• Korean Journal of Microbiology
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• v.45 no.2
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• pp.119-125
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• 2009

To understand the ecological function of heterotrophic bacterial community in water column of large freshwater lakes in the permafrost zone, we investigated the structure and function of bacterial community in Lake Khuvsgul, Mongolia. Species composition of overall bacterial community was analyzed by denaturing gradient gel electrophoresis (DGGE) of 16S rRNA gene fragments, and bacteria that can be cultured at 10oC were isolated and characterized. Based on the depth profile of environmental parameters, thermocline and chemocline were recognized at the 5~10 m zone of the water column. The stratified DGGE profile indicated that the discontinuity of water properties might influence the structure of bacterial community: band profiles in the 0~5 m zone were diverse with large change by depth, but the profile was relatively stable at the $\geq$10 m zone, with predominance of the band identified as Acidovorax facilis. Bacterial cultures were screened for protease, cellulase, amylase and lipase activity, and 23 isolates were selected for high activity of the hydrolytic enzymes. The isolates were identified based on their 16S rRNA gene sequences. In the surface water (zero meter depth), Acidovorax defluvii and Sphingobacterium faecium with high cellulase activity were present. Flavobacterium succinicans, Mycoplana bullata and A. facilis were stably predominant isolates at 2 m, 5 m, and $\geq$10 m depths, respectively. F. succinicans isolates showed high protease activity while M. bullata isolates showed moderate levels of protease and celluase activity. A. facilis isolates showed either cellulase or lipase activity, exclusively to each other. According to the profile of growth rates of the isolates in the temperature range of $0\sim42^{\circ}C$, the surface-zone (0~5 m) isolates were facultative psychrophiles while isolates from $\geq$10 m depth were typical mesophiles. This stratification is believed to be due to stratified availability of organic materials to the bacterial decomposers. In the water column below the chemoline, the environment is extremely oligotrophic so that the trait of rapid growth in low temperature might not be demanded by deep-lake decomposers. The stratified distribution of community composition and decomposer activity in Lake Khuvsgul implies that ecological functions of bacterial community in lakes of cold region are sharply divided by water column stratification.

• Korean Journal of Microbiology
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• v.51 no.2
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• pp.97-107
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• 2015

Wetlands constitute a transitional zone between terrestrial and aquatic ecosystems and have unique characteristics such as frequent inundation, inflow of nutrients from terrestrial ecosystems, presence of plants adapted to grow in water, and soil that is occasionally oxygen deficient due to saturation. These characteristics and the presence of vegetation determine physical and chemical properties that affect decomposition rates of organic matter (OM). Decomposition of OM is associated with activities of various extracellular enzymes (EE) produced by bacteria and fungi. Extracellular enzymes convert macromolecules to simple compounds such as labile organic carbon (C), nitrogen (N), phosphorus (P), and sulfur (S) that can be easily taken up by microbes and plants. Therefore, the enzymatic approach is helpful to understand the decomposition rates of OM and nutrient cycling in wetland soils. This paper reviews the physical and biogeochemical factors that regulate extracellular enzyme activities (EEa) in wetland soils, including those of ${\beta}$-glucosidase, ${\beta}$-N-acetylglucosaminidase, phosphatase, arylsulfatase, and phenol oxidase that decompose organic matter and release C, N, P, and S nutrients for microbial and plant growths. Effects of pH, water table, and particle size of OM on EEa were not significantly different among sites, whereas the influence of temperature on EEa varied depending on microbial acclimation to extreme temperatures. Addition of C, N, or P affected EEa differently depending on the nutrient state, C:N ratio, limiting factors, and types of enzymes of wetland soils. Substrate quality influenced EEa more significantly than did other factors. Also, drainage of wetland and increased temperature due to global climate change can stimulate phenol oxidase activity, and anthropogenic N deposition can enhance the hydrolytic EEa; these effects increase OM decomposition rates and emissions of $CO_2$ and $CH_4$ from wetland systems. The researches on the relationship between microbial structures and EE functions, and environmental factors controlling EEa can be helpful to manipulate wetland ecosystems for treating pollutants and to monitor wetland ecosystem services.

