NZ621711B2 - Novel yeast and method for producing ethanol using same - Google Patents
Novel yeast and method for producing ethanol using same Download PDFInfo
- Publication number
- NZ621711B2 NZ621711B2 NZ621711A NZ62171112A NZ621711B2 NZ 621711 B2 NZ621711 B2 NZ 621711B2 NZ 621711 A NZ621711 A NZ 621711A NZ 62171112 A NZ62171112 A NZ 62171112A NZ 621711 B2 NZ621711 B2 NZ 621711B2
- Authority
- NZ
- New Zealand
- Prior art keywords
- xylose
- yeast
- glucose
- ethanol
- raw material
- Prior art date
Links
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 title claims abstract description 188
- 240000004808 Saccharomyces cerevisiae Species 0.000 title claims abstract description 63
- 238000004519 manufacturing process Methods 0.000 title claims abstract description 29
- SRBFZHDQGSBBOR-SQOUGZDYSA-N Xylose Natural products O[C@@H]1CO[C@@H](O)[C@@H](O)[C@@H]1O SRBFZHDQGSBBOR-SQOUGZDYSA-N 0.000 claims abstract description 144
- 229960003487 Xylose Drugs 0.000 claims abstract description 72
- WQZGKKKJIJFFOK-GASJEMHNSA-N D-Glucose Natural products OC[C@H]1OC(O)[C@H](O)[C@@H](O)[C@@H]1O WQZGKKKJIJFFOK-GASJEMHNSA-N 0.000 claims abstract description 63
- 239000008103 glucose Substances 0.000 claims abstract description 63
- WQZGKKKJIJFFOK-VFUOTHLCSA-N β-D-glucose Chemical compound OC[C@H]1O[C@@H](O)[C@H](O)[C@@H](O)[C@@H]1O WQZGKKKJIJFFOK-VFUOTHLCSA-N 0.000 claims abstract description 32
- 239000002994 raw material Substances 0.000 claims description 31
- 239000007788 liquid Substances 0.000 claims description 27
- 239000002028 Biomass Substances 0.000 claims description 20
- 239000000413 hydrolysate Substances 0.000 claims description 6
- 150000002772 monosaccharides Chemical class 0.000 claims description 6
- 241000222120 Candida <Saccharomycetales> Species 0.000 claims description 2
- 229940035295 Ting Drugs 0.000 claims description 2
- 235000014680 Saccharomyces cerevisiae Nutrition 0.000 description 58
- 238000000855 fermentation Methods 0.000 description 19
- 230000004151 fermentation Effects 0.000 description 19
- QTBSBXVTEAMEQO-UHFFFAOYSA-N acetic acid Chemical compound CC(O)=O QTBSBXVTEAMEQO-UHFFFAOYSA-N 0.000 description 18
- IJGRMHOSHXDMSA-UHFFFAOYSA-N nitrogen Chemical compound N#N IJGRMHOSHXDMSA-UHFFFAOYSA-N 0.000 description 16
- 239000000243 solution Substances 0.000 description 14
- 235000000346 sugar Nutrition 0.000 description 14
- 150000008163 sugars Chemical class 0.000 description 10
- 241000191335 [Candida] intermedia Species 0.000 description 8
- 229910052757 nitrogen Inorganic materials 0.000 description 8
- GZCGUPFRVQAUEE-KCDKBNATSA-N D-(+)-Galactose Natural products OC[C@@H](O)[C@H](O)[C@H](O)[C@@H](O)C=O GZCGUPFRVQAUEE-KCDKBNATSA-N 0.000 description 7
- WQZGKKKJIJFFOK-QTVWNMPRSA-N D-mannopyranose Chemical compound OC[C@H]1OC(O)[C@@H](O)[C@@H](O)[C@@H]1O WQZGKKKJIJFFOK-QTVWNMPRSA-N 0.000 description 7
- WQZGKKKJIJFFOK-PHYPRBDBSA-N α-D-galactose Chemical compound OC[C@H]1O[C@H](O)[C@H](O)[C@@H](O)[C@H]1O WQZGKKKJIJFFOK-PHYPRBDBSA-N 0.000 description 7
- OKTJSMMVPCPJKN-UHFFFAOYSA-N carbon Chemical compound [C] OKTJSMMVPCPJKN-UHFFFAOYSA-N 0.000 description 6
- 229910052799 carbon Inorganic materials 0.000 description 6
- 229940041514 Candida albicans extract Drugs 0.000 description 5
- 239000001913 cellulose Substances 0.000 description 5
- 229920002678 cellulose Polymers 0.000 description 5
- 239000004615 ingredient Substances 0.000 description 5
- 244000005700 microbiome Species 0.000 description 5
- 125000002924 primary amino group Chemical group [H]N([H])* 0.000 description 5
- 230000000754 repressing Effects 0.000 description 5
- 239000012138 yeast extract Substances 0.000 description 5
- 229920001817 Agar Polymers 0.000 description 4
- 229920002488 Hemicellulose Polymers 0.000 description 4
- HEBKCHPVOIAQTA-SCDXWVJYSA-N Xylitol Chemical compound OC[C@H](O)[C@@H](O)[C@H](O)CO HEBKCHPVOIAQTA-SCDXWVJYSA-N 0.000 description 4
- 229960002675 Xylitol Drugs 0.000 description 4
- 239000008272 agar Substances 0.000 description 4
- 238000011156 evaluation Methods 0.000 description 4
