JP6429513B2 - Effective use of sugar alcohol mixture - Google Patents

Description

 The present invention relates to a method for producing yeast cells, wherein yeast is cultured using maltitol and sorbitol, which are sugar alcohols, as sugar sources (carbon sources). Furthermore, it is related with the manufacturing method of a yeast cell body which culture | cultivates yeast using a sugar alcohol manufacturing process liquid for a culture medium.

 In recent years, interest in oral care such as prevention of obesity and lifestyle-related diseases and prevention of tooth decay and bad breath has increased, and sugar alcohols that have low calorie and non-cariogenic functions have attracted attention as raw materials for various sugarless foods. , Demand is growing. Sugar alcohol is produced by adding hydrogen to various sugars obtained from corn or potato starch as a raw material, or chickenpox. In the production process, a mixture containing a large amount of maltitol and sorbitol is produced. Among these, sorbitol is limited to microorganisms that can assimilate this, such as bacteria belonging to the genus Gluconobactor, which have excellent ability to oxidize various sugars and sugar alcohols. There has been no report that maltitol, which is another main component, can be assimilated by microorganisms.

 On the other hand, in order to obtain yeast cells that are raw materials for seasonings and the like, yeast is often cultured, but glucose is generally used as a sugar to be added to the medium. It accounts for part of the production cost.

Soemphol W. et al .: Characterization of genes involved in D-sorbitol oxidation in thermotolerant Gluconobacter frateurii. Bioscience, Biotechnology, and Biochemistry 76 (8), 1497-505. 2012

 An object of the present invention is to produce yeast cells using maltitol or sorbitol as a sugar source, and to produce yeast cells using a mixed solution containing maltitol or sorbitol as a sugar source.

The present inventor is able to assimilate maltitol and sorbitol, which are main components of sugar alcohol waste liquor, by edible yeast, Candida utilis, that is, culturing yeast cells using them as a sugar source. We have found out that we can do it and have reached the present invention.
That is, the present invention
(1) A method for culturing Candida utilis, characterized in that maltitol or sorbitol is assimilated by Candida utilis.
(2) The present invention provides a cell body of Candida utilis obtained by the culture method of (1) above.

 According to the present invention, maltitol and sorbitol, which have been difficult to assimilate by microorganisms, can be used as a cultured sugar source for Candida utilis. This makes it possible to replace glucose for the medium with a mixed solution containing maltitol and sorbitol produced in the sugar alcohol production process. Furthermore, since there are few microorganisms that can assimilate maltitol and sorbitol, the risk of contamination, which is the most problematic in yeast culture, can be reduced compared to the case of using other sugar sources.

Hereinafter, the present invention will be described in detail.
Maltitol and sorbitol used in the present invention are compounds belonging to sugar alcohols.
In the process of industrially producing sugar alcohol, a mixed solution containing maltitol and sorbitol is produced.
In the present invention, maltitol and sorbitol are assimilated by Candida utilis, which is a kind of yeast.

Known yeasts include the genera Saccharomyces, Schizosaccharomyces, Candida, Yarrowia, Pichia, Hansenula, and Kluyveromyces.
So-called edible yeasts include what are commonly called brewer's yeast (Saccharomyces pastorianus, Saccharomyces cerevisiae), baker's yeast (Saccharomyces cerevisiae), and torula yeast (Candida utilis). Among them, yeast strains commonly used for seasoning yeast extracts include Saccharomyces cerevisiae and Candida utilis.

The present invention has been found that among the various yeast strains described above, the edible yeast Candida utilis has the ability to assimilate maltitol and sorbitol. The Candida utilis strain used in the present invention is not particularly required to be genetically modified or transformed, and may be a commonly used strain.

In the present invention, maltitol and sorbitol can be added to the medium as part or all of the sugar source during the cultivation of Candida utilis.
The sugar alcohol to be added may be maltitol and sorbitol, which are commercially available products, or a mixed solution containing maltitol and sorbitol obtained from the sugar alcohol production process. When a mixed solution containing maltitol and sorbitol is used as a sugar source, the mixing ratio of maltitol and sorbitol may be arbitrary, and the content can be adjusted at the time of medium composition, and thus may be arbitrary.

In addition to these sugar sources, the medium composition used for culture in the present invention only needs to contain a nitrogen source, a sulfur source, a phosphate, and a mineral.

 In the present invention, a strain having an assimilability of sugar alcohol refers to a yeast containing a sugar alcohol-containing medium such as SM medium (0.67 w / v% yeast nitrogen base, 2 w / v% maltitol, pH 6.5). In this case, the growth of the cells was observed when cultured for 3 days.

The present invention will be specifically described below with reference to examples and comparative examples.
The present invention is not limited to these.

<Strain capable of assimilating maltitol>
<Example 1>
Candida utilis NBRC0988 was precultured overnight in 5 ml of YPD medium (yeast extract 1 w / v%, polypeptone 2 w / v%, glucose 2 w / v%), and 100 ml of SD medium (0.67 w / v% yeast). Nitrogen base, 2w / v% glucose, pH adjusted to 6.5) and SM medium (0.67w / v% yeast nitrogen base, 2w / v% maltitol, pH adjusted to 6.5) 0.5v each / v% inoculated and cultured at 30 ° C. and 225 rpm. The sugar source of SD medium is glucose, and the sugar source of SM medium is maltitol. FIG. 1A shows a graph in which the OD ₆₀₀ of the culture broth was measured as an indicator of cell growth and the change with time was plotted.