• Korean Journal of Microbiology
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• v.52 no.3
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• pp.278-285
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• 2016

The bifunctional PheA protein, having chorismate mutase and prephenate dehydratase (CMPD) activities, is one of the key regulatory enzymes in the aromatic amino acid biosynthesis in Escherichia coli, and is negatively regulated by an end-product, phenyalanine. Therefore, PheA protein has been thought as useful for protein engineering to utilize mass production of essential amino acid phenylalanine. To obtain feedback resistant PheA protein against phenylalanine, we mutated by using random mutagenesis, extensively screened, and obtained $pheA^{FBR}$ gene encoding a feedback resistant PheA protein. The mutant PheA protein contains substitution of Leu to Phe at the position of 118, displaying that higher affinity (about $290{\mu}M$) for prephenate in comparison with that (about $850{\mu}M$) of wild type PheA protein. Kinetic analysis showed that the saturation curve of $PheA^{FBR}$ against phenyalanine is hyperbolic rather than that of $PheA^{WT}$, which is sigmoidal, indicating that the L118F mutant enzyme has no cooperative effects in prephenate binding in the presence of phenylalanine. In vitro enzymatic assay showed that the mutant protein exhibited increased activity by above 3.5 folds compared to the wild type enzyme. Moreover, L118F mutant protein appeared insensitive to feedback inhibition with keeping 40% of enzymatic activity even in the presence of 10 mM phenylalanine at which the activity of wild type $PheA^{WT}$ was not observed. The substitution of Leu to Phe in CMPD may induce significant conformational change for this enzyme to acquire feedback resistance to end-product of the pathway by modulating kinetic properties.

• Journal of the East Asian Society of Dietary Life
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• v.26 no.5
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• pp.435-444
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• 2016

This study was conducted to investigate the effects of natural preservatives (G3, G3-1, F3, and F3-1) using Cordyceps militaris on improvement of food quality and safety of noodle during storage. Wheat flour noodle were prepared using three different concentrations of natural preservatives (0.100, 0.200, and 0.400%). Changes in microbial populations, pH value, titratable acidity, and sensory evaluation were measured during storage at $12{\pm}2^{\circ}C$ for 3 days. Overall, use of natural preservatives resulted in lower levels of total mesophilic bacteria, coliform, yeast and mold in noodle compared to the control. In particular, natural preservatives using $2{\times}$ MIC concentrations (0.400%) of F3 and F3-1 were effective at maintaining levels of total mesophilic bacteria for noodle during storage. The pH values of noodle made with F3 and F3-1 were higher than the others. The titratable acidity of noodle with natural preservatives did not significantly change during storage. In sensory evaluation, appearance, color, and overall acceptability of noodle with F3 and F3-1 were preferred than the control. These results could provide useful information for developing an alternative preservation method to improve food quality and shelf-life of noodle using natural preservatives.

• Food Science and Preservation
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• v.23 no.5
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• pp.605-613
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• 2016

The quality and shelf life of sliced root of Platycodon grandiflorum (Doraji) treated by ozon-microbubble-heat shock (OMH) were investigated by combining modified-atmosphere packaging [MAP, ($50%O_2+15%CO_2+35%N_2$)]. The study was based on microbiological (total viable bacteria, fungi, Enterobacteriaceae and coliforms numbers), physicochemical and sensory changes. OMH treatment was effective in reducing microbial populations of the sliced Doraji, especially Enterobacteriaceae and coliforms reduced by 2 log CFU/g. However OMH-MAP treatment remained the aerobe and fungi numbers. Regarding the color, OMH-MAP delayed the change of Hunter $b^*$ and the sliced Doraji by OMH-MAP treatment exhibited lower decrease of flavor and overall acceptability compared to those by polypropylene packaging after tap water treatment (Control). The OMH and $50%O_2+15%CO_2$ MAP treatment gave better sensory quality and extended shelf-life for sliced Doraji (~3 days longer shelf-life than Control). Flavor was significantly related to overall acceptability at both Control and OMH-MAP, whereas total coliforms prevalence was associated with overall acceptability at only OMH-MAP. Therefore microbubble-heat shock treatment may improve microbial safety and sliced Doraji by OMH treatment can stored under $50%O_2+15%CO_2$ treatment for up to 7 days. Thus, OMH and MAP treatment may be used in maintaining the storage quality and marketability of sliced Doraji.