- 150000002500 ions Chemical class 0.000 description 4
- 239000011259 mixed solution Substances 0.000 description 4
- 230000000284 resting Effects 0.000 description 4
- 239000000811 xylitol Substances 0.000 description 4
- 235000010447 xylitol Nutrition 0.000 description 4
- 239000002253 acid Substances 0.000 description 3
- 239000007864 aqueous solution Substances 0.000 description 3
- 235000013305 food Nutrition 0.000 description 3
- 230000003301 hydrolyzing Effects 0.000 description 3
- 239000000463 material Substances 0.000 description 3
- 238000000034 method Methods 0.000 description 3
- 239000000203 mixture Substances 0.000 description 3
- 239000008363 phosphate buffer Substances 0.000 description 3
- HEMHJVSKTPXQMS-UHFFFAOYSA-M sodium hydroxide Chemical compound [OH-].[Na+] HEMHJVSKTPXQMS-UHFFFAOYSA-M 0.000 description 3
- 241000196324 Embryophyta Species 0.000 description 2
- 230000002401 inhibitory effect Effects 0.000 description 2
- 230000035772 mutation Effects 0.000 description 2
- 230000002269 spontaneous Effects 0.000 description 2
- 239000010902 straw Substances 0.000 description 2
- 239000000126 substance Substances 0.000 description 2
- 230000001629 suppression Effects 0.000 description 2
- XSQUKJJJFZCRTK-UHFFFAOYSA-N urea Chemical compound NC(N)=O XSQUKJJJFZCRTK-UHFFFAOYSA-N 0.000 description 2
- ZPWVASYFFYYZEW-UHFFFAOYSA-L Dipotassium phosphate Chemical compound [K+].[K+].OP([O-])([O-])=O ZPWVASYFFYYZEW-UHFFFAOYSA-L 0.000 description 1
- 229960001031 Glucose Drugs 0.000 description 1
- 239000007836 KH2PO4 Substances 0.000 description 1
- VTHJTEIRLNZDEV-UHFFFAOYSA-L Magnesium hydroxide Chemical compound [OH-].[OH-].[Mg+2] VTHJTEIRLNZDEV-UHFFFAOYSA-L 0.000 description 1
- 241001465754 Metazoa Species 0.000 description 1
- GNSKLFRGEWLPPA-UHFFFAOYSA-M Monopotassium phosphate Chemical compound [K+].OP(O)([O-])=O GNSKLFRGEWLPPA-UHFFFAOYSA-M 0.000 description 1
- 240000007594 Oryza sativa Species 0.000 description 1
- 235000007164 Oryza sativa Nutrition 0.000 description 1
- 241000235647 Pachysolen tannophilus Species 0.000 description 1
- 235000003534 Saccharomyces carlsbergensis Nutrition 0.000 description 1
- 229940081969 Saccharomyces cerevisiae Drugs 0.000 description 1
- 240000000111 Saccharum officinarum Species 0.000 description 1
- 235000007201 Saccharum officinarum Nutrition 0.000 description 1
- 241000235060 Scheffersomyces stipitis Species 0.000 description 1
- HDTRYLNUVZCQOY-LIZSDCNHSA-N Trehalose Chemical compound O[C@@H]1[C@@H](O)[C@H](O)[C@@H](CO)O[C@@H]1O[C@@H]1[C@H](O)[C@@H](O)[C@H](O)[C@@H](CO)O1 HDTRYLNUVZCQOY-LIZSDCNHSA-N 0.000 description 1
- HDTRYLNUVZCQOY-WSWWMNSNSA-N Trehalose Natural products O[C@@H]1[C@@H](O)[C@@H](O)[C@@H](CO)O[C@@H]1O[C@@H]1[C@H](O)[C@@H](O)[C@@H](O)[C@@H](CO)O1 HDTRYLNUVZCQOY-WSWWMNSNSA-N 0.000 description 1
- 240000008529 Triticum aestivum Species 0.000 description 1
- 240000008042 Zea mays Species 0.000 description 1
- 235000002017 Zea mays subsp mays Nutrition 0.000 description 1
- NBIIXXVUZAFLBC-UHFFFAOYSA-K [O-]P([O-])([O-])=O Chemical compound [O-]P([O-])([O-])=O NBIIXXVUZAFLBC-UHFFFAOYSA-K 0.000 description 1
- 238000009825 accumulation Methods 0.000 description 1
- 150000001413 amino acids Chemical class 0.000 description 1
- 230000015572 biosynthetic process Effects 0.000 description 1
- 239000006227 byproduct Substances 0.000 description 1
- 239000004202 carbamide Substances 0.000 description 1
- 238000005119 centrifugation Methods 0.000 description 1
- 235000005822 corn Nutrition 0.000 description 1
- 235000005824 corn Nutrition 0.000 description 1
- 229910000396 dipotassium phosphate Inorganic materials 0.000 description 1
- 235000019797 dipotassium phosphate Nutrition 0.000 description 1
- 238000004821 distillation Methods 0.000 description 1
- 230000000694 effects Effects 0.000 description 1
- 238000005516 engineering process Methods 0.000 description 1
- 238000005755 formation reaction Methods 0.000 description 1
- 150000004676 glycans Polymers 0.000 description 1
- 150000002402 hexoses Chemical class 0.000 description 1
