Metabolite Profiling during Fermentation of Makgeolli by the Wild Yeast Strain Saccharomyces cerevisiae Y98-5

Abstract

Makgeolli is a traditional Korean alcoholic beverage. The flavor of makgeolli is primarily determined by metabolic products such as free sugars, amino acids, organic acids, and aromatic compounds, which are produced during the fermentation of raw materials by molds and yeasts present in nuruk, a Korean fermentation starter. In this study, makgeolli was brewed using the wild yeast strain Saccharomyces cerevisiae Y98-5, and temporal changes in the metabolites during fermentation were analyzed by ultra-high-performance liquid chromatography-quadrupole-time-of-flight mass spectrometry. The resultant data were analyzed by partial least squares-discriminant analysis (PLS-DA). Various metabolites, including amino acids, organic acids, sugar alcohols, small peptides, and nucleosides, were obviously altered by increasing the fermentation period. Changes in these metabolites allowed us to distinguish among makgeolli samples with different fermentation periods (1, 2, 3, 6, 7, and 8 days) on a PLS-DA score plot. In the makgeolli brewed in this study, the amounts of tyrosine (463.13 µg/mL) and leucine (362.77 µg/mL) were high. Therefore, our results indicate that monitoring the changes in metabolites during makgeolli fermentation might be important for brewing makgeolli with good nutritional quality.

Full text

353
Mycobiology
Metabolite Profiling during Fermentation of
Makgeolli by the Wild Yeast Strain Saccharomyces
cerevisiae Y98-5
Hye Ryun Kim *, Jae-Ho Kim , Byung Hak Ahn and Dong-Hoon Bai
Traditional Alcoholic Beverage Research Team, Korea Food Research Institute, Seongnam 463-746, Korea
Department of Food Engineering, Dankook University, Cheonan 330-714, Korea
Abstract Makgeolli is a traditional Korean alcoholic beverage. The flavor of makgeolli is primarily determined by metabolic
products such as free sugars, amino acids, organic acids, and aromatic compounds, which are produced during the fermentation
of raw materials by molds and yeasts present in nuruk, a Korean fermentation starter. In this study, makgeolli was brewed using
the wild yeast strain Saccharomyces cerevisiae Y98-5, and temporal changes in the metabolites during fermentation were analyzed
by ultra-high-performance liquid chromatography-quadrupole-time-of-flight mass spectrometry. The resultant data were analyzed
by partial least squares-discriminant analysis (PLS-DA). Various metabolites, including amino acids, organic acids, sugar alcohols,
small peptides, and nucleosides, were obviously altered by increasing the fermentation period. Changes in these metabolites
allowed us to distinguish among makgeolli samples with different fermentation periods (1, 2, 3, 6, 7, and 8 days) on a PLS-DA
score plot. In the makgeolli brewed in this study, the amounts of tyrosine (463.13 μg/mL) and leucine (362.77 μg/mL) were high.
Therefore, our results indicate that monitoring the changes in metabolites during makgeolli fermentation might be important for
brewing makgeolli with good nutritional quality.
Keywords Fermentation, Makgeolli, Metabolite, Saccharomyces cerevisiae
Makgeolli is a traditional Korean alcoholic beverage. It is
brewed from rice and nuruk (a Korean fermentation starter)
and roughly filtered before serving. Makgeolli is mainly
consumed by the general public [1]. In the case of makgeolli,
the entire fermented material is homogenized and consumed
as it stands, unlike alcoholic beverages that are more finely
filtered (e.g., cheongju or yakju). Thus, makgeolli includes
the vitamin B group, essential amino acids, glutathione, as
well as proteins, oligosaccharides, and live yeast. Accordingly,
it has nutritional characteristics that are different from
those of other alcoholic beverages [2]. With the recent
increase in the consumption of makgeolli, studies on the
functional effects and flavor components of makgeolli have
also increased. For example, it has been reported that
makgeolli has anticancer effects [3, 4], effects on blood
circulation and lipids [5], antihypertensive activity [6, 7],
fibrinolytic and superoxide dismutase-like activity [8], and
antibacterial/antioxidant activity [9]. Studies on the volatile
flavor components of takju (a type of makgeolli) have shown
that they depend on the type of yeast [10] and the raw
material [11]. In addition, many studies have focused on
the strains used for makgeolli fermentation. These studies
include those conducted for the selection of koji (Aspergillus
spp.) and yeast for the improvement of fermentation
characteristics and cheongju quality [12]; isolation and
identification of a yeast strain that produces abundant
glutathione (a biologically active substance) and determination
of the optimal production conditions [13]; screening of
brewing yeasts and saccharifying molds for foxtail millet
wine-making and examination of the brewing characteristics
of the selected stains [14, 15]; determination of changes in
microflora during fermentation of takju and yakju [16];
isolation and identification of yeast strains with high
viability that produce a high concentration of ethanol [17];
isolation and characterization of ethanol-tolerant yeast
[18]; and finally, research on the production of biologically
active substances such as an antihypertensive angiotensin-
Research Article
Mycobiology 2014 December, 42(4): 353-360
http://dx.doi.org/10.5941/MYCO.2014.42.4.353
pISSN 1229-8093 • eISSN 2092-9323
© The Korean Society of Mycology
*Corresponding author
E-mail: hrkim@kfri.re.kr
Received July 28, 2014
Revised August 18, 2014
Accepted September 22, 2014
This is an Open Access article distributed under the terms of the
Creative Commons Attribution Non-Commercial License (http://
creativecommons.org/licenses/by-nc/3.0/) which permits unrestricted
non-commercial use, distribution, and reproduction in any medium,
provided the original work is properly cited.