• Food Science and Preservation
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• v.10 no.2
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• pp.154-157
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• 2003

To examine the quality changes of three typical fishes under usual storage conditions during marketing, we determined the total bacterial counts and pH values during storage of mackerel, croaker, and saury. Mackerels were stored at 0$^{\circ}C$ and on ice at 19$^{\circ}C$, which is the usual storage condition in a local market and croakers and saury were stored at 0$^{\circ}C$ and 4$^{\circ}C$. Total bacterial counts of mackerel, croaker, and saury were 3,2${\times}$10$^3$, 2.9${\times}$10$^3$, and 2.8 x 10$^4$CFU/g at the time of storage respectively. Total bacterial counts of mackerel stored on ice at 19$^{\circ}C$ increased during storage and reached to 8.4 x 10$\^$6/ CFU/g at day 6, while those stored at 0$^{\circ}C$ decreased up to 2 days of storage and increased to 5.6 ${\times}$ 10$^4$CFU/g. For croaker and saury, total bacterial counts at 0$^{\circ}C$ were 2.5 ${\times}$ 10$\^$5/ and 2.1 x 10$\^$5/ CFU/g at day 6, respectively, while those stored at 4$^{\circ}C$ had 3.6 x 10$\^$6/ and 2.6 ${\times}$ 10$\^$5/ CFU/g. the pH value or mackerel was 5.56 at the time or storage, yet it increased to 6.04. The pH changes of croaker and saury had a similar pattern with that of mackerel, which increased with time of storage. These results suggest that storage of fishes at 0$^{\circ}C$ should be better than those at 4$^{\circ}C$ or on ice at 19$^{\circ}C$ in terms of microbial safety as well as quality and shelf-life of fishes.

• Korean Journal of Microbiology
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• v.50 no.4
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• pp.345-350
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• 2014

The aim of this study was to investigate the effect of high temperature on the viability of probiotic organisms (Bacillus subtilis, Lactobacillus plantarum, and Saccharomyces cerevisiae) mixed with animal feed under controlled conditions by simulating a farm feed bin in the summer. Following inoculation of probiotics into the feed, the pH and probiotic viability were monitored during an 8-day incubation at room temperature. Sterile and non-sterile feeds displayed different patterns of pH changes, with increased pH in non-sterile feed at 2 days, but a pattern of decreasing pH at 4 days. The viabilities of S. cerevisiae and B. subtilis after mono/co-inoculation were maintained without substantial changes during the incubation, whereas L. plantarum viability tended to decline. In both non-sterile and sterile feeds, the probiotics were maintained or grew without any antagonistic effects. Probiotic viability was also tested upon a shift to high temperature ($60^{\circ}C$). There was no distinct change in pH between sterile and non-sterile feeds after the temperature shift. L. plantarum and S. cerevisiae could not survive at the high temperature, whereas B. subtilis displayed normal growth, and it inhibited the growth of contaminant microbes. Fungal growth was not observed in non-sterile feed 2 days after supplementation with B. subtilis. Therefore, heat resistant B. subtilis could be safely used in feed bins to inhibit microbial contamination, even at high temperatures. The prevention of elevated temperature in feed bins is necessary for the utilization of L. plantarum and S. cerevisiae during the summer season.

• Korean Journal of Microbiology
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• v.32 no.2
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• pp.155-162
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• 1994