- 238000011031 large scale production Methods 0.000 description 1
- 239000000347 magnesium hydroxide Substances 0.000 description 1
- 229910001862 magnesium hydroxide Inorganic materials 0.000 description 1
- 229910052943 magnesium sulfate Inorganic materials 0.000 description 1
- CSNNHWWHGAXBCP-UHFFFAOYSA-L magnesium sulphate Substances [Mg+2].[O-][S+2]([O-])([O-])[O-] CSNNHWWHGAXBCP-UHFFFAOYSA-L 0.000 description 1
- 235000019341 magnesium sulphate Nutrition 0.000 description 1
- 239000002207 metabolite Substances 0.000 description 1
- 238000002156 mixing Methods 0.000 description 1
- 229910000402 monopotassium phosphate Inorganic materials 0.000 description 1
- 235000019796 monopotassium phosphate Nutrition 0.000 description 1
- 150000002972 pentoses Chemical class 0.000 description 1
- 239000010452 phosphate Substances 0.000 description 1
- 229920001282 polysaccharide Polymers 0.000 description 1
- 239000005017 polysaccharide Substances 0.000 description 1
- 150000004804 polysaccharides Polymers 0.000 description 1
- 239000000047 product Substances 0.000 description 1
- 238000011084 recovery Methods 0.000 description 1
- 230000002829 reduced Effects 0.000 description 1
- 235000009566 rice Nutrition 0.000 description 1
- 238000010792 warming Methods 0.000 description 1
- 239000002699 waste material Substances 0.000 description 1
- 235000021307 wheat Nutrition 0.000 description 1
- 239000002023 wood Substances 0.000 description 1
Classifications
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N1/00—Microorganisms, e.g. protozoa; Compositions thereof; Processes of propagating, maintaining or preserving microorganisms or compositions thereof; Processes of preparing or isolating a composition containing a microorganism; Culture media therefor
- C12N1/14—Fungi; Culture media therefor
- C12N1/16—Yeasts; Culture media therefor
- C12N1/165—Yeast isolates
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12P—FERMENTATION OR ENZYME-USING PROCESSES TO SYNTHESISE A DESIRED CHEMICAL COMPOUND OR COMPOSITION OR TO SEPARATE OPTICAL ISOMERS FROM A RACEMIC MIXTURE
- C12P7/00—Preparation of oxygen-containing organic compounds
- C12P7/02—Preparation of oxygen-containing organic compounds containing a hydroxy group
- C12P7/04—Preparation of oxygen-containing organic compounds containing a hydroxy group acyclic
- C12P7/06—Ethanol, i.e. non-beverage
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12P—FERMENTATION OR ENZYME-USING PROCESSES TO SYNTHESISE A DESIRED CHEMICAL COMPOUND OR COMPOSITION OR TO SEPARATE OPTICAL ISOMERS FROM A RACEMIC MIXTURE
- C12P7/00—Preparation of oxygen-containing organic compounds
- C12P7/02—Preparation of oxygen-containing organic compounds containing a hydroxy group
- C12P7/04—Preparation of oxygen-containing organic compounds containing a hydroxy group acyclic
- C12P7/06—Ethanol, i.e. non-beverage
- C12P7/08—Ethanol, i.e. non-beverage produced as by-product or from waste or cellulosic material substrate
- C12P7/10—Ethanol, i.e. non-beverage produced as by-product or from waste or cellulosic material substrate substrate containing cellulosic material
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12R—INDEXING SCHEME ASSOCIATED WITH SUBCLASSES C12C - C12Q, RELATING TO MICROORGANISMS
- C12R2001/00—Microorganisms ; Processes using microorganisms
- C12R2001/645—Fungi ; Processes using fungi
- C12R2001/72—Candida
-
- Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
- Y02—TECHNOLOGIES OR APPLICATIONS FOR MITIGATION OR ADAPTATION AGAINST CLIMATE CHANGE
- Y02E—REDUCTION OF GREENHOUSE GAS [GHG] EMISSIONS, RELATED TO ENERGY GENERATION, TRANSMISSION OR DISTRIBUTION
- Y02E50/00—Technologies for the production of fuel of non-fossil origin
- Y02E50/10—Biofuels, e.g. bio-diesel
Abstract
Yeast, designated 4-6-4T2 and deposited under the accession number of FERM BP-11509, capable of producing ethanol with high efficiency within a short time from glucose and xylose in the co-presence of glucose and xylose; and a method for producing ethanol using the yeast.
