and the mixture was fermented at 18 ° C. for 7 days. The obtained fermentation broth was distilled under reduced pressure using a rotary evaporator. The ingredients of the brandy thus obtained are shown in Table 5.

The brandy using 7-C-8 had a high concentration of ethyl caproate and was sensory different from the conventional brandy.

Example 5 Mutant yeast 7-C-8 (FERM P-8452) and brewer's yeast Saccha
Beer was brewed using romyces uvarum (IFO 0565).

Water 1 was added to 160 g of malt to heat and saccharify, and then filtered to obtain wort. After adding 2 g of hops and boiling, the hops were removed, and after cooling, water was added to bring the total amount to 1. This wort 1
Add 2 g of each yeast as wet cell weight to 15 ℃
Fermented for 10 days. The beer ingredients obtained here are the sixth
Shown in the table.

As shown in Table 6, the beer brewed with 7-C-8 was a new type of beer, which had a high ethyl caproate concentration and was organoleptically different from conventional beers.

Example 6 Mutant yeast 7-C-8 (FERM P-8452) and brewer's yeast Saccha
Whiskey was produced using romyces uvarum (IFO 0565). Water 1 was added to 160 g of malt, saccharified by heating and then filtered to obtain wort. After cooling this, water was added to bring the total amount to 1. 2 g of each yeast was added to this wort 1 in terms of wet cell weight, and fermentation was carried out at 15 ° C. for 10 days. This fermentation liquid was distilled under reduced pressure with a rotary evaporator to obtain whiskey. The whiskey ingredients obtained here are
Shown in the table.

As shown in Table 7, whiskeys made from 7-C-8 are
It has a high concentration of ethyl caproate, and it has been organoleptically different from whiskey until now.

Example 7 Using the mutant yeast 7-C-8 (FERM P-8452) and the Shochu No. 2 yeast of the Brewing Society, shochu was produced with the charge composition shown in Table 8.

2 g of cultured yeast as wet cell weight in the preparation of Table 8
Was added, and 1-stage preparation was performed. After fermentation at 15 ° C. for 14 days, the mash was distilled under reduced pressure on a rotary evaporator. Table 9 shows the components of the obtained shochu.

The shochu using 7-C-8 had a high concentration of ethyl caproate and was a new type of shochu with a sensory sense.

Claims (2)

[Claims] 1. A method for producing an alcoholic beverage or bread, which comprises using a mutant yeast which has been produced by mutation to produce a large amount of caproic acid and / or ethyl caproate among aroma components. 2. A mutant yeast that produces a large amount of caproic acid and / or ethyl caproate among aroma components due to mutation is obtained from a strain grown in a cerulenin-containing medium by subjecting various yeasts to a mutation treatment. Claims No. 1
A method for producing an alcoholic beverage or bread according to the item.