• Title/Summary/Keyword: Biological $H_2$ production

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UV-induced Photodamage - attenuating Properties of Water Extract from Lentinuls edodes (피부각질형성세포에서 표고버섯 물 추출물의 피부노화 억제 효과)

  • Lee, Jung Im;Oh, Jung Hwan;Park, So Young;Kim, Hye Ran;Jung, Kyung Im;Jeon, Byung-Jin;Kim, Dongmin;Kong, Chang-Suk
    • Journal of Life Science
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    • v.30 no.10
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    • pp.877-885
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    • 2020
  • Lentinuls edodes has been used for traditional food and medicine around Asia, and a variety of biological effects have been reported. In this study, L. edodes water extract (LWE) was investigated for its anti-photodamage effect in HaCaT keratinocytes. To perform the necessary assays, L. edodes was extracted with distilled water for 8 hr at 40℃ in an extract tank. Anti-photodamage activity was assessed using a scratch wound healing assay, cell proliferation, and a reactive oxygen species (ROS) scavenging test and by measuring the mRNA and protein expression levels of matrix metalloproteinases (MMPs) and type I procollagen. MMPs and collagen expression are major markers of UV-induced photodamage in skin. Prior to photodamage analysis, the total polyphenol and β-glucan contents of the LWE were evaluated and found to be 4.64 mg GAE/g DW and 165.96 mg/g, respectively. Treatment with LWE induced cell migration and cell proliferation in UV-irradiated HaCaT cells, and LWE effectively scavenged the ROS induced by H2O2 and UVB irradiation in HaCaT cells. UVB irradiation induced ROS generation and led to increased production of MMP-1 and MMP-9 and to decreased collagen production in human keratinocytes. Treatment with LWE upregulated the expression levels of MMP-1, MMP-9, and type I procollagen in UVB-irradiated HaCaT cells. This study suggests that LWE could be used to develop cosmetic materials with anti-photodamage effects.

Solid Fermentation of Medicinal Herb Using Phellinus baumii Mycelium and Anti-thrombin and Anti-oxidation Activity of its Methanol Extract (장수상황버섯 균사체를 이용한 한약재의 고체발효 및 메탄올 추출물의 트롬빈 저해 활성과 항산화 활성)

  • Shin, Yong-Kyu;Jang, Han-Su;Kim, Jong-Sik;Ryu, Hee-Young;Kim, Jong-Kuk;Kwun, In-Sook;Sohn, Ho-Yong
    • Microbiology and Biotechnology Letters
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    • v.36 no.3
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    • pp.201-208
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    • 2008
  • To produce bioactivity-strengthen medicinal herbs, the 36 medicinal herbs which have antioxidation or blood circulation activity, were solid fermented using Phellinus baumii mycelium. Most of medicinal herbs, except Chrysanthemum indicum (flower), Zizyphus jujuba Miller (fructus), Aconitum koreanum R. Raymond (root), Magnolia denu-data (flower), and Polygonatum sibiricum Redt (root bark), showed good fermentation at $25^{\circ}C$ for 20 days under 90% of relative humidity. The poor fermentations of the herbs could be explained by lack of nutrient, structural rigidity, and the content of antifungal substance. After fermentation, the average water content of herbs were increased to $67.21{\pm}11.43%$ from $30.84{\pm}15.67%$, but the average pH and average methanol extraction ratio were slightly decreased to $11.16{\pm}7.06%$ and $4.83{\pm}0.73$ from $13.91{\pm}12.22%$ and $5.06{\pm}0.87$, respectively. The analysis of thrombin inhibition and DPPH scavenging activity of the methanol extracts of herbs showed that thrombin inhibition activities of the fermented Drynaria fortunei Kunze, Melia azedarach var. japonica, Prunus persica and Orostachys japonicus, and DPPH scavenging activities of the fermented Polygala tenuifolia, Scrophularia buergeriana, Angelica dahurica, Drynariafortunei Kunze, Cyperus rotundus, and Boschniakia rossica were increased as compared with those activities of non-fermented its cognate herbs. Our results suggest that the production of bioactivity-strengthen medicinal herbs is possible by solid fermentation of Phellinus baumii mycelium, as fermented Drynaria fortunei Kunze showed increased antioxidant and thrombin inhibitory activities than those of non-fermented herbs.