Description
Description
[Title of Invention] Novel Yeast and Method for Producing
l Using Same
[Technical Field]
The present invention relates to a novel yeast and a
method for producing l using the same.
[Background Art]
In recent years, large-scale production of
bioethanol has been conducted throughout the world as a
countermeasure against global warming. A main raw
material for bioethanol is edible biomass such as corn
biomass or sugarcane biomass. Such edible biomass is
matic in terms of a competition between the use as
a raw al for bioethanol and the use as a food
material.
In order to avoid such a problem, it has been
d to develop a technique of producing ethanol from
a cellulosic biomass of wood, non-edible herb and the
like, which are not used as food materials (particularly,
cellulosic biomass obtained from agricultural residues,
forestry residues, etc.).
In such a technique, cellulose or hemicellulose
which is comprised in cellulosic biomass, or a
polysaccharide as a partially decomposed product of such
cellulose or hemicellulose, is hydrolyzed to obtain a
saccharified on comprising, as main ients,
hexose (glucose, mannose and galactose) and pentose
(xylose). Subsequently, sugar(s) comprised in the
rified solution are fermented by microorganisms, so
as to obtain ethanol.
Saccharomyces cerevisiae is known as a yeast capable
of efficiently producing ethanol from glucose and mannose.
However, there are only several types of microorganisms,
which are capable of efficiently producing ethanol from
xylose or galactose.
For instance, Pichia stipitis, Candida ae and
Pachysolen tannophilus (Non Patent Literature 1), and
Candida intermedia (Non Patent Literature 2), are known
as a few examples of microorganisms e of ing
ethanol not only from glucose but also from xylose.
[Citation List]
[Non Patent Literature]
[Non Patent Literature 1]
Yablochkova, EN., Bolotnikova, OI., Mikhailova, NP.,
Nemova, NN. And Ginak, AI. Applied Biochemistry and
Microbiology, Vol. 39, 6 (2003)
[Non Patent Literature 2]
Y. Morikawa, et al., hnology and
Bioengineering, Vol: XXVII, 3 (1984)
[Summary of Invention]
[Problem to be solved by the Invention]
However, the yeast described in Non Patent
Literature 1 has been problematic in that, when ethanol
fermentation is d out in the presence of both
glucose and xylose comprised in a raw material liquid
derived from cellulosic biomass, almost no xylose is
ed until glucose has been almost completely
consumed as a result of catabolite repression caused by
glucose, and thus that a long period of time is required
for fermentation, and ethanol productivity is thereby
lowered.
The present inventor conducted studies regarding the
yeast described in Non Patent Literature 2. As a result,
it was found that catabolite repression is hardly caused
by glucose, depending on production conditions, and that
although there may be a case in which ethanol
fermentation can be carried out from both e and
xylose when it is carried out in the presence of the
glucose and the xylose, the yeast described in Non Patent
Literature 2 is problematic in that xylose consumption
efficiency is poor, and in that the yeast described in
Non Patent Literature 2 does not have a ient
ability to produce l from xylose.
ore, the present invention relates to
provision of a novel yeast having an ability to
efficiently produce ethanol from glucose and xylose in a
short time in the coexistence of the glucose and the
xylose and a method for producing ethanol using the novel
yeast.
[Solution to Problem]
Hence, as a result of intensive studies, the t
inventor found that a specific yeast has an ability to
efficiently produce ethanol from glucose and xylose in a
short time in the coexistence of the glucose and the
xylose, thereby completing the present ion.
Specifically, the present invention provides a yeast,
which was designated as Candida intermedia 44T2 and
was deposited as FERM BP-11509.
In addition, the present invention also es a
method for producing ethanol, which comprises a step of
fermenting a raw material liquid comprising one or more
monosaccharides selected from glucose and xylose using
the aforementioned yeast.