354 Kim et al.
converting-enzyme inhibitor [19] and an antidementia β-
secretase inhibitor [20] from Saccharomyces cerevisiae.
However, there has been no analysis of the metabolite
profile during the fermentation of makgeolli.
Therefore, the aim of this study was to analyze changes
in the metabolite profile during fermentation of makgeolli
brewed with koji and yeast isolated from traditional Korean
nuruk.
MATERIALS AND METHODS
Strains and chemicals. Ye a st s is ol a te d f r om nuruk were
used in this study. Saccharomyces cerevisiae Y98-5 was
collected from the Gongju area of Chungnam province
[21]. Koji (saccharogenic power [sp] 85) was purchased
from Seoul Jangsoo, Inc. (Jincheon, Korea). The amino acids
standard and organic acids were obtained from Sigma-
Aldrich Co. (St. Louis, MO, USA). All reagents used for ultra-
high-performance liquid chromatography-quadrupole-time-
of-flight mass spectrometry (UHPLC-Q-TOF MS) analyses
were of high-performance liquid chromatography grade.
Makgeolli brewing. The first brewing (yeast, 0.02% and
koji [sp 85] : distilled water = 38 : 62) was performed to
reach 36% of the total makgeolli volume and was followed
by fermentation at 25
o
C for 2 days. The second brewing
(steamed non-glutinous rice : water = 32 : 68; 64% of the
total makgeolli volume) was then performed, followed by
fermentation at 25
o
C for 8 days. After compression, makgeolli
was prepared by filtration through a 120-mesh filter. Makgeolli
brewing was performed in triplicate.
Chemical analysis. The concentration of soluble solids
was measured with a handheld refractometer (ATAGO
Pocket PAL-1; ATAGO Co. Ltd., Tokyo, Japan) and recorded
in Brix units (% sucrose). The pH was measured with a
model D-51 pH meter (HORIBA, Kyoto, Japan).
Metabolite extraction. To extract metabolites for UHPLC-
Q-TOF MS analysis, 0.9 mL of 50% MeOH (internal standard
reserpine, 10 ppm) was added to 0.1 mL of makgeolli; after
vortexing for 5 min, the mixture was kept at 4
o
C for 16 hr.
Next, centrifugation was performed at 14,000 rpm at 4
o
C
for 20 min; the supernatant was then collected and the
metabolites were extracted.
Metabolomic analysis. For analysis of the metabolome,
we used an Agilent (Santa Clara, CA, USA) UHPLC-Q-
TOF MS system (UHPLC, Agilent 1290 Infinity; MS, Agilent
6520 with Jet Stream Technology) controlled by MassHunter
Workstation Data Acquisition software v. B. 05.00 (Agilent).
Using the ESI + Jet Stream method, in the positive ionization
mode, the gas temperature was set at 325
o
C, the drying gas
(N
2
) flow at 8 L/mL, the nebulizer pressure at 30 psi, the
capillary voltage at 4,000 V, the skimmer voltage at 65 V,
and the fragmentor voltage at 70 V. In the negative ionization
mode, the gas temperature was set at 325
o
C, the drying gas
flow at 8L/mL, the nebulizer pressure at 30 psi, the capillary
voltage at 3,500 V, the skimmer voltage at 65 V, and the
fragmentor voltage at 50 V. For the mobile phase of UHPLC,
a gradient of 5 mM ammonium acetate in water (A) and
0.1% formic acid in acetonitrile (B) was used. Using a
ZORBAX HILIC Plus (2.1 × 100 mm, 3.5 μm; Agilent)
column, the analysis was performed at a flow rate of 0.3 mL/
min and a column temperature of 30
o
C. The data were
aligned and normalized using Mass Profiler Professional
(Agilent), and multivariate statistical analysis was performed
using SIMCA-P+ 12.0.1 (Umetrics, Umea, Sweden).