Degradation of Poultry Feathers by Bacillus amyloliquefaciens Y10 With Plant Growth-promoting Activity and Biological Activity of Feather Hydrolyzates (식물 성장 촉진 활성을 가진 Bacillus amyloliquefaciens Y10에 의한 가금 우모의 분해 및 생산된 우모 분해산물의 생리활성)

  • Yedam Kim;Young Seok Lee;Youngsuk Kim;Jinmyeong Song;Yeongbeen Bak;Gyulim Park;O-Mi Lee;Hong-Joo Son
    • Journal of Life Science
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    • v.34 no.5
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    • pp.304-312
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    • 2024
  • This study was conducted to characterize strain Y10, isolated from discarded chicken feathers. Strain Y10 was identified as Bacillus amyloliquefaciens through phenotypic and 16S rRNA gene analysis. B. amyloliquefaciens Y10 exhibited plant growth-promoting activities, including the production of fungal cell-degrading enzymes (cellulase, lipase, protease, and pectinase), siderophores, ammonia, and indoleacetic acid. Furthermore, strain Y10 was able to inhibit the mycelial growth of several phytopathogenic fungi. When 0.1% sucrose as a carbon source and 0.05% casein as a nitrogen source were added to the basal medium, adjusted to pH 10, and cultured at 35℃, the degradation rate of chicken feathers by strain Y10 was about two times higher than that of the basal medium, with the feathers almost completely degraded in four days. Strain Y10 also degraded various keratin substrates, including duck feathers, wool, and human nails. It was confirmed that the feather hydrolyzates prepared using strain Y10 exhibited antioxidant activities, such as 1,1-diphenyl-2-picrylhydrazyl radical scavenging activity (EC50 = 0.38 mg/ml) and superoxide dismutase-like activity (EC50 = 183.7 mg/ml). These results suggest that B. amyloliquefaciens Y10 is a potential candidate for the development of bioinoculants and feed additives applicable to the agricultural and livestock industries, as well as the microbiological treatment of keratin waste.

Antioxidant and anti-inflammatory effects of red garlic compositions (홍마늘 조성물의 항산화 및 항염증 효과)

  • Kang, Min Jung;Kim, Dong-Gyu;Shin, Jung Hye
    • Food Science and Preservation
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    • v.24 no.3
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    • pp.446-454
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    • 2017
  • Garlic (Allium sativum L.) and traditional herb has several functional properties and strong biological activities, making it useful as a functional food material. We investigated the antioxidant and anti-inflammatory activity of mixed compounds from red garlic and supplementary materials, including ginger (Zingiber officinale Roscoe), doraji (Platycodon grandiflorum), quince (Chaenomeles sinensis), citrus peel (Citri Pericarpium), and mint (Mentha arvensis). The extracts were prepared with water (W) and ethanol (E) at $70^{\circ}C$ (W-70, E-70) and $95^{\circ}C$ (W-95, E-95) for 3 h. The total content of phenolic compounds was the highest in E-70 (608.60 mg/100 g). Alliin, one of the active ingredients in red garlic, was contained at 1.18-1.29 mg/g and 0.81-0.85 mg/g in water and ethanol extract, respectively. Another active ingredient of red garlic, S-allyl-cysteine (SAC) had higher content in the water extract than in the ethanol extracts. DPPH radical scavenging activity was higher in E-70 (15.96-73.65%) at $313-5,000{\mu}g/mL$. ABTS radical scavenging activity was also higher in E-70 (5.71-77.19%) than in the others. The ROS production rate showed the same tendency as the NO production, with more efficacy in E-95. The expression level of iNOS and $IL-1{\beta}$ was decreased in the E-95 significantly at the concentration of $1,000{\mu}g/mL$ compared to the lipopolysaccharide (LPS) treated group. Based on the above results, the antioxidative and anti-inflammatory activities of the extracts of red garlic and supplementary materials were expressed by different useful substances. The contents of these useful substances were different according to the extraction solvent and temperature.