[Effects of Invention]
The yeast of the present invention has an ability to
efficiently produce ethanol from glucose and xylose in a
short time in the coexistence of the e and the
xylose. Moreover, using such a yeast, ethanol can be
efficiently produced in a short time even in the case of
using a raw material d from cellulosic biomass
comprising glucose or xylose.
Therefore, ing to the method for producing
ethanol of the present invention, ethanol can be
efficiently produced in a short time from a cellulosic
biomass-derived raw material.
[Brief Description of Drawings]
[Figure 1] Figure 1 is a view showing a change over time
in individual ients when glucose and xylose are
fermented with a parent strain.
[Figure 2] Figure 2 is a view showing a change over time
in individual ingredients when e and xylose are
fermented with a 44T2 strain.
[Figure 3] Figure 3 is a view showing a change over time
in individual ingredients when xylose is fermented with a
parent strain.
[Figure 4] Figure 4 is a view showing a change over time
in individual ingredients when xylose is fermented with a
44T2 strain.
[Description of Embodiments]
<Yeast>
The yeast of the present invention is a yeast, which
was designated as Candida intermedia 44T2 and was
deposited at the International Patent Organism Depositary
(IPOD), al Institute of Technology and Evaluation
(NITE), Incorporated Administrative Agency (address:
Tsukuba Central 6, Higashi 11, Tsukuba, Ibaraki, Japan,
postal code: 305-8566) under ion No. FERM BP-11509
on September 20, 2012. This yeast was obtained by
ting Candida intermedia 0601" used as a
parent strain to spontaneous mutation according to an
ordinary method and ing a strain having a higher
ethanol production ability than the parent strain. It is
to be noted that the aforementioned "NBRC10601" is a
yeast available from the ational Patent Organism
tary (IPOD), National Institute of Technology and
Evaluation (NITE), Incorporated Administrative Agency.
The yeast of the present invention has an ability to
ently produce ethanol from glucose and xylose in a
short time in the coexistence of the glucose and the
xylose.
Herein, the phrase "in the coexistence of glucose
and xylose" is used to mean that the yeast of the t
invention coexists in a raw material liquid (fermentation
liquor) comprising at least glucose and xylose. As
mentioned above, the conventional yeast has not had
ient xylose consumption efficiency, or it has had
an ability to produce ethanol from either one of glucose
and xylose. In the case of the conventional yeast, when
both glucose and xylose were t, almost no xylose
was consumed, until glucose had been completely consumed
by catabolite repression. In contrast, the yeast of the
present invention has an ability to efficiently e
ethanol from both glucose and xylose in a short time,
even when both the glucose and the xylose are present.
Moreover, while the yeast of the present invention
efficiently produces l from a raw material liquid
comprising glucose and xylose in a short time, it
produces almost no l as a by-product. Furthermore,
the yeast of the present invention has the same
properties as those of its parent strain, other than such
an ability to produce ethanol from sugar(s).
<Method for Producing Ethanol>
The method for producing l of the present
invention is characterized in that it comprises a step of
fermenting a raw material liquid comprising one or more
monosaccharides selected from glucose and xylose, using
the aforementioned yeast (hereinafter also referred to as
a "fermentation step"). According to this production
method, ethanol can be efficiently produced in a short
time.
The amount of the yeast used is not particularly
limited. The yeast is used at a ratio of generally
approximately from 0.01 to 100 mass parts, and preferably
from 0.1 to 10 mass parts, based on 1 mass part of the
mentioned ccharide(s).
The above-mentioned raw material liquid (for example,
a saccharified solution) preferably ses glucose and
xylose. Moreover, in addition to these ccharides,
the raw material liquid may also se other
monosaccharides es) such as mannose or galactose.
The production method of the present invention is hardly
affected by catabolite repression. Hence, according to
the present production method, even if the raw material
liquid comprises the aforementioned monosaccharides
(xylose, mannose, or galactose) as well as glucose,
ethanol can be efficiently produced.
When the above-mentioned raw material liquid
ses glucose and , with regard to the content
ratios of glucose and xylose, xylose is used at a ratio
of preferably 0.1 to 5 mass parts, and more ably
0.5 to 3 mass parts, based on 1 mass part of glucose.
Using the yeast of the present invention, even if the
content ratios of glucose and xylose are within the above
described , ethanol can be produced not only from
glucose but also from xylose. Moreover, each of the
contents of glucose and xylose in the raw material liquid
is preferably 1 to 100 g/L, and more ably 5 to 50
g/L. Using the yeast of the present invention, even if
each of the contents of glucose and xylose is within the
above-described ranges, ethanol can be produced.