RESULTS AND DISCUSSION
Changes in chemical properties during fermentation.
Makgeolli was brewed using S. cerevisiae Y98-5 (isolated
from nuruk) and koji as fermenting agents. Koji consists of
non-glutinous rice inoculated with Aspergillus species. Fig.
1 shows the changes in the soluble solids content and pH
during fermentation of the makgeolli. The pH was 3.08 on
the first day of fermentation and then gradually increased,
reaching 3.7 upon the completion of fermentation. The pH
was similar to that of makgeolli brewed using koji made
from different rice varieties, as reported in Kwon et al.
[22]. Furthermore, it was similar to the pH of nuruk mash
prepared using Aspergillus oryzae and Aspergillus kawachii,
as reported in Han et al. [23].
The soluble solids content was 4.3% during the early stage
of fermentation. It then increased, reaching a maximum
value (10.6%) on the seventh day of fermentation, before
decreasing to 10.0% upon the completion of fermentation.
The soluble solids content reflects the amount of sugar
remaining after two processes: amylolysis of rice starch by
the koji mold at the early stage of fermentation, and use of
the resulting sugar as a carbon source by S. cerevisiae Y98-
5 for propagation and alcohol fermentation (final ethanol
content was 15%). In the case of the makgeolli brewed with
Fig. 1. Changes of soluble solids content (◆) and pH (○)
during fermentation of makgeolli brewed with Saccharomyces
cerevisiae Y98-5. Each data point represents the mean ± SD
(n = 3).

Metabolite Profiling of Makgeolli during Fermentation 355
S. cerevisiae Y98-5 and koji as fermenting agents, abnormal
fermentation did not occur. The pH was 3.7 and the soluble
solids content was 10% upon the completion of fermentation.
During the making of makgeolli by dilution with water
after the completion of fermentation, a pH and soluble
solids content suitable for drinking were maintained.
Fig. 2. Tot a l ion c h r om a t ogr a m s of makgeolli brewed with Saccharomyces cerevisiae Y98-5, an amino acids standard, and koji.
IS, internal standard; A, adenine; G, guanine.
Fig. 3. Electron ionization mass spectra of [M + H]
+
ions of adenine, guanine, hypoxanthine, xanthine, arabitol, and erythritol
at a collision energy of 70 eV.

356 Kim et al.
Metabolomic profiling of makgeolli during fermentation.
The metabolome of the S. cerevisiae Y98-5 makgeolli
during fermentation was analyzed using UHPLC-Q-TOF
MS, and 296 metabolites were detected. Most metabolites
had a mass value less than 800. Fig. 2 shows the total ion
chromatogram (TIC) for the metabolome on the eighth
day of fermentation as well as TICs for the koji and amino
acids standard. The TIC for the makgeolli on the eighth day
of fermentation was broadly divided into peaks between 1
and 2.5 min, a peak at 2.9 min (internal standard), peaks
between 3.5 and 6.5 min, a peak at 7.2 min, and a peak at
9.87 min. The peaks between 1 and 2.5 min were identified
as adenine, guanine, hypoxanthine, and xanthine, which
originated from the yeast cells inoculated during the
brewing of makgeolli and the fungus in the koji, and as
arabitol and erythritol, which are sugar alcohols (Fig. 3).
These peaks increased in the makgeolli TIC compared with
the koji TIC. The peaks between 3.5 and 6.5 min were
identified as dipeptides, such as Ser-Val and Glu-Val, and
tripeptides, such as Phe-Arg-Asn and Val-Arg-Val (Fig. 4).
The pattern of peaks between 4.2 and 8.2 min was similar
to the peak pattern of the amino acids standard. The results
indicated the presence of 16 amino acids and the nonprotein
amino acid γ-amino-n-butyric acid (GABA). The peak
observed for the makgeolli at 9.87 min was due to fermentation
and was attributed to an [M + H]
+
ion at m/z 257.1027.