Limno-Biological Investigation of Lake Ok-Jeong (옥정호의 육수생물학적 연구)

  • SONG Hyung-Ho
    • Korean Journal of Fisheries and Aquatic Sciences
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    • v.15 no.1
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    • pp.1-25
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    • 1982
  • Limnological study on the physico-chemical properties and biological characteristics of the Lake Ok-Jeong was made from May 1980 to August 1981. For the planktonic organisms in the lake, species composition, seasonal change and diurnal vertical distribution based on the monthly plankton samples were investigated in conjunction with the physico-chemical properties of the body of water in the lake. Analysis of temperature revealed that there were three distinctive periods in terms of vertical mixing of the water column. During the winter season (November-March) the vertical column was completely mixed, and no temperature gradient was observed. In February temperature of the whole column from the surface to the bottom was $3.5^{\circ}C$, which was the minimum value. With seasonal warming in spring, surface water forms thermoclines at the depth of 0-10 m from April to June. In summer (July-October) the surface mixing layer was deepened to form a strong thermocline at the depth of 15-25 m. At this time surface water reached up to $28.2^{\circ}C$ in August, accompanied by a significant increase in the temperature of bottom layer. Maximum bottom temperature was $r5^{\circ}C$ which occurred in September, thus showing that this lake keeps a significant turbulence Aehgh the hypolimnial layer. As autumn cooling proceeded summer stratification was destroyed from the end of October resulting in vertical mixing. In surface layer seasonal changes of pH were within the range from 6.8 in January to 9.0 in guutuost. Thighest value observed in August was mainly due to the photosynthetic activity of the phytoplankton. In the surface layer DO was always saturated throughout the year. Particularly in winter (January-April) the surface water was oversaturated (Max. 15.2 ppm in March). Vertical variation of DO was not remarkable, and bottom water was fairly well oxygenated. Transparency was closely related to the phytoplankton bloom. The highest value (4.6 m) was recorded in February when the primary production was low. During summer transparency decreased hand the lowest value (0.9 m) was recorded in August. It is mainly due to the dense blooming of gnabaena spiroides var. crassa in the surface layer. A. The amount of inorganic matters (Ca, Mg, Fe) reveals that Lake Ok-Jeong is classified as a soft-water lake. The amount of Cl, $NO_3-N$ and COD in 1981 was slightly higher than those in 1980. Heavy metals (Zn, Cu, Pb, Cd and Hg) were not detectable throughout the study period. During the study period 107 species of planktonic organisms representing 72 genera were identified. They include 12 species of Cyanophyta, 19 species of Bacillariophyta, 23 species of Chlorophyta, 14 species of Protozoa, 29 species of Rotifera, 4 species of Cladocera and 6 species of Copepoda. Bimodal blooming of phytoplankton was observed. A large blooming ($1,504\times10^3\;cells/l$ in October) was observed from July to October; a small blooming was present ($236\times10^3\;cells/l$ in February) from January to April. The dominant phytoplankton species include Melosira granulata, Anabaena spiroides, Asterionella gracillima and Microcystis aeruginota, which were classified into three seasonal groups : summer group, winter group and the whole year group. The sumner group includes Melosira granulate and Anabaena spiroides ; the winter group includes Asterionella gracillima and Synedra acus, S. ulna: the whole year group includes Microtystis aeruginosa and Ankistrodesmus falcatus. It is noted that M. granulate tends to aggregate in the bottom layer from January to August. The dominant zooplankters were Thermocpclops taihokuensis, Difflugia corona, Bosmina longirostris, Bosminopsis deitersi, Keratelle quadrata and Asplanchna priodonta. A single peak of zooplankton growth was observed and maximum zooplankton occurrence was present in July. Diurnal vertical migration was revealed by Microcystis aeruginosa, M. incerta, Anabaena spiroides, Melosira granulata, and Bosmina longirostris. Of these, M. granulata descends to the bottom and forms aggregation after sunset. B. longirostris shows fairly typical nocturnal migration. They ascends to the surface after sunset and disperse in the whole water column during night. Foully one species of fish representing 31 genera were collected. Of these 13 species including Pseudoperilnmpus uyekii and Coreoleuciscus splendidus were indigenous species of Korean inland waters. The indicator species of water quality determination include Microcystis aeruginosa, Melosira granulata, Asterionelta gracillima, Brachionus calyciflorus, Filinia longiseta, Conochiloides natans, Asplanchna priodonta, Difflugia corona, Eudorina elegans, Ceratium hirundinella, Bosmina longirostris, Bosminopsis deitersi, Heliodiaptomus kikuchii and Thermocyclops taihokuensis. These species have been known the indicator groups which are commonly found in the eutrophic lakes. Based on these planktonic indicators Lake Ok-Jeong can be classified into an eutrophic lake.