Furthermore, the content ratio of glucose with t to
sugar(s) comprised in the raw material liquid is not
particularly limited. Glucose is contained at a mass
percentage of preferably 5% to 99 %, and more preferably
% to 70 %, based on the total mass of the sugar(s)
comprised in the raw material liquid.
The total content of monosaccharides comprised in
the above described raw material liquid is preferably 1
to 100 g/L.
The raw material liquid used in ethanol production
may be any one of a raw material liquid derived from
animals, a raw material liquid derived from plants, and a
raw material liquid derived from rial wastes.
Among these, a raw material liquid derived from plants is
preferable. From the viewpoint of preventing a
ition n the use as a raw material for
bioethanol and the use as a food material, a cellulosic
biomass hydrolysate is more preferable. The term
"cellulosic biomass" is used herein to mean biomass
comprising cellulose and hemicellulose. Glucose is
obtained by hydrolyzing cellulose comprised in such
biomass, whereas glucose, , mannose, and galactose
are obtained by hydrolyzing hemicellulose comprised
therein.
From the viewpoint of economic advantage in ethanol
production, such cellulosic biomass is preferably
obtained from agricultural residues (rice straw, wheat
straw, etc.), forestry residues (lumbers, etc.), and the
like.
The pH (30°C) of the above described raw material
liquid is ably 3.5 to 6.5, more preferably 4 to 6,
r preferably 4.5 to 5.5, and particularly
preferably 4.5 to 5. According to the yeast of the
t invention, ethanol production is possible even in
such a low pH range. Thus, even in the case of using a
saccharified solution comprising acetic acid or the like
(an acid hydrolysate of biomass cellulose), l can
be ed.
It has been reported that a saccharified on
with a low pH value obtained by hydrolyzing cellulosic
biomass using an acid comprises an inhibitory substances
that inhibit ethanol production by microorganisms, such
as acetic acid (Nigam JN. Journal of Applied Microbiology,
Vol. 90, 208-215 (2001)). It has also been reported that,
in particular, ethanol fermentation by microorganisms
capable of producing ethanol from xylose is inhibited in
a low pH range such as pH 5 or less (Palmqvist E., Hahn-
Hagardal B. Bioresource Technology Vol. 74, 25-33 (2000)).
The above bed production method is carried out
at a temperature of preferably 20°C to 35°C, and more
preferably 25°C to 30°C.
Moreover, in the above described production method,
while the above described yeast may be d to grow
under its growing ions, ethanol may be produced.
Alternatively, the yeast may be left in the state of
resting cells, namely, the yeast may be left under
conditions in which a nitrogen source is reduced to an
amount insufficient for the growth of the yeast and a
carbon source used as a raw material for l is
abundant, so that ethanol can be produced under
conditions in which the growth of the yeast is suspended.
Among these conditions, it is preferable to produce
ethanol by the yeast which is in the state of resting
cells, since ethanol production is hardly affected by
ethanol production inhibitory substances, such as acetic
acid or ous acid, under such conditions. When
ethanol production is carried out by the yeast which is
in the state of resting cells, the concentration of the
yeast is preferably 5 to 100 g/L. Moreover, when l
is ed by the yeast which is in the state of resting
cells, it is not necessary to add a nitrogen source, a
yeast extract, etc. to the raw material liquid. It is
preferable that the raw material liquid be adjusted to
have the aforementioned pH value by addition of a
phosphate , sodium hydroxide or the like. The
recovery of ethanol may be carried out according to
ordinary means such as distillation.
Furthermore, in the above described production
method, from the viewpoint of the amount of cells, it is
preferable that a pre-culture be carried out before the
aforementioned fermentation step. As a medium used in
such a pre-culture, a medium comprising glucose and one
or more sugars selected from mannose, galactose and
xylose is preferable. As such a medium, a medium
comprising the entioned cellulosic biomass
hydrolysate may be used.
The tration of the aforementioned sugars in
total is preferably 1 to 100 g/L, and more preferably 10
to 50 g/L. The content ratio of one or more sugars
ed from mannose, galactose and xylose to glucose is
not particularly limited.
When a cellulosic biomass hydrolysate is used as a
carbon source in the above bed pre-culture, the
hydrolysate is used at a volume percentage of generally
% or less, and preferably 10% or less, based on the
volume of the . Other ingredients comprised in the
medium are not particularly limited. Examples of such
other ingredients include: nitrogen sources suitable for
growth, such as amino acid, urea, polypeptone, and an
amino ree nitrogen base; and a yeast extract. The
temperature applied to the pre-culture is preferably 10°C
to 37°C, and more preferably 25°C to 30°C. The pH applied
to the pre-culture is preferably 4 to 7, and more
preferably 4.5 to 6.5. In addition, the pre-culture is
preferably d out under aerobic conditions.