Cells of Acinetobacter sp. strain JC1 DSM 3803, an aerobic monoxide-oxidizing bacterium, growing on glucose exhibited high catalase activity at the mid-exponential growth phase. The enzyme activity decreased gradually after then until the early stationary phase, increased again at the mid-stationary phase, and then decreased again thereafter. Cells growing on glucose was found to contain three kinds of catalses. Cat1, Cat2 and Cat3. The activities of Cat1 and Cat3 did change significantly during growth, but that of Cat2 exhibited significant variation. Cat3 was found to present only in cells growing on glucose, but not in cells growing on carbon monoxide of methanol. The activities of call and Cat3 in cell-free extracts were stable upon treatment with ethanol and chloroform, but decreased to some extent when the enzymewere treated with 2mM $H_2O_2$ and/or 3-amino-1,2,4-triazole (AT). Cat2 was found to be extremely sensitive to the ethanol-chloroform and $H_2O_2$ treatments, but was insensitive to the AT treatment. Cat1 exhibited enzyme activity after incubation for 1 min at 80$^{\circ}C$. Cat2 and Cat3 did not show enzyme activity after incubation for 1 min at 60$^{\circ}C$ and 70$^{\circ}C$, respectively. Cat2 was found to have peroxidase activity. Cat3 was purified to homogenity in seven steps. The molecular weight of the native enzyme was estimated to be 150,000. Sodium dodecyl sulfate-gel electrophoresis revealed two identical subunits of molecular weight 65,000. The enzyme was found to show two $K_m$ values of 39 mM and 58mM. The optimal pH for the enzyme activity was 7.0, but the activities at pH 6.0, 8.0, and 9.0, were found to be comparable to that at the optimal pH. The optimal temperature for the enzyme activity was found to be 40$^{\circ}C$. The enzyme also exhibited strong activity at 20$^{\circ}C$, 30$^{\circ}C$, and 50$^{\circ}C$. The purified enzyme was not affected by the ethanol-chloroform treatment. The enzyme, howerver, showed less than 10% of the original activity when it was treated with 12 mN AT, 0.1 mM $NaN_3$ of 1mM KCN.

• Korean Journal of Microbiology
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• v.53 no.2
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• pp.83-96
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• 2017

A constructed sea stream in Yeongdo, Busan, Republic of Korea is mostly static due to the lifted stream bed and tidal characters, and receives domestic wastewater nearby, causing a consistent odor production and water quality degradation. Bioaugmentation of a microbial consortium was proposed as an effective and economical restoration technology to restore the polluted stream. The microbial consortium activated on site was augmented on a periodic basis (7~10 days) into the most polluted site (Site 2) which was chosen considering the pollution level and tidal movement. Physicochemical parameters of water qualities were monitored including pH, temperature, DO, ORP, SS, COD, T-N, and T-P. COD and microbial community analyses of the sediments were also performed. A significant reduction in SS, COD, T-N, and COD (sediment) at Site 2 occurred showing their removal rates 51%, 58% and 27% and 35%, respectively, in 13 months while T-P increased by 47%. In most of the test sites, population densities of sulfate reducing bacterial (SRB) groups (Desulfobacteraceae_uc_s, Desulfobacterales_uc_s, Desulfuromonadaceae_uc_s, Desulfuromonas_g1_uc, and Desulfobacter postgatei) and Anaerolinaeles was observed to generally decrease after the bioaugmentation while those of Gamma-proteobacteria (NOR5-6B_s and NOR5-6A_s), Bacteroidales_uc_s, and Flavobacteriales_uc_s appeared to generally increase. Aerobic microbial communities (Flavobacteriaceae_uc_s) were dominant in St. 4 that showed the highest level of DO and least level of COD. These microbial communities could be used as an indicator organism to monitor the restoration process. The alpha diversity indices (OTUs, Chao1, and Shannon) of microbial communities generally decreased after the augmentation. Fast uniFrac analysis of all the samples of different sites and dates showed that there was a similarity in the microbial community structures regardless of samples as the augmentation advanced in comparison with before- and early bioaugmentation event, indicating occurrence of changing of the indigenous microbial community structures. It was concluded that the bioaugmentation could improve the polluted water quality and simultaneously change the microbial community structures via their niche changes. This in situ remediation technology will contribute to an eco-friendly and economically cleaning up of polluted streams of brine water and freshwater.