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Studies on the physio-chemical properties and the cultivation of oyster mushroom(Pleurotus ostreatus) (느타리버섯의 생리화학적성질(生理化學的性質) 및 재배(栽培)에 관(關)한 연구(硏究))

  • Hong, Jai-Sik
    • Applied Biological Chemistry
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    • v.21 no.3
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    • pp.150-184
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    • 1978
  • Nutritional characteristics and physio-chemical properties of mycelial growth and fruitbody formation of oyster mushroom(Pleurotus ostreatus)in synthetic media, the curtural condition for the commerical production in the rice straw and poplar sawdust media, and the changes of the chemical components of the media and mushroom during the cultivation were investigated. The results can be summarized as follows: 1. Among the carbon sources mannitol and sucrose gave rapid mycelial growth and rapid formation of fruit-body with higher yield, while lactose and rhamnose gave no mycelial growth. Also, citric acid, succinic acid, ethyl alcohol and glycerol gave poor fruit-body formation, and acetic acid, formic acid, fumaric acid, n-butyl alcohol, n-propyl alcohol and iso-butyl alcohol inhibited mycelial growth. 2. Among the nitrogen sources peptone gave rapid mycelial growth and rapid formation of fruit-body with higher yield, while D,L-alanine, asparatic acid, glycine and serine gave very poor fruit-body formation, and nitrite nitrogens, L-tryptophan and L-tyrosine inhibited mycelial growth. Inorganic nitrogens and amino acids added to peptone were effective for fruit-body growth, and thus addition of ammonium sulfate, ammonium tartarate, D,L-alanine and L-leucine resulted in about 10% increase fruit-body yield. L-asparic acid about 15%, L-arginine about 20%, L-glutamic acid, and L-lysine about 25%. 3. At C/N ratio of 15.23 fruit-body formation was fast, but the yield decreased, and at C/N ratio of 11.42 fruit-body formation was slow, but the yield increased. Also, at the same C/N ratio the higher the concentration of mannitol and petone, the higher yield was produced. Thus, from the view point of both yield of fruit-body and time required for fruiting the optimum C/N ratio would be 30. 46. 4. Thiamine, potassium dihydrogen phosphate and magnecium sulfate at the concentration of $50{\mu}g%$. 0.2% and 0.02-0.03%, respectively, gave excellent mycelial and fruit-body growth. Among the micronutrients ferrous sulfate, zinc sulfate and manganese sulfate showed synergetic growth promoting effect but lack of manganese resulted in a little reduction in mycelial and fruit-body growth. The optimum concentrati on of each these nutrients was 0.02mg%. 5. Cytosine and indole acetic acid at 0.2-1mg% and 0.01mg%, respectively, increased amount of mycelia, but had no effect on yield of fruit-body. The other purine and pyrimidine bases and plant hormones also had no effect on mycelial and fruit-belly yield. 6. Illumination inhibited mycelial growth, but illumination during the latter part of vegetative growth induced primordia formation. The optimum light intensity and exposure time was 100 to 500 lux and 6-12 hours per day, respectively. Higher intensity of light was injurous, and in darkness only vegetative growth without primordia formation was continued. 7. The optimum temperature for mycelial growth was $25^{\circ}C$ and for fruit-body formation 10 to $15^{\circi}C$. The optimum pH range was from 5.0 to 6.5. The most excellent fry it-body formation were produced from the mycelium grown for 7 to 10 days. The lesser the volume of media, the more rapid the formation of fruit-body; and the lower the yield of fruit-body; and the more the volume of media, the slower the formation of fruit-body, and the higher the yield of fruit-body. The primordia formation was inhibited by $CO_2$. 8. The optimum moisture content for mycelial growth was over 70% in the bottle media of rice straw and poplar sawdust. 