A partial least squares-discriminant analysis (PLS-DA)
of the metabolome of Y98-5 makgeolli during the fermentation
period was performed using SIMCA-P+. As shown in Fig. 5,
the makgeolli samples taken at different fermentation times
were clearly distinguishable in the score plot generated by
combining PC1 (30.15% of the total variance) with PC2
(18.40% of the total variance). Based on PC1, the first-,
second-, and third-day fermentation samples were positioned
on the right side of the plot and the sixth-, seventh-, and
eighth-day fermentation samples were positioned on the left
side, indicating that the early and late stages of fermentation
were distinct. Based on PC2, the first-day fermentation
sample was positioned on the lower side of the pot and the
second- and third-day fermentation samples were positioned
on the upper side, indicating that there were differences
between days even within the early stage of fermentation.
The products of mixed-acid fermentation include mostly
ethanol, acetic acid, lactic acid, succinic acid, and formic
acid. If neutral fermentation occurs, 2,3-butanediol is
produced from pyruvate through acetoin. 2,3-Butanediol is
mostly produced by bacteria such as Bacillus and Enterobacter
[24]. In this experiment, 2,3-butanediol was not detected.
Quantitative analyses of makgeolli metabolites during
fermentation. Table 1 summarizes the major metabolites
that were identified during fermentation of makgeolli by
using UHPLC-Q-TOF MS in the positive and negative ion
modes. Sixteen amino acids (including phenylalanine), the
nonprotein amino acid GABA, and four organic acids
(including citric acid) were identified. The quantitative
analysis of the identified materials indicated that the contents
tended to increase as the fermentation period increased
(Table 2). It has been reported that amino acids are
produced by the enzymatic action of microorganisms
during fermentation of the protein contained in rice, the
major raw material in makgeolli production, and that these
Fig. 4. Electron ionization mass spectra of [M + H]
+
, [M + Na]
+
, and [M + K]
+
ions o f Ser-Va l , Gl u -Val, P h e- Ar g- As n , and Va l -
Arg-Val.

Metabolite Profiling of Makgeolli during Fermentation 357
Fig. 5. Partial least squares-discriminant analysis score plot derived from ultra-high-performance liquid chromatography-
quadrupole-time-of-flight mass spectrometry profiles of makgeolli brewed with Saccharomyces cerevisiae Y98-5 during the
fermentation period (■, 1; ●, 2; ◆, 3; □, 6; ○, 7; and ▲, 8 days). PC1 and PC2 account for 30.15% and 18.40% of the
variance, respectively.
Table 1. Identification of major metabolites of Y98-5 makgeolli by UHPLC-Q-TOF MS in the positive and negative ion modes
No. RT Identity Formular [M + H]
+
Exact mass Actual mass Mass error (ppm) MS fragment (ESI)
0104.323 Phenylalanine C9H12NO2 166.0863 166.0867 2.4080120.08, 131.05
0204.418 Tyrosine C9H12NO3 182.0812 182.0825 7.1390136.07, 165.05
0304.549 Leucine C6H14NO2 132.1019 132.10200.757086.09
0404.648 Isoleucine C6H14NO2 132.1019 132.1017 −1.5140 86.09
0504.692 Methionine C5H12NO2S 150.0583 150.0583 0.0000 104.05, 132.10
0605.039 γ-Amino-n-butyric acid C4H10NO2 104.0706 104.0710 3.844086.06, 87.04
0705.109 Valine C5H12NO2 118.0863 118.0859 −3.387072.08
0805.303 Gluatamic acid C5H10NO4 148.0604 148.06104.052084.04, 102.05, 130.05
0905.415 Threonine C4H10NO3 120.0655 120.0655 0.0000 74.06, 102.05
10 5.55 Aspartic acid C4H8NO4 134.0448 134.0448 0.0000 88.03, 116.03
11 05.572 Serine C3H8NO3 106.0499 106.0496 −2.829060.04, 88.04
12 05.714 Alanine C3H8NO2 90.055 090.0544 −6.663044.049
13 05.955 Glycine C2H6NO2 076.0393 076.0379 −18.412048.05, 59.06
14 06.047 Proline C5H10NO2 116.0706 116.0703 −2.585070.06
15 08.457 Arginine C6H13N2O4S2 175.1190175.1199 5.1390156.07
16 08.872 Histidine C6H10N3O2 156.0768 156.0775 4.4850110.07
17 09.099 Lysine C6H15N2O2 147.1128 147.1131 2.0390121.05, 130.08
Formular [M −H]
18 07.324 Malic acid C4H5O5 133.0142 133.0139 −2.255075.0, 87.0, 114.9
19 07.638 Lactic acid C3H5O3 089.0244 089.0248 4.493044.99, 87.00
20 09.730 Citric acid C6H7O7 191.0197 191.0188 −4.712068.99, 112.98
21 11.359 Succinic acid C4H5O4 117.0193 117.0192 −0.855068.99, 112.98
UHPLC-Q-TOF MS, ultra-high-performance liquid chromatography-quadrupole-time-of-flight mass spectrometry; ESI, electrospray ionization.