• Applied Biological Chemistry
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• v.10
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• pp.69-100
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• 1968

1. In order to investigate on the microflora and enzyme activity of mold wheat 'Nuruk' , the major source of microorganisms for the brewing of Takju (a Korean Sake), two samples of Nuruk, one prepared at the College of Agriculture, Chung Nam University (S) and the other perchased at a market (T), were taken for the study. The molds, aerobic bacteria, lactic acid bacteria, and yeasts were examined and counted. The yeasts were classified by the treatment with TTC (2, 3, 5 triphenyltetrazolium chloride) agar that yields a varied shade of color. The amylase and protease activities of Nuruk were measured. The results were as the followings. a) In the Nuruk S found were: Aspergillus oryzae group, $204{\times}10^5$; Black Aspergilli, $163{\times}10^5$; Rhizogus, $20{\times}10^5$; Penicillia, $134{\times}10^5$; Areobic bacteria, $9{\times}10^6-2{\times}10^7$; Lactic acid bacteria, $3{\times}10^4$ In the Nuruk T found were: Aspergillus oryzae group, $836{\times}10^5$; Black Aspergilli, $286{\times}10^5$; Rhizopus, $623{\times}10^5$; Penicillia, $264{\times}10^5$; Aerobic bacteria, $5{\times}10^6-9{\times}10^6$; Lactic acid bacteria, $3{\times}10^4$ b) Eighty to ninety percent of the aerobic bacteria in Nuruk S appeared to belong to Bacillus subtilis while about 70% of those in Nuruk T seemed to be spherical bacteria. In both Nuruks about 80% of lactic acid bacteria were observed as spherical ones. c) The population of yeasts in 1g. of Nuruk S was about $6{\times}10^5$, 56.5% of which were TTC pink yeasts, 16% of which were TTC red pink yeasts, 8% of which were TTC red yeasts, 19.5% of which were TTC white yeasts. In Nuruk T(1g) the number of yeasts accounted for $14{\times}10^4$ and constituted of 42% TTC pink. 21% TTC red pink 28% TTC red and 9% TTC white. d) The enzyme activity of 1g Nuruk S was: Liquefying type Amylase, $D^{40}/_{30},=256$ W.V. Saccharifying type Amylase, 43.32 A.U. Acid protease, 181 C.F.U. Alkaline protease, 240C.F.U. The enzyme activity of 1g Nuruk T was: Liquefying type Amylase $D^{40}/_{30},=32$ W.V. Saccharifying type amylase $^{30}34.92$ A.U. Acid protease, 138 C.F.U. Alkaline protease 31 C.F.U. 2. During the fermentation of 'Takju' employing the Nuruks S and T the microflora and enzyme activity throughout the brewing were observed in 12 hour intervals. TTC pink and red yeasts considered to be the major yeasts were isolated and cultured. The strains ($1{\times}10^6/ml$) were added to the mashes S and T in which pH was adjusted to 4.2 and the change of microflora was examined during the fermentation. The results were: a) The molds disappeared from each sample plot since 2 to 3 days after mashing while the population of aerobic bacteria was found to be $10{\times}10^7-35{\times}10^7/ml$ inS plots and $8.2{\times}10^7-12{\times}10^7$ in plots. Among them the coccus propagated substantially until some 30 hours elasped in the S and T plots treated with lactic acid but decreased abruptly thereafter. In the plots of SP. SR. TP. and TR the coccus had not appeared from the beginning while the bacillus showed up and down changes in number and diminished by 1/5-1/10 the original at the end stage. b) The lactic acid bacteria observed in the S plot were about $7.4{\times}10^7$ in number per ml of the mash in 24 hours and increased up to around $2{\times}10^8$ until 3-4 days since. After this period the population decreased rapidly and reached about $4{\times}10^5$ at the end, In the plot T the lactic acid becteria found were about $3{\times}10^8$ at the period of 24 fours, about $3{\times}10$ in 3 days and about $2{\times}10^5$ at the end in number. In the plots SP. SR. TP, and TR the lactic acid bacteria observed were as less as $4{\times}10^5$ at the stage of 24 hours and after this period the organisms either remained unchanged in population or ceased to exist. c) The maiority of lactic acid bacteria found in each mash were spherical and the change in number displayed a tendency in accordance with the amount of lactic acid and alcohol produced in the mash. d) The yeasts had showed a marked propagation since the period of 24 hours when the number was about $2{\times}10^8$ ㎖ mash in the plot S. $4{\times}10^8$ in 48 hours and $5-7{\times}10^8$ in the end period were observed. In the plot T the number was $4{\times}10^8$ in 24 hours and thereafter changed up and down maintaining $2-5{\times}10^8$ in the range. e) Over 90% of the yeasts found in the mashes of S and T plots were TTC