10% addition of rice bran to the media exhibited excellent mycelial growth and fruit-body formation, and the addition of calciumcarbonate alone was effective, but the addition of calcium carbonate was ineffective in the presence of rice bran. 9. In the cultivation experiments the total yield of mushroom from the rice straw media was $14.99kg/m^2$, and from the sawdust media $6.52kg/m^2$, 90% of which was produced from the first and second cropping period. The total yield from the rice straw media was about 2.3 times as high as that from the sawdust media. 10. Among the chemical components of the media little change was observed in the content of ash on the dry weight basis, and organic matter content decreased as the cultivation progressed. Moisture content, which was about 79% at the time of spawning, decreased a little during the period of mycelial propagation, after which no change was observed. 11. During the period from spawning to the fourth cropping about 16.7% of the dry matter, about 19.3% of organic matter, and about 40% of nitrogen were lost from the rice straw media; about 7.5% of dry mallet, about 7.6% of organic matter, and about 20% of nitrogen were lost from the sawdust media. For the production of 1kg of mushroom about 232g of organic matter and about 7.0g of nitrogen were consumed from the rice straw media; about 235g of organic matter and about 6.8g of nitrogen were consumed from the sawdust media, 1㎏ of mushroom from either of media contains 82.4 and 82.3g of organic matter and 5.6 and 5.4g of nitrogen, respectively. 12. Total nitrogen content of the two media decreased gradually as the cultivation progressed, and total loss of insoluble nitrogen was greater than that of soluble nitrogen. Content of amino nitrogen continued to increase up to the third cropping time, after which it decreased. 13. In the rice straw media 28.0 and 13.8% of the total pentosan and ${\alpha}$-cellulose, respectively, lost during the whole cultivation period was lost during the period of mycelial growth; in the sawdust media 24.1 and 11.9% of the total pentosan and ${\alpha}$-cellulose, respectively, was lost during the period of mycelial growth. Lignin content in the media began to decrease slightly from the second cropping time, while the content of reduced sugar, trehalose and mannitol continued to increase. C/N ratio of the rice straw media decreased from 33.2 at spawining to 30.0 at ending; that of the sawdust media decreased from 61.3 to 60.0. 14. In both media phosphorus, potassium, manganese and zinc decreased, at magnesium, calcium and copper showed irregular changes, and iron had a tendency to be increased. 15. Enzyme activities are much higher in the rice straw media than in the sawdust media. CMC saccharifying and liquefying activity gradually increased from after mycelial propagation to the second cropping, after which it decreased in both media. Xylanase activity rapidly and greatly increased during the second cropping period rather than the first period. At the start of the third cropping period the activity decreased rapidly in the rice straw media, which was not observed in the sawdust media. Protease activity was highest after mycelial propagation, after which it gradually decreased. The pH of the rice straw media decreased from 6.3 at spawning to 5.0 after fourth cropping; that of the sawdust media decreased from 5.7 to 4.9. 16. The contents of all the components except crude fibre of the mushroom from the rice straw media were higher than those from the sawdust media. Little change was observed in the content of the components of mushroom cropped from the first to the third period, but slight decrease was noticed at the fourth cropping.

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Microbiological and Enzymological Studies on Takju Brewing (탁주(濁酒) 양조(釀造)에 관(關)한 미생물학적(微生物學的) 및 효소학적(酵素學的) 연구(硏究))

  • Kim, Chan-Jo
    • 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.

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