358 Kim et al.
constituents affect the taste of makgeolli [25]. It is known
that makgeolli must contain free amino acids that produce
a balance of sour, savory, sweet, and bitter tastes, and that
higher amino acid contents are better [26]. At the early
stage of Y98-5 makgeolli fermentation, the major amino
acids were leucine, glutamic acid, tyrosine, and phenylalanine.
Upon the completion of fermentation, the contents of alanine,
proline, and glycine had increased, and these amino acids
were identified as major amino acids of the makgeolli,
along with the major amino acids identified at the early
stage of fermentation. Although the major amino acids
identified upon the completion of Y98-5 makgeolli fermentation
included tyrosine and leucine, these amino acids were not
among the major amino acids identified in yakju by Lee
[27] (arginine, alanine, glutamic acid, serine, and glycine)
or by Cheong et al. [28] (alanine, proline, phenylalanine,
and glutamic acid). Tyrosine is the starting material for the
production of neurotransmitters, including dopamine, and
thus has been proposed as a treatment for depression. The
high amount of leucine observed at the early stage of
fermentation may be due to differences in the raw rice,
koji, and yeast used in this study compared with previous
studies. Leucine can be used to produce glucose when
there is no intake of food and thus helps to maintain and
regulate blood glucose levels. Among the amino acids,
leucine had the highest content of 463.13 μg/mL on the
eighth day of Y98-5 makgeolli fermentation, and the tyrosine
content was 362.77 μg/mL, the second highest.
Glutamic acid aids sugar and fat metabolism and produces
an umami taste [29]. The concentration of glutamic acid in
the Y98-5 makgeolli on the eighth day of fermentation was
309.3 μg/mL. The concentration of alanine, which plays an
important role in metabolism, aids liver detoxication, and
produces a sweet taste, was 193.39μg/mL. The concentration
of phenylalanine was 232.83 μg/mL; this amino acid is
effective for weight loss and lipid profile improvement
because it elicits the secretion of cholecystokinin in the
intestine, which reduces hunger by stimulating the satiety
center of the brain. Valine, a 2-amino valeric acid, can be
obtained from the hydrolysis of albumin and is synthesized
through the amination of 2-oxoisovaleric acid by valine
aminotransferase. It is a precursor of pantothenic acid and
has properties similar to those of leucine. Thus, isolation of
pure valine from protein hydrolysates is relatively difficult.
Valine protects liver function and has a slightly bitter taste
in addition to a sweet taste [30]. On the eighth day of
fermentation, the concentration of valine was 106.67 μg/mL.
The concentration of methionine, an amino acid that plays
an important role as a precursor of S-adenosylmethionine
and promotes fat metabolism, was 90.85 μg/mL. A low
amount of GABA, a primary inhibitory neurotransmitter
in the brain [31], was observed during makgeolli fermentation.
The highest amount, 15.45 μg/mL, was observed on the
seventh day of fermentation.