pink type while both TTC red pink and TTC red types held range of $2{\times}10-3{\times}10^7$ throughout the entire fermentation. f) The population of TTC pink yeasts in the plot SP was as $5{\times}10^8$ much as that is, twice of that of S plot at the period of 24 hours. The predominance in number continued until the middle and later stages but the order of number became about the same at the end. g) Total number of the yeasts observed in the plot SR showed little difference from that of the plot SP. The TTC red yeasts added appeared considerably in the early stage but days after the change in number was about the same as that of the plot S. In the plot TR the population of TTC red yeasts was predominant over the T plot in the early stage which there was no difference between two plots there after. For this reason even in the plot w hers TTC red yeasts were added TTC pink yeasts were predominant. TTC red yeasts observed in the present experiment showed continuing growth until the later stage but the rate was low. h) In the plot TP TTC pink yeasts were found to be about $5{\times}10^8$ in number at the period of 2 days and inclined to decrease thereafter. Compared with the plot T the number of TTC pink yeasts in the plot TP was predominant until the middle stage but became at the later stage. i) The productivity of alcohol in the mash was measured. The plot where TTC pink yeasts were added showed somewhat better yield in the earely stage but at and after the middle stage the difference between the yeast-added and the intact mashes was not recognizable. And the production of alcohol was not proportional to the total number of yeasts present. j) Activity of the liquefying amylase was the highest until 12 hours after mashing, somewhat lowered once after that, and again increased around 36-48 hours after mashing. Then the activity had decreased continuously. Activity of saccharifying amylase also decreased at the period of 24 hours and then increased until 48 hours when it reached the maximum. Since, the activity had gradually decreased until 72 hours and rapidly so did thereafter. k) Activity of alkaline protease during the fermentation of mash showed a tendency to decrease continusously although somewhat irregular. Activity of acid protease increased until hours at the maximum, then decreased rapidly, and again increased, the vigor of acid protease showed better shape than that of alkaline protease throughout. 3. TTC pink yeasts that were predominant in number, two strains of TTC red pink yeasts that appeared throughout the brewing, and TTC red yeasts were identified and the physiological characters examined. The results were as described below. a) TTC pinkyeasts (B-50P) and two strains of TTC red pink yeasts (B-54 RP & B-60 RP) w ere identified as the type of Saccharomyces cerevisiae and TTC pink red yeasts CB-53 R) were as the type of Hansenula subpelliculosa. b) The fermentability of four strains above mentioned were measured as follows. Two strains of TTC red pink yeasts were the highest, TTC pink yeasts were the lowest in the fermantability. The former three strains were active in the early stage of fermentation and found to be suitable for manufacturing 'Takju' TTC red yeasts were found to play an important role in Takju brewing due to its strong ability to produce esters although its fermentability was low. c) The tolerance against nitrous acid of strains of yeast was marked. That against lactic acid was only 3% in Koji extract, and TTC red yeasts showed somewhat stronger resistance. The tolerance against alcohol of TTC pink and red pink yeasts in the Hayduck solution was 7% while that in the malt extract was 13%. However, that of TTC red yeasts was much weaker than others. Liguefying activity of gelatin by those four strains of yeast was not recognized even in 40 days. 4. Fermentability during Takju brewing was shown in the first two days as much as 70-80% of total fermentation and around 90% of fermentation proceeded in 3-4 days. The main fermentation appeared to be completed during :his period. Productivity of alcohol during Takju brewing was found to be apporximately 65% of the total amount of starch put in mashing. 5. The reason that Saccharomyces coreanuss found be Saito in the mash of Takju was not detected in the present experiment is considered due to the facts that Aspergillus oryzae has been inoculated in the mold wheat (Nuruk) since around 1930 and also that Koji has been used in Takju brewing, consequently causing they complete change in microflora in the Takju brewing. This consideration will be supported by the fact that the original flavor and taste have now been remarkably changed.