<Comparative Example 1>
In Example 1, it carried out similarly to Example 1 except having used Saccharomyces cerevisiae NBRC101557 instead of Candida utilis. The result is shown in FIG. 1B.

<Comparative Example 2>
In Example 1, it carried out like Example 1 except having used Pichia pastoris NBRC0948 instead of Candida utilis. The result is shown in FIG. 1C.

<Comparative Example 3>
In Example 1, it carried out like Example 1 except having used Kluyveromyces marxianus NBRC1777 instead of Candida utilis. The result is shown in FIG. 1D.

<Comparative example 4>
In Example 1, Schizosaccharomyces pombe NBRC0340 was used instead of Candida utilis, and YE medium (yeast extract 0.5%, glucose 3%) was used instead of YPD medium for pre-culture, as in Example 1. went. The result is shown in FIG. 1E.

As a result of Example 1 to Comparative Example 4, as can be seen from the graphs of FIGS. 1A to 1E, Candida utilis showed high maltitol utilization ability. In contrast, none of Saccharomyces cerevisiae, Pichia pastoris, Kluyveromyces marxianus, and Schizosaccharomyces pombe could assimilate maltitol.

<Specificity of maltitol utilization>
<Example 2>
Candida utilis NBRC0988 was pre-cultured overnight in 5 ml YPD medium (yeast extract 1%, polypeptone 2%, glucose 2%) in 100 ml SM medium (0.67w / v% yeast nitrogen base, 2w / v % Maltitol, pH adjusted to 6.5), 0.5% (v / v) was inoculated, cultured at 30 ° C. and 225 rpm, OD ₆₀₀ of the culture was measured, and the change with time was observed.

<Comparative Example 5>
In Example 2, a test was conducted in the same manner as in Example 2 using a medium containing glucose, fructose, sucrose, and maltose instead of maltitol in the SM medium.
The OD ₆₀₀ after 48 hours from the start of the culture was measured, and the results are shown in Table 1. Maltitol in Example 2 and glucose, fructose, sucrose, and maltose in Comparative Example 5 grew, but OD ₆₀₀ per gram of sugar was highest in maltitol, and Candida utilis had a high ability to utilize sugar alcohol. The result showed that.

<Various sugar alcohol utilization>
<Example 3>
Candida utilis NBRC0988 pre-cultured overnight in 5 ml YPD medium into 100 ml SM medium (0.67 w / v% yeast nitrogen base, 2 w / v% maltitol, pH adjusted to 6.5) 0.5 v / v% was inoculated, cultured at 30 ° C. and 225 rpm, the OD ₆₀₀ of the culture was measured, and the change with time was observed.

<Comparative Example 6>
In Example 3, the test was performed in the same manner as in Example 3 except that sorbitol and mannitol were used instead of maltitol in the SM medium.
The OD ₆₀₀ after 48 hours from the start of the culture was measured, and the results are shown in Table 2. Although there was a difference in the growth of bacterial cells depending on the type of sugar alcohol, growth was confirmed with any sugar alcohol.

<Difference in growth rate between strains>
<Example 4>
Candida utilis NBRC0987 and Candida utilis NBRC0988 were pre-cultured overnight in 5 ml of YPD medium (yeast extract 1%, polypeptone 2%, glucose 2%) in 100 ml SD medium and SM medium 0.5% (v / V) Inoculated, cultured at 30 ° C. and 225 rpm, OD ₆₀₀ of the culture was measured, and changes with time were observed.
These two strains are standard strains of Candida utilis and can be obtained from NBRC.

FIG. 2A shows the time course of the growth of Candida utilis NBRC0987, and FIG. 2B shows the time course of the growth of Candida utilis NBRC0988. When the sugar source is maltitol, the growth start of NBRC0987 was delayed from that of NBRC0988, but since all strains assimilated maltitol, strains belonging to Candida utilis are mostly maltitol. It was shown that it possessed the assimilation property.

The graph which contrasted the growth in Example 1, Candida utilis in case sugar sources are glucose and maltitol. The graph which contrasted the growth in case the glucose sources are glucose and maltitol in the comparative example 1 and Saccharomyces cerevisiae. The graph which compared the growth in case the glucose source is glucose and maltitol in the comparative example 2 and Pichia pastoris. The graph which contrasted the growth in case the glucose source is glucose and maltitol in the comparative example 3, Kluyveromyces marxianus. The graph which compared the growth in case the glucose sources are glucose and maltitol in the comparative example 4, Schizosaccharomyces pombes. The growth graph which used maltitol as a sugar source in Example 4, Candida utilis NBRC0987. The growth graph which used maltitol as a sugar source in Example 4, Candida utilis NBRC0988.

As described above, according to the present invention, Candida utilis can assimilate maltitol and sorbitol, and therefore can be cultured using a sugar alcohol process liquid containing a large amount thereof as a sugar source.

Claims (1)

A method for culturing Candida utilis, characterized in that maltitol or sorbitol is assimilated by Candida utilis without performing genetic recombination .