Organic acids are major constituents that contribute to
the taste of fermented alcoholic beverages such as sake and
wine [32, 33]. The major organic acid of Y98-5 makgeolli
was citric acid. The maximum amount of citric acid, which
has a fresh sour taste, was 5.088 mg/mL on the seventh
day of fermentation. Lactic, succinic, and malic acids
Table 2 . Quantitative analysis of major Y98-5 makgeolli metabolites during fermentation, using UHPLC-Q-TOF MS
Makgeolli metabolite (μg/mL)
1 day 2 days 3 days 6 days 7 days 8 days
Phenylalanine 112.04 ± 0.36 167.64 ± 2.750203.95 ± 1.56 289.63 ± 7.070310.44 ± 7.750323.83 ± 4.860
Tyrosine 120.93 ± 1.52 195.01 ± 4.080243.14 ± 1.69 327.75 ± 11.00 344.31 ± 6.750362.77 ± 7.480
Leucine 182.98 ± 3.00 184.03 ± 4.760229.01 ± 3.35 392.45 ± 12.85 431.21 ± 4.210463.13 ± 9.460
Isoleucine 049.73 ± 2.74 49.63 ± 1.81 055.10 ± 3.38 76.96 ± 1.95 81.89 ± 1.73 86.47 ± 1.48
Methionine 038.45 ± 0.34 40.63 ± 0.45 043.13 ± 0.42 75.55 ± 2.06 83.82 ± 1.70 90.85 ± 1.89
γ-Amino-n-butyric acid 012.36 ± 0.37 13.59 ± 0.88 011.41 ± 0.79 14.77 ± 0.73 15.45 ± 1.38 014.9 ± 0.27
Val i n e 045.43 ± 0.18 53.85 ± 0.20 060.72 ± 0.53 92.74 ± 1.28 103.69 ± 2.450106.67 ± 1.770
Gluatamic acid 152.33 ± 3.46 184.80 ± 6.570229.50 ± 3.89 282.43 ± 9.250298.79 ± 5.940309.30 ± 1.180
Threonine 040.02 ± 0.26 43.00 ± 0.78 044.48 ± 0.30 62.71 ± 1.50 66.70 ± 1.09 69.66 ± 1.99
Aspartic acid 058.62 ± 3.03 55.54 ± 1.23 067.56 ± 1.13 96.76 ± 4.22 105.43 ± 1.320113.45 ± 3.930
Serine 045.01 ± 0.98 51.27 ± 6.97 054.69 ± 2.78 92.00 ± 3.28 092.03 ± 13.30 104.19 ± 4.080
Alanine 094.59 ± 1.04 107.16 ± 3.070134.81 ± 2.15 118.32 ± 1.310196.72 ± 2.030193.39 ± 2.010
Glycine 072.22 ± 0.08 115.76 ± 0.680122.01 ± 2.44 150.04 ± 5.380152.08 ± 2.480157.1 ± 1.37
Proline 067.82 ± 1.33 140.63 ± 3.890179.22 ± 1.63 227.70 ± 4.700242.98 ± 7.260257.62 ± 4.530
Arginine 004.59 ± 0.16 06.10 ± 0.05 05.83 ± 0.77 09.76 ± 1.63
Histidine 090.58 ± 9.63 106.37 ± 10.44 097.67 ± 3.18 119.78 ± 3.720126.96 ± 1.710129.80 ± 11.41
Lysine 030.13 ± 7.77 51.89 ± 3.53 060.47 ± 5.01 99.86 ± 7.09 127.04 ± 2.840128.79 ± 5.460
Malic acid 049.4 ± 1.2 93.3 ± 8.6 232.3 ± 6.0 494.8 ± 27.1 514.1 ± 24.1 529.06 ± 21.10
Lactic acid 074.4 ± 4.5 268.5 ± 6.300535.9 ± 11.7 717.5 ± 45.4 722.7 ± 59.6 730.8 ± 41.1
Citric acid .04298 ± 12.2 0.5046 ± 120.5 0.5203 ± 40.3 5159 ± 76.1 .5088 ± 54.8 .5070 ± 66.3
Succinic acid 116.1 ± 4.3 360.4 ± 13.7 515.4 ± 6.6 628.9 ± 40.3 648.4 ± 32.1 679.3 ± 17.8
UHPLC-Q-TOF MS, ultra-high-performance liquid chromatography-quadrupole-time-of-flight mass spectrometry.

Metabolite Profiling of Makgeolli during Fermentation 359
(529~730 μg/mL) were the next most abundant organic
acids, in that order; the concentrations of all three tended
to increase until the eighth day of fermentation.
In conclusion, in makgeolli brewed with S. cerevisiae
Y98-5 (isolated from traditional Korean nuruk) and koji, the
amino acid, GABA, and organic acid contents increased
during the fermentation period, and the citric acid content
reached a maximum on the seventh day of fermentation.
The amounts of tyrosine, which is involved in stimulating
and invigorating the brain, and leucine, which functions in
blood sugar regulation, were high. For the metabolomic
study of traditional alcoholic beverages, more metabolite
libraries are needed. Although wine yeasts and baker’s
yeasts are currently imported from foreign countries and
used for the brewing of makgeolli, the above results
demonstrate the nutritional superiority of a domestic yeast
isolated from traditional Korean nuruk. The results of this
study could form the basis for the invigoration of domestic
yeast.
ACKNOWLEDGEMENTS
This study was supported by a grant from the Korean
Traditional Food Globalization Research and Development
Projects (E01444500-01) of the Korea Food Research
Institute.
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