[Examples]
Hereinafter, the present ion will be described
in detail in the following examples. However, these
examples are not intended to limit the scope of the
present invention.
Example 1
In accordance with the following procedures, the
yeast Candida intermedia 0601," which was deposited
at the ational Patent Organism Depositary (IPOD),
National Institute of Technology and Evaluation ,
Incorporated Administrative Agency, was used as a parent
strain, and the yeast Candida intermedia "NBRC10601" was
subjected to acclimation and spontaneous mutation,
thereby obtaining a yeast strain 2.
First, the pH of an acetic acid aqueous solution
comprising glucose and xylose (each 1 mass %) was
adjusted to be 5.0 with magnesium hydroxide, and 20% of
this solution was then mixed with 80% of a liquid medium
(1% yeast extract, 2% amino acid-free nitrogen base).
Thereafter, 1% xylose was added to 10 mL of the obtained
mixed solution, and one um loop of the yeast
Candida intermedia "NBRC10601" was then inoculated into
the mixed solution. The thus obtained mixture was
cultured at 30°C for 3 days to obtain a e solution.
Subsequently, in the same manner as described above,
50% of an acetic acid aqueous solution comprising glucose
and xylose (each 1 mass %) that had been adjusted to have
pH 5.0 was mixed with 50% of a liquid medium. Thereafter,
to 10 mL of this mixed solution, 100 mL of the culture
solution obtained as a result of the aforementioned
culture for 3 days was added, and the thus obtained
mixture was further cultured for 7 days. Thereafter, in
the same manner as described above, 80% of an acetic acid
aqueous solution comprising glucose and xylose (each 1
mass %) that had been adjusted to have pH 5.0 was mixed
with 20% of a medium. To 10 mL of this mixed solution,
100 mL of the culture solution obtained as a result of
the entioned culture for 7 days was added, and the
obtained e was further cultured for 30 days, so as
to prepare an acclimated strain solution.
The prepared acclimated strain solution was diluted
by 1000 times, and the thus diluted solution was applied
onto a YNB agar medium (5% glucose, 1% yeast t, 2%
amino acid-free nitrogen base, 2% agar), and the
resultant was then cultured at 25°C for 4 days.
Thereafter, a strain that formed a colony was obtained.
The obtained strain was applied onto a YNB agar
medium (2% trehalose, 1% yeast extract, 2% amino acidfree
nitrogen base, 2% agar), and the resultant was
cultured 25°C for 3 days. Thereafter, formation of a
colony was confirmed, and the culture was then stored at
4°C. Colonies, which had grown at 4°C, were selected, and
thereafter, using the selected colonies, an ethanol
tion test was carried out in a phosphate buffer
(2.5% xylose, 0.1 M KH2PO 4, pH = 5.0, 0.006 M MgSO4×7H2O).
A strain having an y to e ethanol, which was
higher than the parent strain, was selected.
Thus, a yeast of interest was selected, and was
designated as Candida intermedia 44T2. The present
yeast strain was deposited at the ational Patent
Organism Depositary (IPOD), National Institute of
logy and Evaluation (NITE), orated
Administrative Agency. The accession number is FERM BP-
11509.
Example 2
The parent strain (NBRC10601) and the strain 44T2
were each inoculated into a YNB medium (0.5% glucose, 2%
xylose, 1% yeast extract, 2% amino ree nitrogen
base), and the obtained mixtures were each subjected to a
shaking culture at 30°C at 120 rpm (24 to 48 hours, OD660
= 18 to 25). The weight of a dry yeast strain in 1 mL of
the culture on was calculated according to the
following relational expression (1), which was obtained
empirically. (According to the expression (1), the
weight of a dry yeast strain could be calculated to be
0.0062 g in the case of OD660 = 20, for example.)
Weight of dry yeast strain (g/mL) = 0.00032 ´ OD660 -
0.00017 ... (1)
A necessary amount of culture solution was recovered
by centrifugation (3000 rpm, 2 minutes) such that the
amount of the yeast strain became 2% in terms of dry
weight. Thereafter, a fermentation solution prepared by
mixing the sugars shown in Tables 1 to 4 below (raw
materials for ethanol) into a phosphate buffer
(KH2PO4/K2HPO4: 0.01M, pH = 5.0) was added to each yeast
strain, followed by performing fermentation. A change
over time in the amount of ethanol produced was confirmed.
The results are shown in Tables 1 to 4 and Figures 1 to 4.
It is to be noted that the s G and X used in the
tables and the s indicate glucose and xylose,
respectively.
[Table 1]
Fermentation
Cell mass used: NBRC10601, Sugars: G (2.5%) + X (2.5%)
[Table 2]
Fermentation
Cell mass used: 44T2, Sugars: G (2.5%) + X (2.5%)
[Table 3]
Fermentation
Cell mass used: 601, Sugars: X (5%)
[Table 4]
Fermentation
Cell mass used: 44T2, Sugars: X (5%)
As shown in Table 1 and Figure 1, when ethanol was
produced by the parent strain (2%) using glucose and
xylose (each 2.5%) as carbon sources, both the glucose
and the xylose were consumed at the initial stage of
fermentation and ethanol was produced. However, at the
time at which all glucose was consumed, ethanol
tion was terminated, and also, ption of
xylose was almost ated. From these results, it is
found that the parent strain was hardly affected by
catabolite sion caused by glucose, but that it had
a low ability to produce ethanol from xylose.
In contrast, as shown in Table 2 and Figure 2, when
ethanol was produced by the strain 44T2 (2%) using
glucose and xylose (each 2.5%) as carbon sources, both
the glucose and the xylose were consumed at the initial
stage of fermentation, as in the case of the parent
strain. Not only ethanol was produced from both the
glucose and the xylose, but also consumption of xylose
progressed at a nearly nt rate even after
completion of glucose consumption. As a result, not only
glucose but also xylose was sufficiently consumed, and
together with this phenomenon, the amount of ethanol
produced was also increased. From these s, it is
found that the strain 44T2 inherited the property of
being hardly affected by catabolite repression caused by
glucose from the parent strain, and at the same time, the
strain 44T2 had an y to produce ethanol from
xylose that had been significantly improved when compared
with the parent strain.
In addition, as shown in Table 3 and Figure 3, when
ethanol was produced by the parent strain (2%) using
xylose (5%) as a carbon source, 30% or more of xylose
remained even after 24 hours of fermentation.
In contrast, as shown in Table 4 and Figure 4, when
ethanol was produced by the strain 2 (2%) using
xylose (5%) as a carbon source, 90% or more of xylose was
consumed after 24 hours of fermentation, and ethanol was
produced in an amount approximately 1.4 times higher than
that in the case of using the parent strain.
The reason why there is a difference between the
strain 2 and the parent strain in terms of ability
to produce ethanol from xylose has not been necessarily
clarified. From the above Figures 1 to 4, however, it is
assumed that production of ethanol from xylose would be
suppressed or inhibited by accumulation of xylitol which
is an intermediate metabolite during the production of
ethanol from xylose.
That is to say, from the comparison between Figures
1 and 3 each showing a change over time in fermentation
using the aforementioned parent strain, and Figures 2 and
4 each showing a change over time in tation using
the aforementioned strain 44T2, it is considered that
a large amount of xylitol is produced by the parent
strain, and with such an increase in the amount of
xylitol produced, consumption of xylose is lowered, and
l production tends to be terminated.
Thus, it is d that the reason that ethanol can
be efficiently produced using the yeast of the t
invention would be that the present yeast is hardly
affected by catabolite sion caused by glucose, and
also that production of l is suppressed and thereby
ethanol productivity is hardly inhibited by such xylitol.
Example 3
The yeasts shown in Table 5 (approximately 2% dry
weight) were each added to a fermentation solution (0.01
M phosphate buffer, pH 5.0) each comprising the sugars
shown in the same table. Twelve hours later, the amount
of l produced (% (v/v)) was determined. The
results are shown in Table 5.
[Table 5]
From Table 5, it is found that the strain 44T2
exhibits an ability to e ethanol, which is higher
than that of the parent strain or known yeast strains,
regardless of the types of sugars used.
Claims (5)
- [Claim 1] A yeast, which was designated as Candida edia 44T2 and was deposited as FERM BP-11509.
- [Claim 2] A method for producing ethanol, which comprises a step of ting a raw material liquid comprising at least one monosaccharides selected from glucose and xylose, using the yeast according to claim 1.
- [Claim 3] The method according to claim 2, wherein the raw material liquid comprises glucose and .
- [Claim 4] The method according to claim 2 or 3, wherein the pH of the raw material liquid is 3.5 to 6.5.
- [Claim 5] The method according to any one of claims 2 to 4, wherein a cellulosic biomass hydrolysate is used as the raw material liquid.
Applications Claiming Priority (3)
Application Number | Priority Date | Filing Date | Title |
---|---|---|---|
JP2011240158A JP5715930B2 (en) | 2011-11-01 | 2011-11-01 | Novel yeast and ethanol production method using the same |
JP2011-240158 | 2011-11-01 | ||
PCT/JP2012/077428 WO2013065541A1 (en) | 2011-11-01 | 2012-10-24 | Novel yeast and method for producing ethanol using same |
Publications (2)
Publication Number | Publication Date |
---|---|
NZ621711A NZ621711A (en) | 2015-05-29 |
NZ621711B2 true NZ621711B2 (en) | 2015-09-01 |
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